Groups of five mice were intranasally infected with IAV ( 1918 ) , and recombinant DDX3 protein was administered intranasally . Two days post - infection , the expression level of IFNβ in the lungs was analyzed by semi - quantitative RT - PCR ( left panel ) and qPCR ( right panel ) . A p value of < 0 . 05 was considered significant ( * * p < 0 . 01 ) .

O O O B-ORGANISM O O O O B-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM O O O B-GENEPROD O O O O O O O O O O O O B-EXP_ASSAY O O B-GENEPROD O O B-TISSUE O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O O O B-EXP_ASSAY O O O O O O O O O O O O O O O O O O O O O O O O O O

Groups of five mice were intranasally infected with IAV (1918), and recombinant DDX3 protein was administered intranasally. Two days post-infection, the expression level of IFNβ in the lungs was analyzed by semi-quantitative RT-PCR (left panel) and qPCR (right panel). A p value of <0.05 was considered significant (** p < 0.01).


( B ) Left , orthogonal view of a 20μm tumor section labelled with antibodies against EpCAM ( in red ) and DNA in blue . Right , dot plot bar representing cell heights in cells from tumor sections , n = 60 cells from 1 tumor .

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(B) Left, orthogonal view of a 20μm tumor section labelled with antibodies against EpCAM (in red) and DNA in blue. Right, dot plot bar representing cell heights in cells from tumor sections, n=60 cells from 1 tumor.


( C ) On the left , images of low magnification views of healthy tissues labelled with antibodies against pericentrin ( PCNT ) and CDK5RAP2 , shown in red and green , respectively . DNA is in blue . The white dashed squares represent the regions shown in higher magnifications on the right . One centrosome was considered as such when PCNT and CDK5RAP2 signals co - localized ( Inset 1 ) . Lack of co - localization ( Insets 2 and 3 ) was noticed and discarded during quantification .

O O O O O O O O O O O O O O O O O O O B-GENEPROD O B-GENEPROD O O B-GENEPROD O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-SUBCELLULAR O O O O O B-GENEPROD O B-GENEPROD O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O O O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O O O O

(C) On the left, images of low magnification views of healthy tissues labelled with antibodies against pericentrin (PCNT) and CDK5RAP2, shown in red and green, respectively. DNA is in blue. The white dashed squares represent the regions shown in higher magnifications on the right. One centrosome was considered as such when PCNT and CDK5RAP2 signals co-localized (Inset 1). Lack of co-localization (Insets 2 and 3) was noticed and discarded during quantification.


( D ) Super - resolution microscopy of healthy tissues labelled for the centriole marker CEP135 ( in green ) and PCNT ( in red ) .

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(D) Super-resolution microscopy of healthy tissues labelled for the centriole marker CEP135 (in green) and PCNT (in red).


( E - F ) Representative images of two different fields of the same tumor ( E ) and of a tumor section to highlight different centrososome phenotypes , labelled as described in ( C ) . The pink dashed line highlights an area of cells without centrosomes . The white dashed squares represent the regions shown in higher magnifications in insets in ( G and I ) and the pink dashed linesurrounds nuclei without centrosomes . ( G ) Insets from ( F ) . ( H ) Examples of tumor sections showing small ( left ) and large ( right ) regions with nuclei without centrosomes .

O O O O O O O O O O O O O O O O O O O O O O O O O O B-SUBCELLULAR O O O O O O O O O O O O O O O O O O O O B-SUBCELLULAR O O O O O O O O O O O O O O O O O O O O O O O O O B-SUBCELLULAR O B-SUBCELLULAR O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-SUBCELLULAR O B-SUBCELLULAR O

(E-F) Representative images of two different fields of the same tumor (E) and of a tumor section to highlight different centrososome phenotypes, labelled as described in (C). The pink dashed line highlights an area of cells without centrosomes. The white dashed squares represent the regions shown in higher magnifications in insets in (G and I) and the pink dashed linesurrounds nuclei without centrosomes. (G) Insets from (F). (H) Examples of tumor sections showing small (left) and large (right) regions with nuclei without centrosomes.


( I ) Insets from ( F ) showing different types of centrosome amplification events .

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(I) Insets from (F) showing different types of centrosome amplification events.


( J ) Super - resolution microscopy of tumor tissues labelled as in ( C ) .

O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O

(J) Super-resolution microscopy of tumor tissues labelled as in (C).


( K ) Maximum Z - projection of a tumor section labelled with EpCAM antibodies ( Red ) and DNA in blue . The dashed line shows the absence of EpCAM staining , arrows show a disorganized region .

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(K) Maximum Z-projection of a tumor section labelled with EpCAM antibodies (Red) and DNA in blue. The dashed line shows the absence of EpCAM staining, arrows show a disorganized region.


( A ) Images of EOC tumors labelled with antibodies against PCNT ( red ) and CDKRAP2 ( green ) . Nuclei are shown in blue . The colored arrows reflect different centrosome number . The list of centrosome behaviors annotated it is not exhaustive to allow comprehension of the events . # TT72 , shows a cyst - like arrangement of EOC nuclei where it is impossible to correspond centrosomes to individual nuclei . The white bars with arrows of different colors highlight possible interpretations of the same condition . In # TT79 , regions without centrosomes can be identified . In the inset , a single centrosome localized in the lumen can be identified and it is not possible to know to which nucleus ( white dashed lines ) it corresponds to . # TT88 presents only very small clusters of centrosomes , few nuclei without centrosomes and many conditions with 2 centrosomes . In # TT61 , regions without centrosomes can be distinguished , in addition to nuclei associated with single centrosomes . In # TT46 , clusters containing large ( inset 1 ) or small clusters ( inset 3 ) can be identified . In cyst - like arrangements several centrosomes can be identified ( inset 2 ) , suggesting that all cells have at least one centrosome or , in contrast just one centrosome ( inset 4 ) , illustrating lack of centrosomes .

O O O O O B-DISEASE O O O O O B-GENEPROD O O O O B-GENEPROD O O O O B-SUBCELLULAR O O O O O O O O O O B-SUBCELLULAR O O O O O B-SUBCELLULAR O O O O O O O O O O O O O O O O O O O O O O O B-DISEASE B-SUBCELLULAR O O O O O O B-SUBCELLULAR O O B-SUBCELLULAR O O O O O O O O O O O O O O O O O O O O O O O B-SUBCELLULAR O O O O O O O O O O B-SUBCELLULAR B-EXP_ASSAY O O B-TISSUE O O O O O O O O O O O O B-SUBCELLULAR O O O O O O O O O O O O O O O O O B-SUBCELLULAR O O B-SUBCELLULAR O B-SUBCELLULAR O O O O O B-SUBCELLULAR O O O O O O O B-SUBCELLULAR O O O O O O O B-SUBCELLULAR O O O B-SUBCELLULAR O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-SUBCELLULAR O O O O O O O O O O O O O O O O B-SUBCELLULAR O O O O O O B-SUBCELLULAR O O O O O O O O B-SUBCELLULAR O

(A) Images of EOC tumors labelled with antibodies against PCNT (red) and CDKRAP2 (green). Nuclei are shown in blue. The colored arrows reflect different centrosome number. The list of centrosome behaviors annotated it is not exhaustive to allow comprehension of the events. #TT72, shows a cyst-like arrangement of EOC nuclei where it is impossible to correspond centrosomes to individual nuclei. The white bars with arrows of different colors highlight possible interpretations of the same condition. In #TT79, regions without centrosomes can be identified. In the inset, a single centrosome localized in the lumen can be identified and it is not possible to know to which nucleus (white dashed lines) it corresponds to. #TT88 presents only very small clusters of centrosomes, few nuclei without centrosomes and many conditions with 2 centrosomes. In #TT61, regions without centrosomes can be distinguished, in addition to nuclei associated with single centrosomes. In #TT46, clusters containing large (inset 1) or small clusters (inset 3) can be identified. In cyst-like arrangements several centrosomes can be identified (inset 2), suggesting that all cells have at least one centrosome or, in contrast just one centrosome (inset 4), illustrating lack of centrosomes.


( B - C ) Plots showing the frequency of centrosome amplification events ( B ) and the frequency of nuclei without centrosomes ( C ) in tumor tissues . Note that in B and C , the order of the tumors is conserved between the two plots to allow for comparison . For each tumor , 10 random fields were chosen and analysed , with an average of 5248 nuclei counted per tumor .

O O O O O O O O O O B-SUBCELLULAR O O O O O O O O O B-SUBCELLULAR O B-SUBCELLULAR O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-SUBCELLULAR O O O O

(B-C) Plots showing the frequency of centrosome amplification events (B) and the frequency of nuclei without centrosomes (C) in tumor tissues. Note that in B and C, the order of the tumors is conserved between the two plots to allow for comparison. For each tumor, 10 random fields were chosen and analysed, with an average of 5248 nuclei counted per tumor.


( D ) Tumor sections labelled with CEP135 ( green ) and PCNT ( red ) on the left and CEP135 ( green ) and CEP192 ( red ) on the right . The white squares represent the regions shown in higher magnifications on the right .

O O O O O O O B-GENEPROD O O O O B-GENEPROD O O O O O O O B-GENEPROD O O O O B-GENEPROD O O O O O O O O O O O O O O O O O O O O O

(D) Tumor sections labelled with CEP135 (green) and PCNT (red) on the left and CEP135 (green) and CEP192 (red) on the right. The white squares represent the regions shown in higher magnifications on the right.


( B ) Stills of a time - lapse movie of Ctrl spheroids . Time is shown in hours ( hrs ) . ( C ) z - view of Ctrl cells as shown in ( B ) . Note the red - colored cancer cells at the beginning of the movie on top of the mesothelial layer ( green ) while at later time points , they have cleared through the mesothelial cells .

O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-CELL_TYPE O O O O O O O O O O O O O O O B-CELL_TYPE I-CELL_TYPE O

(B) Stills of a time-lapse movie of Ctrl spheroids. Time is shown in hours (hrs). (C) z- view of Ctrl cells as shown in (B). Note the red-colored cancer cells at the beginning of the movie on top of the mesothelial layer (green) while at later time points, they have cleared through the mesothelial cells.


( D ) Graph bars of the normalized clearance in A . U . of iOVCAR8 ( left ) and iSKOV3 ( right ) spheroids after the indicated treatments . The two cell lines show different clearance capacity . For each cell line , three independent experiments were performed . For iOVCAR8 , we analyzed 47 spheroids in DMSO and 56 spheroids in centrinone . For iSKOV3 we analyzed 56 spheroids in DMSO and 48 spheroids in centrinone . Dots represent the mean and the shadow ± SEM . Statistical significance was assessed with an ANCOVA test . Dots represent the mean and the shadow ± SEM . Statistical significance was assessed with an ANOVA test .

O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O O O O O O O O O O O B-SMALL_MOLECULE O O O O B-SMALL_MOLECULE O O O O O O O O B-SMALL_MOLECULE O O O O B-SMALL_MOLECULE O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

(D) Graph bars of the normalized clearance in A.U. of iOVCAR8 (left) and iSKOV3 (right) spheroids after the indicated treatments. The two cell lines show different clearance capacity. For each cell line, three independent experiments were performed. For iOVCAR8, we analyzed 47 spheroids in DMSO and 56 spheroids in centrinone. For iSKOV3 we analyzed 56 spheroids in DMSO and 48 spheroids in centrinone. Dots represent the mean and the shadow±SEM. Statistical significance was assessed with an ANCOVA test. Dots represent the mean and the shadow±SEM. Statistical significance was assessed with an ANOVA test.


