EMBO/sd-character-level-ner huggingface dataset:
Figure 2(A) UBQLN4 co-localizes with cytoplasmic aggregates after puromycin treatment. HeLa cells expressing Flag-tagged UBQLN4 were treated with 5 µg/mL puromycin for 2 h and then followed by 1% Triton X-100 extraction and 4% paraformaldehyde fixation. Fixed cells were stained with anti-polyubiquitin FK2 (shown in red to detect ubiquitin-positive cytoplasmic aggregates, ALIS) and anti-Flag antibodies (shown in green). (B, C) Puromycin-treated cells with UBQLN4 siRNA contain increased number of ALIS. At 72 h after transfection of the two distinct siRNA duplexes for UBQLN4 (UBQLN4 siRNA#1 and #2) or control siRNA (5 nM each), the cells were treated with 5 µg/mL puromycin for 2 h, and then subjected to immunostaining with an anti-polyubiquitin FK2 antibody (shown in green, B) as in (A). The number of ALIS was determined. The quantified data in (C) represent mean ± SEM. n = 76 cells for control siRNA, n = 89 cells for UBQLN4 siRNA#1, and n = 81 cells for UBQLN4 siRNA #2. P‐values were calculated by Welch's t-test between the siRNA‐transfected and the respective control condition (*P < 0.01). (B, C) Puromycin-treated cells with UBQLN4 siRNA contain increased number of ALIS. At 72 h after transfection of the two distinct siRNA duplexes for UBQLN4 (UBQLN4 siRNA#1 and #2) or control siRNA (5 nM each), the cells were treated with 5 µg/mL puromycin for 2 h, and then subjected to immunostaining with an anti-polyubiquitin FK2 antibody (shown in green, B) as in (A). The number of ALIS was determined. The quantified data in (C) represent mean ± SEM. n = 76 cells for control siRNA, n = 89 cells for UBQLN4 siRNA#1, and n = 81 cells for UBQLN4 siRNA #2. P‐values were calculated by Welch's t-test between the siRNA‐transfected and the respective control condition (*P < 0.01). (D) Depletion of UBQLN4 makes HeLa cells more sensitive to puromycin-induced cell death. After 48 h of UBQLN4 siRNA treatment, the cells were treated with 5 µg/mL puromycin for 15.5 h and then viability was measured. The data represent mean ± SD calculated from 3 independent experiments (n=3). P‐values were calculated by Student's t-test compared with puromycin-treated control siRNA cells (*P < 0.01).
a) Colocalization of GFP–LC3 and LAMP1.figf5(a) Colocalization of GFP-LC3 and LAMP1. HeLa cells expressing wild-type or mutant UVRAG were transfected with GFP-LC3 and incubated under normal conditions or treated with rapamycin (2 μM). Autophagosome