( F ) Representative images of DMSO ( left ) and centrinone ( right ) spheroids . Mean ± SD . Red nuclei represent false colored invading nuclei . ( G ) Dot plot showing the quantification of the number of nuclei detected on the bottom side of the BM . Three or four positions from three BM inserts were analyzed in each condition . Mean ± SD . Statistical significance was assessed with the Mann - Whitney test . For all the experiments , the centrosome number was verified in parallel to confirm the decreased centrosome conditions compared to DMSO .

O O O O O O B-SMALL_MOLECULE O O O O B-SMALL_MOLECULE O O O O O O O O O O B-SUBCELLULAR O O O O B-SUBCELLULAR O O O O O O O O O O O O O B-SUBCELLULAR O O O O O O O B-TISSUE O O O O O O O B-TISSUE O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-SUBCELLULAR O O O O O O O O O B-SUBCELLULAR O O O B-SMALL_MOLECULE O

(F) Representative images of DMSO (left) and centrinone (right) spheroids. Mean±SD. Red nuclei represent false colored invading nuclei. (G) Dot plot showing the quantification of the number of nuclei detected on the bottom side of the BM. Three or four positions from three BM inserts were analyzed in each condition. Mean±SD. Statistical significance was assessed with the Mann-Whitney test. For all the experiments, the centrosome number was verified in parallel to confirm the decreased centrosome conditions compared to DMSO.


Vero and Caco - 2 cells were infected with SARS - CoV - 2 at MOIs of 0 . 2 for 8 h . Infected cells were then permeabilized and immunostained for the intracellular SARS - CoV - 2 nucleoprotein ( NP , red ) . Nuclei were stained with Hoechst ( blue ) before imaging by fluorescence confocal microscopy . Scale bars : 100 µm .

B-CELL_LINE O B-CELL_LINE I-CELL_LINE I-CELL_LINE O O O O B-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM O O O O O O O O O O O O O O O O B-EXP_ASSAY O O O B-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM B-GENEPROD O B-GENEPROD O O O O B-SUBCELLULAR O B-EXP_ASSAY O B-SMALL_MOLECULE O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O O O O O O O

Vero and Caco-2 cells were infected with SARS-CoV-2 at MOIs of 0.2 for 8 h. Infected cells were then permeabilized and immunostained for the intracellular SARS-CoV-2 nucleoprotein (NP, red). Nuclei were stained with Hoechst (blue) before imaging by fluorescence confocal microscopy. Scale bars: 100 µm.


Infection of Vero and Caco - 2 cells was monitored over 24 h using the same flow cytometry - based assay used for the experiment shown in panel D . Infection is given as the total fluorescence associated with the NP protein - positive cells . MFI , mean fluorescence intensity . n = 3 .

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Infection of Vero and Caco-2 cells was monitored over 24 h using the same flow cytometry-based assay used for the experiment shown in panel D. Infection is given as the total fluorescence associated with the NP protein-positive cells. MFI, mean fluorescence intensity. n = 3.


Cells were pretreated with the indicated concentrations of aprotinin ( A ) and SB412515 ( B ) , which are inhibitors of TMPRSS2 and cathepsin L , respectively . Infection of Calu - 3 , Caco - 2 , A549 * , and Vero cells with SARS - CoV - 2 at MOIs of 0 . 3 , 0 . 4 , 0 . 2 , and 0 . 3 , respectively , was achieved in the continuous presence of the drug . Infected cells were quantified by flow cytometry as described in Fig . 1D , and data were normalized to samples where inhibitors had been omitted . n = 2 - 4 biological replicates .

O O O O O O O O B-SMALL_MOLECULE O O O O B-SMALL_MOLECULE O O O O O O O O B-GENEPROD O B-GENEPROD I-GENEPROD O O O O O B-CELL_LINE I-CELL_LINE I-CELL_LINE O B-CELL_LINE I-CELL_LINE I-CELL_LINE O O O O O B-CELL_LINE O O B-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O O O O O O O O O O O O O O O O O O

Cells were pretreated with the indicated concentrations of aprotinin (A) and SB412515 (B), which are inhibitors of TMPRSS2 and cathepsin L, respectively. Infection of Calu-3, Caco-2, A549*, and Vero cells with SARS-CoV-2 at MOIs of 0.3, 0.4, 0.2, and 0.3, respectively, was achieved in the continuous presence of the drug. Infected cells were quantified by flow cytometry as described in Fig. 1D, and data were normalized to samples where inhibitors had been omitted. n = 2-4 biological replicates.


SARS - CoV - 2 particles were bound to A549 * and Vero cells ( MOIs 0 . 2 and 0 . 3 , respectively ) ( C ) or Calu - 3 and Caco - 2 cells ( 0 . 6 and 0 . 5 , respectively ) ( D ) on ice for 90 min and subsequently warmed rapidly to 37 ° C to allow infectious penetration . SB412515 ( 10 µM , C ) or aprotinin ( 30 µM , D ) was added at different times post warming to block further proteolytic activation . Infection was analyzed by flow cytometry , and data were normalized to samples where protease inhibitors had been omitted . n = 2 .

B-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM O O O O O O O B-CELL_LINE O O O O O O O O O O O O O O O O O B-CELL_LINE I-CELL_LINE I-CELL_LINE O B-CELL_LINE I-CELL_LINE I-CELL_LINE O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-SMALL_MOLECULE O O O O O O O B-SMALL_MOLECULE O O O O O O O O O O O O O O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O O O O O O O O O

SARS-CoV-2 particles were bound to A549* and Vero cells (MOIs 0.2 and 0.3, respectively) (C) or Calu-3 and Caco-2 cells (0.6 and 0.5, respectively) (D) on ice for 90 min and subsequently warmed rapidly to 37°C to allow infectious penetration. SB412515 (10 µM, C) or aprotinin (30 µM, D) was added at different times post warming to block further proteolytic activation. Infection was analyzed by flow cytometry, and data were normalized to samples where protease inhibitors had been omitted. n = 2.


Cells were pretreated with endosomal - pH interfering drugs at the indicated concentrations and subsequently infected with SARS - CoV - 2 as described in Fig . 2A and 2B in the continuous presence of NH4Cl ( A ) , chloroquine ( B ) , bafilomycin A1 ( C ) , or concanamycin B ( D ) . The proportion of infected cells was quantified by flow cytometry as described in Fig . 1D , and data were normalized to control samples that were not treated with inhibitors . n = 2 - 6 biological replicates .

O O O O O O O O O O O O O O O O O B-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM O O O O O O O O O O O O O B-SMALL_MOLECULE O O O O B-SMALL_MOLECULE O O O O B-SMALL_MOLECULE I-SMALL_MOLECULE O O O O O B-SMALL_MOLECULE O O O O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

Cells were pretreated with endosomal-pH interfering drugs at the indicated concentrations and subsequently infected with SARS-CoV-2 as described in Fig. 2A and 2B in the continuous presence of NH4Cl (A), chloroquine (B), bafilomycin A1 (C), or concanamycin B (D). The proportion of infected cells was quantified by flow cytometry as described in Fig. 1D, and data were normalized to control samples that were not treated with inhibitors. n = 2-6 biological replicates.


A549 * cells expressing or lacking TMPRSS2 were pretreated with the indicated concentrations of camostat mesylate ( B ) , SB412515 ( C ) , and bafilomycin A1 ( D ) . Infection with SARS - CoV - 2 ( MOI ~ 0 . 2 ) was achieved in the continuous presence of the drug . Infected cells were quantified by flow cytometry as described in Fig . 1D , and data were normalized to samples where inhibitors had been omitted . n = 5 - 6 biological replicates .

O O O O O O B-GENEPROD O O O O O O O B-SMALL_MOLECULE I-SMALL_MOLECULE O O O O B-SMALL_MOLECULE O O O O O B-SMALL_MOLECULE I-SMALL_MOLECULE O O O O O O B-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM O O O O O O O O O O O O O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O O O O O O O O O O O O O O O O O O

A549* cells expressing or lacking TMPRSS2 were pretreated with the indicated concentrations of camostat mesylate (B), SB412515 (C), and bafilomycin A1 (D). Infection with SARS-CoV-2 (MOI ~0.2) was achieved in the continuous presence of the drug. Infected cells were quantified by flow cytometry as described in Fig. 1D, and data were normalized to samples where inhibitors had been omitted. n = 5-6 biological replicates.


The timing of the MG - 132 - sensitive step during SARS - CoV - 2 infectious entry into Calu - 3 and Caco - 2 cells was assayed as detailed in D but using 60 µM MG - 132 for Caco - 2 cells and MOIs of 0 . 5 and 0 . 6 , respectively . n = 2 .

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The timing of the MG-132-sensitive step during SARS-CoV-2 infectious entry into Calu-3 and Caco-2 cells was assayed as detailed in D but using 60 µM MG-132 for Caco-2 cells and MOIs of 0.5 and 0.6, respectively. n = 2.


A549 * cells expressing or lacking TMPRSS2 were pretreated with MG - 132 at the indicated concentrations and subsequently infected with SARS - CoV - 2 ( MOI ~ 0 . 2 ) in the continuous presence of the drug . The proportion of infected cells was quantified by flow cytometry as described in Fig . 1D , and data were normalized to that from control samples for which MG - 132 had been omitted . n = 6 biological replicates .

O O O O O O B-GENEPROD O O O B-SMALL_MOLECULE I-SMALL_MOLECULE I-SMALL_MOLECULE O O O O O O O O B-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM O O O O O O O O O O O O O O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O O O O O O O O O B-SMALL_MOLECULE I-SMALL_MOLECULE I-SMALL_MOLECULE O O O O O O O O O O

A549* cells expressing or lacking TMPRSS2 were pretreated with MG-132 at the indicated concentrations and subsequently infected with SARS-CoV-2 (MOI ~0.2) in the continuous presence of the drug. The proportion of infected cells was quantified by flow cytometry as described in Fig. 1D, and data were normalized to that from control samples for which MG-132 had been omitted. n = 6 biological replicates.


SARS - CoV - 2 was subjected to pretreatment with trypsin and furin for 15 min at 37 ° C prior to infection of Caco - 2 and Vero cells ( MOIs of 0 . 4 and 0 . 3 , respectively ) . Infected cells were quantified by flow cytometry as described in Fig . 1D . Data were normalized to samples not pretreated with trypsin . n = 4 - 6 biological replicates .

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SARS-CoV-2 was subjected to pretreatment with trypsin and furin for 15 min at 37°C prior to infection of Caco-2 and Vero cells (MOIs of 0.4 and 0.3, respectively). Infected cells were quantified by flow cytometry as described in Fig. 1D. Data were normalized to samples not pretreated with trypsin. n = 4-6 biological replicates.


A549 * expressing or lacking TMPRSS2 cells and Vero cells were first infected with SARS - CoV - 2 at MOIs of 0 . 1 and 0 . 2 , respectively , for 24 h and then cocultured for 5 h along with target cells , not infected but prestained with CMFDA , a cytosolic green dye . Cells were subsequently treated with trypsin or furin for 5 min at 37 ° C and left to coculture for an additional hour at 37 ° C . After fixation , nuclei were stained with Hoechst ( blue ) , and infected cells were subjected to immunofluorescence staining against SARS - CoV - 2 nucleoprotein ( magenta ) . Samples were ultimately imaged by confocal fluorescence microscopy . White stars indicate syncytia with at least four nuclei . Scale bars : 100 µm . Images of microscope fields obtained in ( E ) were quantified [ A549 * : n ( no virus ) = 30 , n ( no protease ) = 80 , n ( trypsin ) = 80 , and n ( furin ) = 60 ; A549 * TMPRSS2 : n ( no virus ) = 30 , n ( no protease ) = 80 , n ( trypsin ) = 79 , and n ( furin ) = 60 ; Vero : n ( no virus ) = 40 , n ( no protease ) = 112 , n ( trypsin ) = 114 , and n ( furin ) = 60 ] . The fusion index is given as f = 1 - [ ( number of cells in a field after fusion ) / ( number of nuclei ) ] .

O O O O O B-GENEPROD O O B-CELL_LINE O O O O O B-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-SMALL_MOLECULE O O O O O O O O O O O B-SMALL_MOLECULE O B-SMALL_MOLECULE O O O O O O O O O O O O O O O O O O O O O O O B-SUBCELLULAR O B-EXP_ASSAY O B-SMALL_MOLECULE O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O B-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM B-GENEPROD O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O O B-CELL_TYPE O O O O B-SUBCELLULAR O O O O O O O O O B-EXP_ASSAY O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-SMALL_MOLECULE O O O O O O O B-SMALL_MOLECULE O O O O O O B-GENEPROD O O O O O O O O O O O O O O O O O O O B-SMALL_MOLECULE O O O O O O O B-SMALL_MOLECULE O O O O B-CELL_LINE O O O O O O O O O O O O O O O O O O O B-SMALL_MOLECULE O O O O O O O B-SMALL_MOLECULE O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-SUBCELLULAR O O O

A549* expressing or lacking TMPRSS2 cells and Vero cells were first infected with SARS-CoV-2 at MOIs of 0.1 and 0.2, respectively, for 24 h and then cocultured for 5 h along with target cells, not infected but prestained with CMFDA, a cytosolic green dye. Cells were subsequently treated with trypsin or furin for 5 min at 37°C and left to coculture for an additional hour at 37°C. After fixation, nuclei were stained with Hoechst (blue), and infected cells were subjected to immunofluorescence staining against SARS-CoV-2 nucleoprotein (magenta). Samples were ultimately imaged by confocal fluorescence microscopy. White stars indicate syncytia with at least four nuclei. Scale bars: 100 µm. Images of microscope fields obtained in (E) were quantified [A549*: n (no virus) = 30, n (no protease) = 80, n (trypsin) = 80, and n (furin) = 60; A549* TMPRSS2: n (no virus) = 30, n (no protease) = 80, n (trypsin) = 79, and n (furin) = 60; Vero: n (no virus) = 40, n (no protease) = 112, n (trypsin) = 114, and n (furin) = 60]. The fusion index is given as f = 1 - [(number of cells in a field after fusion)/(number of nuclei)].


SARS - CoV - 2 particles ( MOI of 0 . 3 ) were first subjected to trypsin treatment for 15 min at 37 ° C and then exposed to buffers at the indicated pH for 10 min at 37 ° C , and vice versa . Calu - 3 and Vero cells were then infected at a MOI of 0 . 3 and analyzed by flow cytometry as described in Fig . 1D . n = 4 - 8 biological replicates .

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SARS-CoV-2 particles (MOI of 0.3) were first subjected to trypsin treatment for 15 min at 37°C and then exposed to buffers at the indicated pH for 10 min at 37°C, and vice versa. Calu-3 and Vero cells were then infected at a MOI of 0.3 and analyzed by flow cytometry as described in Fig. 1D. n = 4-8 biological replicates.


SARS - CoV - 2 was first treated with trypsin or thermolysin and then allowed to infect A549 * and Vero cells ( MOIs ~ 0 . 2 and 0 . 3 ) in the continuous presence of SB412515 ( cathepsin L inhibitor ) . Infected cells were quantified by flow cytometry as described in Fig . 1D , and data were normalized to samples where SB412515 had been omitted . n = 4 biological replicates .

B-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM I-ORGANISM O O O O B-SMALL_MOLECULE O B-SMALL_MOLECULE O O O O O O O O B-CELL_LINE O O O O O O O O O O O O O O O O O B-SMALL_MOLECULE O B-GENEPROD I-GENEPROD O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O O O O O B-SMALL_MOLECULE O O O O O O O O O O

SARS-CoV-2 was first treated with trypsin or thermolysin and then allowed to infect A549* and Vero cells (MOIs ~0.2 and 0.3) in the continuous presence of SB412515 (cathepsin L inhibitor). Infected cells were quantified by flow cytometry as described in Fig. 1D, and data were normalized to samples where SB412515 had been omitted. n = 4 biological replicates.


C . Tissue distribution of the Ldb2 protein . Representative western - blot image is shown . Yellow asterisks indicate uncharacterized bands .

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C. Tissue distribution of the Ldb2 protein. Representative western-blot image is shown. Yellow asterisks indicate uncharacterized bands.


D . Expression of Ldb2 in the cerebral cortex . DAPI - stained nuclei visualized ( right ) in the cerebral cortex .

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D. Expression of Ldb2 in the cerebral cortex. DAPI-stained nuclei visualized (right) in the cerebral cortex.


E . Expression of Ldb2 in the amygdala . LA ; lateral amygdala , BLA ; basolateral amygdala , Ce ; central nucleus of amygdala .

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0 0 B-EXP_ASSAY 0 B-GENEPROD 0 0 B-TISSUE 0 B-TISSUE 0 B-TISSUE I-TISSUE 0 B-TISSUE 0 B-TISSUE I-TISSUE 0 B-TISSUE 0 B-TISSUE I-TISSUE I-TISSUE I-TISSUE 0

E. Expression of Ldb2 in the amygdala. LA; lateral amygdala, BLA; basolateral amygdala, Ce; central nucleus of amygdala.


F - G . Expression of Ldb2 in neurons . The merged images obtained with Ldb2 ( stained in green ) and NeuN ( stained in red ) antibodies are shown for cerebral cortex layers I - VI and the amygdala ( F ) . most of Ldb2 - positive cells are NeuN - positive . G . NeuN - positive cells / Ldb2 - positive cells ( left ) and Ldb2 - positive cells / NeuN - positive cells were counted in cerebral cortex layers I - VI .

O O O O B-EXP_ASSAY O B-GENEPROD O B-CELL_TYPE O O O O O O B-GENEPROD O B-EXP_ASSAY O O O O B-GENEPROD O B-EXP_ASSAY O O O O O O O B-TISSUE I-TISSUE I-TISSUE I-TISSUE O O O O B-TISSUE O O O O O O B-GENEPROD O O O O B-GENEPROD O O O O O B-GENEPROD O O O O B-GENEPROD O O O O O O O B-GENEPROD O O O O B-GENEPROD O O O O O O B-TISSUE I-TISSUE I-TISSUE I-TISSUE O O O

F-G. Expression of Ldb2 in neurons. The merged images obtained with Ldb2 (stained in green) and NeuN (stained in red) antibodies are shown for cerebral cortex layers I-VI and the amygdala (F). most of Ldb2-positive cells are NeuN-positive. G. NeuN-positive cells/Ldb2-positive cells (left) and Ldb2-positive cells/NeuN-positive cells were counted in cerebral cortex layers I-VI.


Ldb2 KO mice ( red , n = 10 - 11 ) were compared to control ( blue , n = 10 - 11 ) in Home cage - activity test ( A ) . Voluntary locomotor activities in the home cages were monitored for a week . Data of dark ( left ) and light ( right ) phases are indicated .

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Ldb2 KO mice (red, n = 10-11) were compared to control (blue, n = 10-11) in Home cage-activity test (A). Voluntary locomotor activities in the home cages were monitored for a week. Data of dark (left) and light (right) phases are indicated.


Ldb2 KO mice ( red , n = 10 - 11 ) were compared to control ( blue , n = 10 - 11 ) in Open - field test ( B )

B-GENEPROD O B-ORGANISM O O O O O O O O O O O O O O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O

Ldb2 KO mice (red, n = 10-11) were compared to control (blue, n = 10-11) in Open-field test (B)


Ldb2 KO mice ( red , n = 10 - 11 ) were compared to control ( blue , n = 10 - 11 ) in Acoustic startle response ( C )

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Ldb2 KO mice (red, n = 10-11) were compared to control (blue, n = 10-11) in Acoustic startle response (C)


Ldb2 KO mice ( red , n = 10 - 11 ) were compared to control ( blue , n = 10 - 11 ) in Prepulse - inhibition test ( D )

Ldb2 KO mice (red, n = 10-11) were compared to control (blue, n = 10-11) in Prepulse-inhibition test (D)


Ldb2 KO mice ( red , n = 10 - 11 ) were compared to control ( blue , n = 10 - 11 ) in , Morris water - maze test ( E ) where data in hidden test ( escape latency and moving speed , path length and no movement time ) and probe test ( path length ) are

B-GENEPROD O B-ORGANISM O O O O O O O O O O O O O O O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O B-EXP_ASSAY I-EXP_ASSAY O B-EXP_ASSAY I-EXP_ASSAY O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O O O B-EXP_ASSAY I-EXP_ASSAY O O

Ldb2 KO mice (red, n = 10-11) were compared to control (blue, n = 10-11) in , Morris water-maze test (E) where data in hidden test (escape latency and moving speed, path length and no movement time) and probe test (path length) are shown


Ldb2 KO mice ( red , n = 10 - 11 ) were compared to control ( blue , n = 10 - 11 ) in Y - maze test ( F ) .

B-GENEPROD O B-ORGANISM O O O O O O O O O O O O O O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O O

Ldb2 KO mice (red, n = 10-11) were compared to control (blue, n = 10-11) in Y-maze test (F).


Ldb2 KO mice ( red , n = 10 - 11 ) were compared to control ( blue , n = 10 - 11 ) in Fear - conditioning test ( G ) . Results of contextual ( left ) and cued ( right ) tests are shown .

B-GENEPROD O B-ORGANISM O O O O O O O O O O O O O O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O O O O O O O O O O

Ldb2 KO mice (red, n = 10-11) were compared to control (blue, n = 10-11) in Fear-conditioning test (G). Results of contextual (left) and cued (right) tests are shown.


Ldb2 KO mice ( red , n = 10 - 11 ) were compared to control ( blue , n = 10 - 11 ) in Hot - plate test ( H ) .

Ldb2 KO mice (red, n = 10-11) were compared to control (blue, n = 10-11) in Hot-plate test (H).


Locomotor activity was assessed in the same experimental set as in the open - field test . After acclimation for 120 min , mice ( n = 10 - 12 / group ) were injected ( arrows ) with methylphenidate ( MPD : 3 mg / kg , A ) and their locomotor activity was monitored . In the right panels , cumulative values for pre - and post - injection times are also shown .

B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O O O O O B-ORGANISM O O O O O O O O O O O O O O O B-SMALL_MOLECULE O B-SMALL_MOLECULE O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O O O O O O O O O O O O O

Locomotor activity was assessed in the same experimental set as in the open-field test. After acclimation for 120 min, mice (n = 10-12/group) were injected (arrows) with methylphenidate (MPD: 3 mg/kg, A) and their locomotor activity was monitored. In the right panels, cumulative values for pre- and post-injection times are also shown.


Locomotor activity was assessed in the same experimental set as in the open - field test . After acclimation for 120 min , mice ( n = 10 - 12 / group ) were injected ( arrows ) with MK - 801 ( 0 . 2 mg / kg , B ) and their locomotor activity was monitored . In the right panels , cumulative values for pre - and post - injection times are also shown .

B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O O O O O B-ORGANISM O O O O O O O O O O O O O O O B-SMALL_MOLECULE I-SMALL_MOLECULE I-SMALL_MOLECULE O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

Locomotor activity was assessed in the same experimental set as in the open-field test. After acclimation for 120 min, mice (n = 10-12/group) were injected (arrows) with MK-801 (0.2 mg/kg, B) and their locomotor activity was monitored. In the right panels, cumulative values for pre- and post-injection times are also shown.


C . Popping behavior induced by the injection of MK - 801 ( 0 . 5 mg / kg ) was manually assessed in WT ( n = 4 ) and KO ( n = 6 ) animals . Typically , each episode is constituted by blocks of 5 - 15 successive jumping behaviors . Episode numbers in 20 min ( left ) and total duration ( right ) are shown in a box - whisker plot .

O O B-EXP_ASSAY I-EXP_ASSAY O O O O O B-SMALL_MOLECULE I-SMALL_MOLECULE I-SMALL_MOLECULE O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O O O O O O O O O O O O O O O

C. Popping behavior induced by the injection of MK-801 (0.5 mg/kg) was manually assessed in WT (n = 4) and KO (n = 6) animals. Typically, each episode is constituted by blocks of 5-15 successive jumping behaviors. Episode numbers in 20 min (left) and total duration (right) are shown in a box-whisker plot.


A . Fear - conditioning test . Freezing ( % ) was monitored in the cued ( top ) and contextual ( bottom ) tests before ( pre ) and after ( post ) administration of clozapine ( left ) or haloperidol ( right ) . Data are presented as means ± S . E . M . ( n = 7 - 9 / group ) . * , p < 0 . 05 ; * * * , p < 0 . 001 ( Tukey test ) ; # , p < 0 . 05 ( t - test , KO / saline vs . KO / clozapine ) .

O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O B-EXP_ASSAY O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-SMALL_MOLECULE O O O O B-SMALL_MOLECULE O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-SMALL_MOLECULE O O O O B-SMALL_MOLECULE O O

A. Fear-conditioning test. Freezing (%) was monitored in the cued (top) and contextual (bottom) tests before (pre) and after (post) administration of clozapine (left) or haloperidol (right). Data are presented as means ± S.E.M. (n = 7-9/group). *, p < 0.05; ***, p < 0.001 (Tukey test); #, p < 0.05 (t-test, KO/saline vs. KO/clozapine).


B . Home cage - activity test . Spontaneous locomotor activity of mice chronically fed chow with or without 0 . 24 % ( w / w ) lithium carbonate for two months was monitored for 6 days in the home cage .

O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O B-EXP_ASSAY I-EXP_ASSAY O B-ORGANISM O O O O O O O O O O O O O O O B-SMALL_MOLECULE I-SMALL_MOLECULE O O O O O O O O O O O O O

B. Home cage-activity test. Spontaneous locomotor activity of mice chronically fed chow with or without 0.24% (w/w) lithium carbonate for two months was monitored for 6 days in the home cage.


A . Scatter plots showing Venus - fluorescent intensities and the median amplitude of miniature EPSCs ( mEPSC ) in the LA of the AV - KO mice and their littermates ( AV - WT ) . B . Scatter plots showing Venus - fluorescent intensities and the mean frequency of miniature EPSCs in the LA of the AV - KO and their control littermates ( AV - WT ) . Three typical pictures of Venus - positive cells with various fluorescent intensities are shown . Lines in each panel are regression lines showing inverse correlation of the fluorescent intensity with the mEPSC amplitude ( A ) or the mEPSC frequency ( B ) . The black dotted lines are regression lines for AV - WT mice , the red dotted lines are those for AV - KO mice , and black solid lines are those for all cells .

O O O O O B-GENEPROD O B-EXP_ASSAY I-EXP_ASSAY O O O O O B-EXP_ASSAY I-EXP_ASSAY O B-EXP_ASSAY O O O B-TISSUE O O O O O B-ORGANISM O O O O O O O O O O O O O O B-GENEPROD O B-EXP_ASSAY I-EXP_ASSAY O O O O O B-EXP_ASSAY I-EXP_ASSAY O O B-TISSUE O O O O O O O O O O O O O O O O O O O B-GENEPROD O O O O O B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O O B-EXP_ASSAY O O O O O O B-EXP_ASSAY O O O O O O O O O O O O O O O O B-ORGANISM O O O O O O O O O O O B-ORGANISM O O O O O O O O O O O

A. Scatter plots showing Venus-fluorescent intensities and the median amplitude of miniature EPSCs (mEPSC) in the LA of the AV-KO mice and their littermates (AV-WT). B. Scatter plots showing Venus-fluorescent intensities and the mean frequency of miniature EPSCs in the LA of the AV-KO and their control littermates (AV-WT). Three typical pictures of Venus-positive cells with various fluorescent intensities are shown. Lines in each panel are regression lines showing inverse correlation of the fluorescent intensity with the mEPSC amplitude (A) or the mEPSC frequency (B). The black dotted lines are regression lines for AV-WT mice, the red dotted lines are those for AV-KO mice, and black solid lines are those for all cells.


B . Numbers of Venus - positive neurons in the lateral nucleus ( LA ) and the lateral side of the basal nucleus ( BAl ) of Naive , Unpaired , and Paired groups . Open and filled bars represent AV - WT and AV - KO mice , respectively . Positive neurons of AV - WT was increased only in the Paired group compared to the Naïve and Unpaired groups in the LA ( upper panel ) . AV - KO was increased in the Unpaired as well as the Paired groups compared to the Naïve group Positive neurons both in the Unpaired and Paired AV - KO mice were increased compared to the same groups of the AV - WT mice In the BAl , positive neurons were increased both in the Unpaired and Paired groups compared to the Naïve group in both the AV - WT and AV - KO mice The error bars represent S . E . M . ( n = 5 / group ) .

O O O O B-GENEPROD O O B-CELL_TYPE O O B-TISSUE I-TISSUE O B-TISSUE O O O O O O O B-TISSUE I-TISSUE O B-TISSUE O O O O O O O O O O O O O O O O O O O O O O B-ORGANISM O O O O B-CELL_TYPE O O O O O O O O O O O O O O O O O O O O B-TISSUE O O O O O O O O O O O O O O O O O O O O O O O O O B-CELL_TYPE O O O O O O O O O B-ORGANISM O O O O O O O O O O O O B-ORGANISM O O B-TISSUE O O B-CELL_TYPE O O O O O O O O O O O O O O O O O O O O O O O O B-ORGANISM O O O O O O O O O O O O O O O O O O

B. Numbers of Venus-positive neurons in the lateral nucleus (LA) and the lateral side of the basal nucleus (BAl) of Naive, Unpaired, and Paired groups. Open and filled bars represent AV-WT and AV-KO mice, respectively. Positive neurons of AV-WT was increased only in the Paired group compared to the Naïve and Unpaired groups in the LA (upper panel). AV-KO was increased in the Unpaired as well as the Paired groups compared to the Naïve group Positive neurons both in the Unpaired and Paired AV-KO mice were increased compared to the same groups of the AV-WT mice In the BAl , positive neurons were increased both in the Unpaired and Paired groups compared to the Naïve group in both the AV-WT and AV-KO mice The error bars represent S.E.M. (n =5/group).


C . Numbers of Venus - positive neurons classified by fluorescence intensities in the LA of the AV - WT and AV - KO mice . Weak positive neurons were increased in the Paired group compared to the Naïve group in AV - WT Both Weak and Middle positive neurons were increased in the Paired group compared to the Naïve group in AV - KO The increment of the Middle Venus - positive neurons was larger in the AV - KO compared to AV -

O O O O B-GENEPROD O O B-CELL_TYPE O O B-EXP_ASSAY I-EXP_ASSAY O O B-TISSUE O O O O O O O O O B-ORGANISM O O O B-CELL_TYPE O O O O O O O O O O O O O O O O O O O O B-CELL_TYPE O O O O O O O O O O O O O O O O O O O O B-GENEPROD O O B-CELL_TYPE O O O O O O O O O O O

C. Numbers of Venus-positive neurons classified by fluorescence intensities in the LA of the AV-WT and AV-KO mice. Weak positive neurons were increased in the Paired group compared to the Naïve group in AV-WT Both Weak and Middle positive neurons were increased in the Paired group compared to the Naïve group in AV-KO The increment of the Middle Venus-positive neurons was larger in the AV-KO compared to AV-WT


Results of coimmunoprecipitation between overexpressed LDB2 and interaction candidates LDB2 vs . LDB2 ( A ) LDB2 ( WT , T83N or P170L ) and candidate interactors were tagged with Flag and HA , respectively . HEK293T cells were transfected with the Flag - construct and HA - construct indicated . After Flag - LDB2 was immunoprecipitated with a Flag antibody , the immunocomplexes were analyzed with western blotting with Flag and HA antibodies . The input samples ( input ) were also analyzed . Green and black arrowhead indicate positions for Flag - Ldb2 and HA - proteins , respectively .

O O B-EXP_ASSAY O O B-GENEPROD O B-EXP_ASSAY O B-GENEPROD O O B-GENEPROD O O O B-GENEPROD O O O O O O O O O O O O O B-GENEPROD O B-GENEPROD O O O B-CELL_LINE O O O O O B-GENEPROD O O O B-GENEPROD O O O O O B-GENEPROD O B-GENEPROD O B-EXP_ASSAY O O B-GENEPROD O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O B-GENEPROD O B-GENEPROD O O O O O O O O O O O O O O O O O O O B-GENEPROD O B-GENEPROD O B-GENEPROD O O O O O

Results of coimmunoprecipitation between overexpressed LDB2 and interaction candidates LDB2 vs. LDB2 (A) LDB2 (WT, T83N or P170L) and candidate interactors were tagged with Flag and HA, respectively. HEK293T cells were transfected with the Flag-construct and HA-construct indicated. After Flag-LDB2 was immunoprecipitated with a Flag antibody, the immunocomplexes were analyzed with western blotting with Flag and HA antibodies. The input samples (input) were also analyzed. Green and black arrowhead indicate positions for Flag-Ldb2 and HA-proteins, respectively.


Results of coimmunoprecipitation between overexpressed LDB2 and interaction candidates LDB2 vs . LHX2 ( B ) LDB2 ( WT , T83N or P170L ) and candidate interactors were tagged with Flag and HA , respectively . HEK293T cells were transfected with the Flag - construct and HA - construct indicated . After Flag - LDB2 was immunoprecipitated with a Flag antibody , the immunocomplexes were analyzed with western blotting with Flag and HA antibodies . The input samples ( input ) were also analyzed . Green and black arrowhead indicate positions for Flag - Ldb2 and HA - proteins , respectively .

Results of coimmunoprecipitation between overexpressed LDB2 and interaction candidates LDB2 vs. LHX2 (B) LDB2 (WT, T83N or P170L) and candidate interactors were tagged with Flag and HA, respectively. HEK293T cells were transfected with the Flag-construct and HA-construct indicated. After Flag-LDB2 was immunoprecipitated with a Flag antibody, the immunocomplexes were analyzed with western blotting with Flag and HA antibodies. The input samples (input) were also analyzed. Green and black arrowhead indicate positions for Flag-Ldb2 and HA-proteins, respectively.


Results of coimmunoprecipitation between overexpressed LDB2 and interaction candidates LDB2 vs . LMO2 / 3 / 4 ( C ) LDB2 ( WT , T83N or P170L ) and candidate interactors were tagged with Flag and HA , respectively . HEK293T cells were transfected with the Flag - construct and HA - construct indicated . After Flag - LDB2 was immunoprecipitated with a Flag antibody , the immunocomplexes were analyzed with western blotting with Flag and HA antibodies . The input samples ( input ) were also analyzed . Green and black arrowhead indicate positions for Flag - Ldb2 and HA - proteins , respectively .

O O B-EXP_ASSAY O O B-GENEPROD O B-EXP_ASSAY O B-GENEPROD O O B-GENEPROD O O O O O O O B-GENEPROD O O O O O O O O O O O O O B-GENEPROD O B-GENEPROD O O O B-CELL_LINE O O O O O B-GENEPROD O O O B-GENEPROD O O O O O B-GENEPROD O B-GENEPROD O B-EXP_ASSAY O O B-GENEPROD O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O B-GENEPROD O B-GENEPROD O O O O O O O O O O O O O O O O O O O B-GENEPROD O B-GENEPROD O B-GENEPROD O O O O O

Results of coimmunoprecipitation between overexpressed LDB2 and interaction candidates LDB2 vs. LMO2/3/4 (C) LDB2 (WT, T83N or P170L) and candidate interactors were tagged with Flag and HA, respectively. HEK293T cells were transfected with the Flag-construct and HA-construct indicated. After Flag-LDB2 was immunoprecipitated with a Flag antibody, the immunocomplexes were analyzed with western blotting with Flag and HA antibodies. The input samples (input) were also analyzed. Green and black arrowhead indicate positions for Flag-Ldb2 and HA-proteins, respectively.


Results of coimmunoprecipitation between overexpressed LDB2 and interaction candidates , LDB2 vs . SSBP2 / 3 / 4 ( D ) LDB2 ( WT , T83N or P170L ) and candidate interactors were tagged with Flag and HA , respectively . HEK293T cells were transfected with the Flag - construct and HA - construct indicated . After Flag - LDB2 was immunoprecipitated with a Flag antibody , the immunocomplexes were analyzed with western blotting with Flag and HA antibodies . The input samples ( input ) were also analyzed . Green and black arrowhead indicate positions for Flag - Ldb2 and HA - proteins , respectively .

O O B-EXP_ASSAY O O B-GENEPROD O B-EXP_ASSAY O O B-GENEPROD O O B-GENEPROD O O O O O O O B-GENEPROD O O O O O O O O O O O O O B-GENEPROD O B-GENEPROD O O O B-CELL_LINE O O O O O B-GENEPROD O O O B-GENEPROD O O O O O B-GENEPROD O B-GENEPROD O B-EXP_ASSAY O O B-GENEPROD O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O B-GENEPROD O B-GENEPROD O O O O O O O O O O O O O O O O O O O B-GENEPROD O B-GENEPROD O B-GENEPROD O O O O O

Results of coimmunoprecipitation between overexpressed LDB2 and interaction candidates , LDB2 vs. SSBP2/3/4 (D) LDB2 (WT, T83N or P170L) and candidate interactors were tagged with Flag and HA, respectively. HEK293T cells were transfected with the Flag-construct and HA-construct indicated. After Flag-LDB2 was immunoprecipitated with a Flag antibody, the immunocomplexes were analyzed with western blotting with Flag and HA antibodies. The input samples (input) were also analyzed. Green and black arrowhead indicate positions for Flag-Ldb2 and HA-proteins, respectively.


Results of coimmunoprecipitation between overexpressed LDB2 and interaction candidates LDB2 vs . RLIM ( E ) . LDB2 ( WT , T83N or P170L ) and candidate interactors were tagged with Flag and HA , respectively . HEK293T cells were transfected with the Flag - construct and HA - construct indicated . After Flag - LDB2 was immunoprecipitated with a Flag antibody , the immunocomplexes were analyzed with western blotting with Flag and HA antibodies . The input samples ( input ) were also analyzed . Green and black arrowhead indicate positions for Flag - Ldb2 and HA - proteins , respectively .

O O B-EXP_ASSAY O O B-GENEPROD O B-EXP_ASSAY O B-GENEPROD O O B-GENEPROD O O O O B-GENEPROD O O O O O O O O O O O O O B-GENEPROD O B-GENEPROD O O O B-CELL_LINE O O O O O B-GENEPROD O O O B-GENEPROD O O O O O B-GENEPROD O B-GENEPROD O B-EXP_ASSAY O O B-GENEPROD O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O B-GENEPROD O B-GENEPROD O O O O O O O O O O O O O O O O O O O B-GENEPROD O B-GENEPROD O B-GENEPROD O O O O O

Results of coimmunoprecipitation between overexpressed LDB2 and interaction candidates LDB2 vs. RLIM (E). LDB2 (WT, T83N or P170L) and candidate interactors were tagged with Flag and HA, respectively. HEK293T cells were transfected with the Flag-construct and HA-construct indicated. After Flag-LDB2 was immunoprecipitated with a Flag antibody, the immunocomplexes were analyzed with western blotting with Flag and HA antibodies. The input samples (input) were also analyzed. Green and black arrowhead indicate positions for Flag-Ldb2 and HA-proteins, respectively.


F . Coimmunoprecipitation of endogenous Ssbp3 and Lmo3 with Ldb2 . Immunocomplexes from mouse brain lysate using an antibody against Ldb2 , Lmo3 or Ssbp3 , or a control antibody were analyzed by western blotting using an anti - Ldb2 ( left ) or anti - Ssbp3 ( right ) antibody .

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F. Coimmunoprecipitation of endogenous Ssbp3 and Lmo3 with Ldb2. Immunocomplexes from mouse brain lysate using an antibody against Ldb2, Lmo3 or Ssbp3, or a control antibody were analyzed by western blotting using an anti-Ldb2 (left) or anti-Ssbp3 (right) antibody.


A . Human neurosphere ( bottom ) induced from iPS cells ( top ) from a healthy control subject .

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A. Human neurosphere (bottom) induced from iPS cells (top) from a healthy control subject.


C . Overlap of ChIP - seq peaks of LDB2 in neurosphere cells and ChIP - seq peaks of EGR1 in human cultured cell lines ECC1 and K562 , at

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C. Overlap of ChIP-seq peaks of LDB2 in neurosphere cells and ChIP-seq peaks of EGR1 in human cultured cell lines ECC1 and K562, at ARC


D . Genome - wide pattern of overlap between the ChIP - seq peaks of LDB2 in neurosphere cells and the ChIP - seq peaks of EGR1 in the ECC1 cell line . Fold enrichment ( log2 ) of ChIP - seq reads against the input reads were calculated for each ChIP - seq peak region ( summit ± 1 , 000 bp ) in each category ( neurosphere specific , common , and ECC1 specific ) and displayed in a heatmap by ngs .

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D. Genome-wide pattern of overlap between the ChIP-seq peaks of LDB2 in neurosphere cells and the ChIP-seq peaks of EGR1 in the ECC1 cell line. Fold enrichment (log2) of ChIP-seq reads against the input reads were calculated for each ChIP-seq peak region (summit ± 1,000 bp) in each category (neurosphere specific, common, and ECC1 specific) and displayed in a heatmap by ngs.plot


E . Annotation of ChIP - seq peaks . ChIP - seq peaks of LDB2 in the neurosphere cells ( all , neurosphere specific , and common were analyzed by the PAVIS

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E. Annotation of ChIP-seq peaks. ChIP-seq peaks of LDB2 in the neurosphere cells (all, neurosphere specific, and common were analyzed by the PAVIS program


Westernblot of producer cell lysates and concentrated reporter particles .

Westernblot of producer cell lysates and concentrated reporter particles.


BHK cells were transfected with either human ACE2 or empty vector , subsequently infected with VSV - reporter particles pseudotyped with chimeric spikes , luciferase was measured and normalized to no pseudotype as a readout for cell entry . Shown are the data for 3 replicates .

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BHK cells were transfected with either human ACE2 or empty vector, subsequently infected with VSV-reporter particles pseudotyped with chimeric spikes, luciferase was measured and normalized to no pseudotype as a readout for cell entry. Shown are the data for 3 replicates.


Primate cells , were infected with VSV - particles pseudotyped with the lineage B chimeric spike panel . Pseudotypes were either left untreated or incubated with trypsin before addition to the cells . Luciferase was measured and normalized to particles produced without pseudotype . Shown are the data for 3 replicates .

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Primate cells, were infected with VSV-particles pseudotyped with the lineage B chimeric spike panel. Pseudotypes were either left untreated or incubated with trypsin before addition to the cells. Luciferase was measured and normalized to particles produced without pseudotype. Shown are the data for 3 replicates.


human cells were infected with VSV - particles pseudotyped with the lineage B chimeric spike panel . Pseudotypes were either left untreated or incubated with trypsin before addition to the cells . Luciferase was measured and normalized to particles produced without pseudotype . Shown are the data for 3 replicates .

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human cells were infected with VSV-particles pseudotyped with the lineage B chimeric spike panel. Pseudotypes were either left untreated or incubated with trypsin before addition to the cells. Luciferase was measured and normalized to particles produced without pseudotype. Shown are the data for 3 replicates.


bat cells were infected with VSV - particles pseudotyped with the lineage B chimeric spike panel . Pseudotypes were either left untreated or incubated with trypsin before addition to the cells . Luciferase was measured and normalized to particles produced without pseudotype . Shown are the data for 3 replicates .

bat cells were infected with VSV-particles pseudotyped with the lineage B chimeric spike panel. Pseudotypes were either left untreated or incubated with trypsin before addition to the cells. Luciferase was measured and normalized to particles produced without pseudotype. Shown are the data for 3 replicates.


Pseudotyped particles were either left untreated and treated with trypsin and subsequently used to infect BHK cells transfected with the coronavirus receptor indicated . Shown are data from 3 replicates .

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Pseudotyped particles were either left untreated and treated with trypsin and subsequently used to infect BHK cells transfected with the coronavirus receptor indicated. Shown are data from 3 replicates.


Expression and pseudotype incorporation of SARS - S - 2019 - nCoV RBD chimeras .

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Expression and pseudotype incorporation of SARS-S-2019-nCoV RBD chimeras.


Pseudotypes were used to infect cells expressing hACE2 , hDPP4 , hAPN , or empty vector , without protease treatment .

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Pseudotypes were used to infect cells expressing hACE2, hDPP4, hAPN, or empty vector, without protease treatment.


VSV pseudotypes were generated with the indicated RBD and used to infect BHKs transfected with either human ACE2 or empty vector . Shown are data for 3 replicates .

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VSV pseudotypes were generated with the indicated RBD and used to infect BHKs transfected with either human ACE2 or empty vector. Shown are data for 3 replicates.


Westernblot of producer cell lysates and concentrated pseudotyped particles .

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Westernblot of producer cell lysates and concentrated pseudotyped particles.


Westernblot of producer cell lysates and concentrated pseudotypes particles .

Westernblot of producer cell lysates and concentrated pseudotypes particles.


Pseudotypes with indicated spike constructs were left untreated or treated with trypsin and subsequently used to infect Huh - 7 . 5 cells . Shown are data for 3 replicates .

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Pseudotypes with indicated spike constructs were left untreated or treated with trypsin and subsequently used to infect Huh-7.5 cells. Shown are data for 3 replicates.


Pseudotypes with indicated spike constructs were left untreated or treated with trypsin and subsequently used to infect BHK cells transfected with human ACE2 . Shown are data for 3 replicates .

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Pseudotypes with indicated spike constructs were left untreated or treated with trypsin and subsequently used to infect BHK cells transfected with human ACE2. Shown are data for 3 replicates.


D . BiFC analysis of interactions between VISP1 or VISP1mUIM and ATG8a or ATG8f in N . benthamiana leaves . Images were taken at 60 hpi . Scale bar = 20 μm .

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D. BiFC analysis of interactions between VISP1 or VISP1mUIM and ATG8a or ATG8f in N. benthamiana leaves. Images were taken at 60 hpi. Scale bar = 20 μm.


E . Co - IP experiments examining protein interactions between ATG8a with VISP1 or VISP1mUIM . N . benthamiana leaves were agroinfiltrated and treated with 2 mM 3 - MA at 48 hpi , and then for IP with anti - Flag beads 12 h later .

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E. Co-IP experiments examining protein interactions between ATG8a with VISP1 or VISP1mUIM. N. benthamiana leaves were agroinfiltrated and treated with 2 mM 3-MA at 48 hpi, and then for IP with anti-Flag beads 12 h later.


Confocal microscopy of YFP - ATG8e - labeled autophagic bodies in root cells of Col - 0 / YFP - ATG8e and VISP1OE / YFP - ATG8e plants . 7 - day - old seedlings were inoculated in liquid ½MS medium with 1 µM ConA for 12 h in dark . Scale bar = 20 μm . Puncta numbers of YFP - ATG8e - labeled autophagic bodies per 10 cells in panel A . Data points represent means of three biological repeats . Error bars indicate SD . * * * p < 0 . 001 ( Student ' s t - test ) .

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Confocal microscopy of YFP-ATG8e-labeled autophagic bodies in root cells of Col-0/YFP-ATG8e and VISP1OE/YFP-ATG8e plants. 7-day-old seedlings were inoculated in liquid ½MS medium with 1 µM ConA for 12 h in dark. Scale bar = 20 μm.   Puncta numbers of YFP-ATG8e-labeled autophagic bodies per 10 cells in panel A. Data points represent means of three biological repeats. Error bars indicate SD. ***p < 0.001 (Student's t-test).


Western blotting analysis of accumulation of ATG8a and ATG8a - PE in leaves of 4 - week - old Col - 0 , VISP1OE , VISP1mUIM / OE and VISP1mARM / OE plants .

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Western blotting analysis of accumulation of ATG8a and ATG8a-PE in leaves of 4-week-old Col-0, VISP1OE, VISP1mUIM/OE and VISP1mARM/OE plants.


Representative TEM images of autophagic structures in inoculated leaves of Col - 0 and VISP1OE plants treated with mock buffer or CMV - 2blm infection at 4 dpi . The leaves was treated with 1 µM ConA at 4 dpi for 12 h in dark before TEM sampling . Magenta arrowheads indicate autophagic bodies inside the vacuoles . Scale bar = 2 μm . Cp , chloroplast ; CW , cell wall ; V , vacuole . Autophagic structure numbers per 10 cells in panel F . Error bars indicate SD . Letters indicate significant differences ( ANOVA , P < 0 . 05 ) .

O O O O B-SUBCELLULAR O O O B-TISSUE O O O O O O O O O O O O B-ORGANISM I-ORGANISM I-ORGANISM O O O O O O B-TISSUE O O O O O B-SMALL_MOLECULE O O O O O O O O O O O O O O O B-SUBCELLULAR I-SUBCELLULAR O O B-SUBCELLULAR O O O O O O O B-SUBCELLULAR O B-SUBCELLULAR O B-SUBCELLULAR O B-SUBCELLULAR I-SUBCELLULAR O O O B-SUBCELLULAR O B-SUBCELLULAR O O O O O O O O O O O O O O O O O O O O O O O O O O O O

Representative TEM images of autophagic structures in inoculated leaves of Col-0 and VISP1OE plants treated with mock buffer or CMV-2blm infection at 4 dpi. The leaves was treated with 1 µM ConA at 4 dpi for 12 h in dark before TEM sampling. Magenta arrowheads indicate autophagic bodies inside the vacuoles. Scale bar = 2 μm. Cp, chloroplast; CW, cell wall; V, vacuole.   Autophagic structure numbers per 10 cells in panel F. Error bars indicate SD. Letters indicate significant differences (ANOVA, P < 0.05).


Long - term symptoms of Col - 0 , VISP1OE1 , VISP1OE2 , rdr6 - 15 , and sgs3 - 1 plants inoculated with CMV - 2blm at 50 dpi . Scale bar = 2 cm .

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Long-term symptoms of Col-0, VISP1OE1, VISP1OE2, rdr6-15, and sgs3-1 plants inoculated with CMV-2blm at 50 dpi. Scale bar = 2 cm.


Northern blotting analyzing accumulation of genomic RNAs ( gRNAs ) in systemically leaves of Col - 0 , OE1 ( VISP1OE1 ) , OE2 ( VISP1OE2 ) , rdr6 - 15 , and sgs3 - 1 plants infected by CMV - 2blm at 14 dpi . Methylene blue - stained rRNA were used as loading controls . Northern blotting analyzing accumulation of RNA3 - derived vsiRNAs in the same samples as shown in panel D .

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Northern blotting analyzing accumulation of genomic RNAs (gRNAs) in systemically leaves of Col-0, OE1 (VISP1OE1), OE2 (VISP1OE2), rdr6-15, and sgs3-1 plants infected by CMV-2blm at 14 dpi. Methylene blue-stained rRNA were used as loading controls.   Northern blotting analyzing accumulation of RNA3-derived   vsiRNAs in the same samples as shown in panel D.


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Long-term systemic symptoms of Col-0, visp1-1, visp1-2, rdr6-15, and sgs3-1 plants inoculated with CMV-2blm at 50 dpi. Scale bar = 2 cm.


Northern blotting detecting accumulation of genomic RNAs and RNA3 - derived vsiRNAs in systemically infected leaves at 14 dpi . Relative accumulation ( RA ) of gRNA3 was calculated from band densities . Northern blotting analyzing accumulation of RNA3 - derived vsiRNAs in the same samples as shown in panel D . Bottom values represent relative accumulation ( RA ) of RNA3 - derived vsiRNAs .

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Northern blotting detecting accumulation of genomic RNAs and RNA3-derived vsiRNAs in systemically infected leaves at 14 dpi. Relative accumulation (RA) of gRNA3 was calculated from band densities.   Northern blotting analyzing accumulation of RNA3-derived vsiRNAs in the same samples as shown in panel D. Bottom values represent relative accumulation (RA) of RNA3-derived vsiRNAs.


Co - immunoprecipitation analyses of the interactions between SGS3 , SGS3ΔCC , or SGS3CC and VISP1 or VISP1mARM . N . benthamiana leaves were agroinfiltrated with constructs as indicated and treated with 2 mM 3 - MA at 48 hpi , and collected for IP with anti - Flag beads 12 h later .

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Co-immunoprecipitation analyses of the interactions between SGS3, SGS3ΔCC, or SGS3CC and VISP1 or VISP1mARM. N. benthamiana leaves were agroinfiltrated with constructs as indicated and treated with 2 mM 3-MA at 48 hpi, and collected for IP with anti-Flag beads 12 h later.


GST pull - down analysis of the in vitro interactions of SGS3CC with VISP1 , VISP1mARM or VISP1mUIM . GST - VISP1 , GST - VISP1mARM , GST - VISP1mUIM or GST were incubated with SGS3CC - His for IP with anti - GST beads , and detected by Western blotting with GST or His antibodies .

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GST pull-down analysis of the in vitro interactions of SGS3CC with VISP1, VISP1mARM or VISP1mUIM. GST- VISP1, GST- VISP1mARM, GST- VISP1mUIM or GST were incubated with SGS3CC-His for IP with anti-GST beads, and detected by Western blotting with GST or His antibodies.


Confocal analysis of co - localization of CFP - NbATG8f - labelled autophagic bodies with the VISP1 - YN and SGS3 - YC bodies in N . benthamiana leaves . The infiltrated leaves were treated with 100 µM E64d or DMSO at 48 hpi and examined for imaging at 60 hpi . Scale bar = 20 μm .

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Confocal analysis of co-localization of CFP-NbATG8f-labelled autophagic bodies with the VISP1-YN and SGS3-YC bodies in N. benthamiana leaves. The infiltrated leaves were treated with 100 µM E64d or DMSO at 48 hpi and examined for imaging at 60 hpi. Scale bar = 20 μm.


Confocal analysis of effect of VISP1 - Flag VISP1mARM - Flag , and VISP1mUIM - Flag on RDR6 - YN and SGS3 - YC formed bodies in N . benthamiana leaves at 60 hpi . EV : empty vector . Scale bar = 20 μm . Numbers of RDR6 - YN and SGS3 - YC formed bodies per 10 cells in E were used for quantification . Data points represent means of three biological repeats . Error bars indicate SD . Letters indicate significant differences ( ANOVA , P < 0 . 05 ) .

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Confocal analysis of effect of VISP1-Flag VISP1mARM-Flag, and VISP1mUIM-Flag on RDR6-YN and SGS3-YC formed bodies in N. benthamiana leaves at 60 hpi. EV: empty vector. Scale bar = 20 μm.   Numbers of RDR6-YN and SGS3-YC formed bodies per 10 cells in E were used for quantification. Data points represent means of three biological repeats. Error bars indicate SD. Letters indicate significant differences (ANOVA, P < 0.05).


Western blotting detecting accumulation of the endogenous SGS3 protein in leaves of 2 - week - old Col - 0 , VISP1OE , visp1 , and sgs3 - 1 Arabidopsis plants with anti - SGS3 antibodies . Western blotting detecting accumulation of the endogenous SGS3 protein in leaves of 2 - week - old Col - 0 , VISP1OE , VISP1mUIM / OE , and VISP1mARM / OE Arabidopsis plants . Western blotting detecting accumulation of the endogenous SGS3 protein in 2 - week - old VISP1 - overexpressing Col - 0 , atg5 - 1 , and atg7 - 1 mutants .

B-EXP_ASSAY I-EXP_ASSAY O O O O O B-GENEPROD O O B-TISSUE O O O O O O O O O O O O O O O B-GENEPROD O O B-ORGANISM O O O O B-GENEPROD O O B-EXP_ASSAY I-EXP_ASSAY O O O O O B-GENEPROD O O B-TISSUE O O O O O O O O O O O O O O O O O O O O B-ORGANISM O O B-EXP_ASSAY I-EXP_ASSAY O O O O O B-GENEPROD O O O O O O O O O B-EXP_ASSAY O O O O B-GENEPROD O O O O B-GENEPROD O O O O

Western blotting detecting accumulation of the endogenous SGS3 protein in leaves of 2-week-old Col-0, VISP1OE, visp1, and sgs3-1 Arabidopsis plants with anti-SGS3 antibodies.   Western blotting detecting accumulation of the endogenous SGS3 protein in leaves of 2-week-old Col-0, VISP1OE, VISP1mUIM/OE, and VISP1mARM/OE Arabidopsis plants.   Western blotting detecting accumulation of the endogenous SGS3 protein in 2-week-old VISP1-overexpressing Col-0, atg5-1, and atg7-1 mutants.


Developmental phenotypes of rosette leaves of 5 - week - old Col - 0 , visp1 - 1 , visp1 - 2 , VISP1OE1 , VISP1OE2 , and sgs3 - 1 seedlings . Arrowheads indicate the 8th and 9th rosette leaves from the top leaves . Scale bar = 2 cm . The length / width ratios of the 8th and 9th rosette leaves of the seedlings shown in A . Every dot represents data from the length / width ratio of one leave ( n = 20 ) . Letters indicate significant differences ( ANOVA , P < 0 . 05 ) .

O O O B-TISSUE I-TISSUE O O O O O O O O O O O O O O O O O O O O O O O B-GENEPROD O O B-ORGANISM O O O O O O O B-TISSUE I-TISSUE O O O B-TISSUE O O O O O O O O O O O O O O O O O B-TISSUE I-TISSUE O O B-ORGANISM O O O O O O O O O O O O O O O O B-TISSUE O O O O O O O O O O O O O O O O O O O O

Developmental phenotypes of rosette leaves of 5-week-old Col-0, visp1-1, visp1-2, VISP1OE1, VISP1OE2, and sgs3-1 seedlings. Arrowheads indicate the 8th and 9th rosette leaves from the top leaves. Scale bar = 2 cm.   The length/width ratios of the 8th and 9th rosette leaves of the seedlings shown in A. Every dot represents data from the length/width ratio of one leave (n = 20). Letters indicate significant   differences (ANOVA, P < 0.05).


Northern blotting detecting accumulation of TAS1 - TasiR255 , TAS2 - TasiR1511 , TAS3 - TasiR5 ′ D8 , miR173 , and miR390 in leaves of Arabidopsis plants shown in panel A . Values represent relative accumulation ( RA ) of siRNAs and values in Col - 0 plants were set as 1 . rRNA served as loading controls .

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Northern blotting detecting accumulation of TAS1-TasiR255, TAS2-TasiR1511, TAS3-TasiR5′D8, miR173, and miR390 in leaves of Arabidopsis plants shown in panel A. Values represent relative accumulation (RA) of siRNAs and values in Col-0 plants were set as 1. rRNA served as loading controls.


B - C Competitor donor derived Gr , B , CD4 T , and CD8 T cells in the peripheral blood . In the control group , competitor donor cells are WT HSCs . In the deficient group , competitor donor cells are uMT - / - or NSG HSCs Data information : Data are shown as percentages of the total number of white blood cells ( WBCs ) . Data were collected at month 7 post transplantation for the uMT - / - group and month 8 post - transplantation for the NSG group , and presented as mean ± SEM . n = 7 mice for the uMT - / - group and n = 6 for the NSG group . * p < 0 . 05 , * * p < 0 . 01 , * * * p < 0 . 001 ( two - tailed and two - sample equal variance Student " s t - test )

O O O O O O B-CELL_TYPE O B-CELL_TYPE O B-CELL_TYPE I-CELL_TYPE O O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O O B-TISSUE I-TISSUE O O O O O O O O O O O B-CELL_TYPE O O O O O O O O O O B-GENEPROD O O O O B-GENEPROD B-CELL_TYPE O O O O O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O B-CELL_TYPE O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-ORGANISM O O B-GENEPROD O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

B-C Competitor donor derived Gr, B, CD4 T, and CD8 T cells in the peripheral blood. In the control group, competitor donor cells are WT HSCs. In the deficient group, competitor donor cells are uMT-/- or NSG HSCs Data information: Data are shown as percentages of the total number of white blood cells (WBCs). Data were collected at month 7 post transplantation for the uMT-/- group and month 8 post-transplantation for the NSG group, and presented as mean ± SEM. n=7 mice for the uMT-/- group and n=6 for the NSG group. *p<0.05, **p<0.01, ***p<0.001 (two-tailed and two-sample equal variance Student"s t-test)


D - E WT donor derived Gr , B , CD4 T , and CD8 T cells in the peripheral blood Data information : Data are shown as percentages of the total number of white blood cells ( WBCs ) . Data were collected at month 7 post transplantation for the uMT - / - group and month 8 post - transplantation for the NSG group , and presented as mean ± SEM . n = 7 mice for the uMT - / - group and n = 6 for the NSG group . * p < 0 . 05 , * * p < 0 . 01 , * * * p < 0 . 001 ( two - tailed and two - sample equal variance Student " s t - test )

O O O O O O B-CELL_TYPE O B-CELL_TYPE O B-CELL_TYPE I-CELL_TYPE O O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O O B-TISSUE I-TISSUE O O O O O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O B-CELL_TYPE O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-ORGANISM O O B-GENEPROD O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

D-E WT donor derived Gr, B, CD4 T, and CD8 T cells in the peripheral blood Data information: Data are shown as percentages of the total number of white blood cells (WBCs). Data were collected at month 7 post transplantation for the uMT-/- group and month 8 post-transplantation for the NSG group, and presented as mean ± SEM. n=7 mice for the uMT-/- group and n=6 for the NSG group. *p<0.05, **p<0.01, ***p<0.001 (two-tailed and two-sample equal variance Student"s t-test)


F - G Total production of Gr , B , CD4 T , and CD8 T cells in the peripheral blood ( derived from WT HSCs , competitor HSCs , helper whole bone marrow cells , and residual host cells ) shown as percentages of the total number of white blood cells ( WBCs ) . Data information : Data are shown as percentages of the total number of white blood cells ( WBCs ) . Data were collected at month 7 post transplantation for the uMT - / - group and month 8 post - transplantation for the NSG group , and presented as mean ± SEM . n = 7 mice for the uMT - / - group and n = 6 for the NSG group . * p < 0 . 05 , * * p < 0 . 01 , * * * p < 0 . 001 ( two - tailed and two - sample equal variance Student " s t - test )

O O O O O O B-CELL_TYPE O B-CELL_TYPE O B-CELL_TYPE I-CELL_TYPE O O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O O B-TISSUE I-TISSUE O O O O B-CELL_TYPE O O B-CELL_TYPE O O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O B-CELL_TYPE O O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O B-CELL_TYPE O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-ORGANISM O O B-GENEPROD O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

F-G Total production of Gr, B, CD4 T, and CD8 T cells in the peripheral blood (derived from WT HSCs, competitor HSCs, helper whole bone marrow cells, and residual host cells) shown as percentages of the total number of white blood cells (WBCs). Data information: Data are shown as percentages of the total number of white blood cells (WBCs). Data were collected at month 7 post transplantation for the uMT-/- group and month 8 post-transplantation for the NSG group, and presented as mean ± SEM. n=7 mice for the uMT-/- group and n=6 for the NSG group. *p<0.05, **p<0.01, ***p<0.001 (two-tailed and two-sample equal variance Student"s t-test)


A - B Total numbers of barcoded WT clones that give rise to granulocytes ( Gr ) , B cells , CD4 T cells , and CD8 T cells Data information : Data were collected at month 7 post - transplantation for the uMT - / - group and month 8 post - transplantation for the NSG group , and presented as mean ± SEM . n = 7 mice for the uMT - / - group and n = 6 for the NSG group . * p < 0 . 05 ( two - tailed and two - sample equal variance Student " s t - test )

O O O B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O B-CELL_TYPE O B-CELL_TYPE O O B-CELL_TYPE I-CELL_TYPE O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O O O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-ORGANISM O O B-GENEPROD O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O O O O O O O O O O O O O O

A-B Total numbers of barcoded WT clones that give rise to granulocytes (Gr), B cells, CD4 T cells, and CD8 T cells Data information: Data were collected at month 7 post-transplantation for the uMT-/- group and month 8 post-transplantation for the NSG group, and presented as mean ± SEM. n=7 mice for the uMT-/- group and n=6 for the NSG group. *p<0.05 (two-tailed and two-sample equal variance Student"s t-test)


C - J Number of HSC clones that produce different amounts of Gr , B , CD4 T , and CD8 T cells . We combined the sequencing data with the flow cytometry data to calculate clonal abundance for each clone as follows : Clonal abundance = 100 % * ( Each cell population ( Gr , B , CD4T or CD8T cells ) % WBCs ) * ( Donor % Each cell population ) * ( GFP % Donor cells ) * ( number of reads for each barcode ) / ( total reads of all barcodes Data information : Data were collected at month 7 post - transplantation for the uMT - / - group and month 8 post - transplantation for the NSG group , and presented as mean ± SEM . n = 7 mice for the uMT - / - group and n = 6 for the NSG group . * p < 0 . 05 ( two - tailed and two - sample equal variance Student " s t - test )

O O O B-EXP_ASSAY O B-CELL_TYPE O O O O O O B-CELL_TYPE O B-CELL_TYPE O B-CELL_TYPE I-CELL_TYPE O O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O O O O O O O O B-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O O O O O O O O O O O O O B-CELL_TYPE O B-CELL_TYPE O B-CELL_TYPE O B-CELL_TYPE O O O B-CELL_TYPE O O O O O O O O O O O B-GENEPROD O O O O O O B-EXP_ASSAY O O O O O O O O O O O O O O O O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-ORGANISM O O B-GENEPROD O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O O O O O O O O O O O O O O

C-J Number of HSC clones that produce different amounts of Gr, B, CD4 T, and CD8 T cells. We combined the sequencing data with the flow cytometry data to calculate clonal abundance for each clone as follows: Clonal abundance =100%*(Each cell population (Gr, B, CD4T or CD8T cells) % WBCs) * (Donor % Each cell population) * (GFP % Donor cells) * (number of reads for each barcode) / (total reads of all barcodes Data information: Data were collected at month 7 post-transplantation for the uMT-/- group and month 8 post-transplantation for the NSG group, and presented as mean ± SEM. n=7 mice for the uMT-/- group and n=6 for the NSG group. *p<0.05 (two-tailed and two-sample equal variance Student"s t-test)


K - L Production of Gr , B , CD4 T , and CD8 T cells by expanding WT clones that produce more than 0 . 1 % of WBCs in each lineage . Data information : Data were collected at month 7 post - transplantation for the uMT - / - group and month 8 post - transplantation for the NSG group , and presented as mean ± SEM . n = 7 mice for the uMT - / - group and n = 6 for the NSG group . * p < 0 . 05 ( two - tailed and two - sample equal variance Student " s t - test )

O O O O O B-CELL_TYPE O B-CELL_TYPE O B-CELL_TYPE I-CELL_TYPE O O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O O O O O O O O O O O O O B-CELL_TYPE O O O O O O O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-ORGANISM O O B-GENEPROD O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O O O O O O O O O O O O O O

K-L Production of Gr, B, CD4 T, and CD8 T cells by expanding WT clones that produce more than 0.1% of WBCs in each lineage. Data information: Data were collected at month 7 post-transplantation for the uMT-/- group and month 8 post-transplantation for the NSG group, and presented as mean ± SEM. n=7 mice for the uMT-/- group and n=6 for the NSG group. *p<0.05 (two-tailed and two-sample equal variance Student"s t-test)


A - B Total amount of progenitors as a percentage of ckit and Il7rα enriched bone marrow ( BM ) cells . Shown are hematopoietic stem cells ( HSCs ) , Flk2 - and Flk2 + multipotent progenitors ( MPP ) , common lymphoid progenitors ( CLP ) , common myeloid progenitors ( CMP ) , megakaryocyte - erythroid progenitors ( MEP ) , and granulocyte - monocyte progenitors ( GMP ) Data information : Data were collected at month 7 post transplantation for the uMT - / - group and month 8 post - transplantation for the NSG group , and presented as mean ± SEM . n = 7 mice for the uMT - / - group and n = 6 for the NSG group . * p < 0 . 05 and * * p < 0 . 01 ( two - tailed and two - sample equal variance Student " s t - test )

O O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O O B-GENEPROD O B-GENEPROD O B-CELL_TYPE I-CELL_TYPE O B-CELL_TYPE O O O O O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O B-CELL_TYPE O O O O O O O B-CELL_TYPE I-CELL_TYPE O B-CELL_TYPE O O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O B-CELL_TYPE O O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O B-CELL_TYPE O O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O B-CELL_TYPE O O O B-CELL_TYPE I-CELL_TYPE I-CELL_TYPE I-CELL_TYPE O B-CELL_TYPE O O O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-ORGANISM O O B-GENEPROD O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

A-B Total amount of progenitors as a percentage of ckit and Il7rα enriched bone marrow (BM) cells. Shown are hematopoietic stem cells (HSCs), Flk2- and Flk2+ multipotent progenitors (MPP), common lymphoid progenitors (CLP), common myeloid progenitors (CMP), megakaryocyte-erythroid progenitors (MEP), and granulocyte-monocyte progenitors (GMP) Data information: Data were collected at month 7 post transplantation for the uMT-/- group and month 8 post-transplantation for the NSG group, and presented as mean ± SEM. n=7 mice for the uMT-/- group and n=6 for the NSG group. *p<0.05 and **p< 0.01 (two-tailed and two-sample equal variance Student"s t-test)


C - D WT donor derived progenitors as a percentage of ckit and Il7rα enriched BM cells . Data information : Data were collected at month 7 post transplantation for the uMT - / - group and month 8 post - transplantation for the NSG group , and presented as mean ± SEM . n = 7 mice for the uMT - / - group and n = 6 for the NSG group . * p < 0 . 05 and * * p < 0 . 01 ( two - tailed and two - sample equal variance Student " s t - test )

O O O O O O O O O O O B-GENEPROD O B-GENEPROD O B-CELL_TYPE I-CELL_TYPE O O O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O B-ORGANISM O O B-GENEPROD O O O O O O O O O O B-GENEPROD O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

C-D WT donor derived progenitors as a percentage of ckit and Il7rα enriched BM cells. Data information: Data were collected at month 7 post transplantation for the uMT-/- group and month 8 post-transplantation for the NSG group, and presented as mean ± SEM. n=7 mice for the uMT-/- group and n=6 for the NSG group. *p<0.05 and **p< 0.01 (two-tailed and two-sample equal variance Student"s t-test)


a , Immunoprecipitation of Flag - beclin 1 constructs with Nef - HA in HeLa cells 24 h post - transfection .

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a, Immunoprecipitation of Flag-beclin 1 constructs with Nef-HA in HeLa cells 24 h post-transfection.


b , GFP - LC3 - positive dots ( autophagosomes ) in MCF7 cells expressing GFP - LC3 and Flag - beclin 1 constructs grown in either normal medium or starved in EBSS for 2 h .

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b, GFP-LC3-positive dots (autophagosomes) in MCF7 cells expressing GFP-LC3 and Flag-beclin 1 constructs grown in either normal medium or starved in EBSS for 2 h.


d , Biochemical assessment of autophagy ( p62 and LC3 immunoblots ) in peptide - treated HeLa cells ( 3 h ) .

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d, Biochemical assessment of autophagy (p62 and LC3 immunoblots) in peptide-treated HeLa cells (3 h).


e , f , Representative images ( e ) and quantification of GFP - LC3 - positive dots ( f ) in peptide - treated HeLa / GFP - LC3 cells ( 30 µM , 3 h ) . Scale bars , 20 µm .

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e, f, Representative images (e) and quantification of GFP-LC3-positive dots (f) in peptide-treated HeLa/GFP-LC3 cells (30 µM, 3 h). Scale bars, 20 µm.


g , GFP - LC3 - positive dots in siRNA - transfected peptide - treated HeLa / GFP - LC3 cells ( 30 µM , 3 h ) .

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g, GFP-LC3-positive dots in siRNA-transfected peptide-treated HeLa/GFP-LC3 cells (30 µM, 3 h).


h , Model of Tat - beclin 1 peptide ( left ) based on corresponding elements of the beclin 1 evolutionarily conserved domain ( ECD ) structure ( centre ) . Essential phenylalanine side chains , magenta ; positions of solubility mutations , pink ; lipid interaction site , yellow . ECD surface representation ( right ) illustrates exposure of corresponding peptide ( cyan ) .

O O O O O O B-GENEPROD I-GENEPROD O O O O O O O O O O B-GENEPROD I-GENEPROD O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

h, Model of Tat-beclin 1 peptide (left) based on corresponding elements of the beclin 1 evolutionarily conserved domain (ECD) structure (centre). Essential phenylalanine side chains, magenta; positions of solubility mutations, pink; lipid interaction site, yellow. ECD surface representation (right) illustrates exposure of corresponding peptide (cyan).


i , p62 and LC3 immunoblots in peptide - treated HeLa cells ( 3 h ) . In b , f , g , bars represent mean ± s . e . m . of triplicate samples ( 50 - 100 cells per sample ) . Similar results were observed in three independent experiments . * P 0 . 05 , * * P 0 . 01 ; t - test .

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i, p62 and LC3 immunoblots in peptide-treated HeLa cells (3 h). In b, f, g, bars represent mean ± s.e.m. of triplicate samples (50-100 cells per sample). Similar results were observed in three independent experiments. *P 0.05, **P 0.01; t-test.


a , HeLa cells were treated with biotin - conjugated peptides ( 30 μM , 3 h ) and proteins bound to peptides were analysed by immunoblot with anti - GAPR - 1 . b - T - B , biotin - Tat - beclin 1 ; b - T - S , biotin - Tat - scrambled .

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a, HeLa cells were treated with biotin-conjugated peptides (30 μM, 3 h) and proteins bound to peptides were analysed by immunoblot with anti-GAPR-1. b-T-B, biotin-Tat-beclin 1; b-T-S, biotin-Tat-scrambled.


b , Immunoprecipitation of Flag - beclin 1 with GAPR - 1 - Myc in HeLa cells 24 h post - transfection .

O O O O B-GENEPROD O B-GENEPROD I-GENEPROD O B-GENEPROD I-GENEPROD I-GENEPROD O B-GENEPROD O B-CELL_LINE O O O O O O O

b, Immunoprecipitation of Flag-beclin 1 with GAPR-1-Myc in HeLa cells 24 h post-transfection.


c , GFP - LC3 - positive dots in GAPR - 1 siRNA - transfected peptide - treated HeLa / GFP - LC3 cells ( 20 & amp ; amp ; mgr ; M , 3 h ) with or without 100 nM bafilomycin A1 . Bars represent mean ± s . e . m . of triplicate samples ( 50 - 100 cells per sample ) . Similar results were observed in three independent experiments . NC , non - silencing control .

O O B-GENEPROD O B-GENEPROD O O O O B-GENEPROD I-GENEPROD I-GENEPROD O O O O O O B-CELL_LINE O B-GENEPROD O B-GENEPROD O O O O O O O O O O O O O O O O O O O O B-SMALL_MOLECULE I-SMALL_MOLECULE O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

c, GFP-LC3-positive dots in GAPR-1 siRNA-transfected peptide-treated HeLa/GFP-LC3 cells (20 &amp;mgr;M, 3 h) with or without 100 nM bafilomycin A1. Bars represent mean ± s.e.m. of triplicate samples (50-100 cells per sample). Similar results were observed in three independent experiments. NC, non-silencing control.


d , Localization of beclin 1 and GM130 ( a Golgi marker ) in HeLa cells stably transduced with empty vector ( left panel ) or GAPR - 1 - Myc ( middle panel ) or transfected with GAPR - 1 siRNA ( right panel ) and treated with peptide ( 20 μM , 1 h ) . Scale bar , 20 μm .

O O O O B-GENEPROD I-GENEPROD O B-GENEPROD O O B-SUBCELLULAR O O O B-CELL_LINE O O O O O O O O O O O B-GENEPROD I-GENEPROD I-GENEPROD O B-GENEPROD O O O O O O O B-GENEPROD I-GENEPROD I-GENEPROD O O O O O O O O O O O O O O O O O O O O O O O

d, Localization of beclin 1 and GM130 (a Golgi marker) in HeLa cells stably transduced with empty vector (left panel) or GAPR-1-Myc (middle panel) or transfected with GAPR-1 siRNA (right panel) and treated with peptide (20 μM, 1 h). Scale bar, 20 μm.


e , Immunoblot of beclin 1 in post - nuclear supernatant ( PNS ) and Golgi - enriched fractions in HeLa cells stably transduced with empty vector or GAPR - 1 - Myc after peptide treatment ( 20 μM , 2 h ) .

O O B-EXP_ASSAY O B-GENEPROD I-GENEPROD O O O O O O O O O B-SUBCELLULAR O O O O B-CELL_LINE O O O O O O O B-GENEPROD I-GENEPROD I-GENEPROD O B-GENEPROD O O O O O O O O O O O

e, Immunoblot of beclin 1 in post-nuclear supernatant (PNS) and Golgi-enriched fractions in HeLa cells stably transduced with empty vector or GAPR-1-Myc after peptide treatment (20 μM, 2 h).


f , WIPI2 dots in cells in the experimental conditions shown in d . Bars represent mean ± s . e . m . for 100 - 150 cells . * P 0 . 05 , * * * P 0 . 001 ; t - test . † † † P 0 . 001 ; two - way ANOVA for comparison of magnitude of changes between groups .

O O B-GENEPROD O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

f, WIPI2 dots in cells in the experimental conditions shown in d. Bars represent mean ± s.e.m. for 100-150 cells. *P 0.05, ***P 0.001; t-test. †††P 0.001; two-way ANOVA for comparison of magnitude of changes between groups.


a , Percentage of cells with small htt103Q aggregates ( left ) and number of aggregates per cell ( right ) in HeLa cells expressing doxycycline ( Dox ) - repressible CFP - htt103Q after daily treatment with doxycycline or peptide ( 20 μM , 4 h per day ) for 2 days . Bars represent mean ± s . e . m . of triplicate samples ( 60 - 120 cells per sample ) . Similar results were observed in three independent experiments .

O O O O O O O B-GENEPROD O O O O O O O O O O O O O O B-CELL_LINE O O B-SMALL_MOLECULE O O O O O B-GENEPROD O B-GENEPROD O O O O B-SMALL_MOLECULE O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

a, Percentage of cells with small htt103Q aggregates (left) and number of aggregates per cell (right) in HeLa cells expressing doxycycline (Dox)-repressible CFP-htt103Q after daily treatment with doxycycline or peptide (20 μM, 4 h per day) for 2 days. Bars represent mean ± s.e.m. of triplicate samples (60-120 cells per sample). Similar results were observed in three independent experiments.


b , Filter trap assays for htt103Q large and small aggregates in HeLa / htt103Q cells .

O O B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O B-GENEPROD O O O O O B-CELL_LINE O B-GENEPROD O O

b, Filter trap assays for htt103Q large and small aggregates in HeLa/htt103Q cells.


c , Viral titres in HeLa cells infected with 0 . 1 plaque - forming units ( p . f . u . ) per cell of SINV , CHIKV or WNV ( strain TX02 ) and treated with peptide ( 10 μM , 4 - 8 h post - infection ) . Values represent geometric mean ± s . e . m . for triplicate samples of supernatants collected 18 h post - infection ( SINV ) or 24 h post - infection ( CHIKV and WNV ) . Similar results were observed in three independent experiments .

O O O O O B-CELL_LINE O O O O O O O O O O O O O O O O O O O O O B-ORGANISM O B-ORGANISM O B-ORGANISM O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O B-ORGANISM O O O O O O O O B-ORGANISM O B-ORGANISM O O O O O O O O O O O

c, Viral titres in HeLa cells infected with 0.1 plaque-forming units (p.f.u.) per cell of SINV, CHIKV or WNV (strain TX02) and treated with peptide (10 μM, 4-8 h post-infection). Values represent geometric mean ± s.e.m. for triplicate samples of supernatants collected 18 h post-infection (SINV) or 24 h post-infection (CHIKV and WNV). Similar results were observed in three independent experiments.


d , Bacteriacolony - forming units ( c . f . u . ) in primary BMDMs infected with L . monocytogenes ΔactA mutant strain DPL - 4029 for 30 min and treated with peptide ( 10 µM from 0 to 2 h post - infection ) . Bars represent mean ± s . e . m . of triplicate samples . Similar results were observed in three independent experiments .

O O B-ORGANISM B-EXP_ASSAY I-EXP_ASSAY I-EXP_ASSAY O O O O O O O O O O B-CELL_TYPE O O B-ORGANISM I-ORGANISM I-ORGANISM O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O O

d, Bacteriacolony-forming units (c.f.u.) in primary BMDMs infected with L. monocytogenes ΔactA mutant strain DPL-4029 for 30 min and treated with peptide (10 µM from 0 to 2 h post-infection). Bars represent mean ± s.e.m. of triplicate samples. Similar results were observed in three independent experiments.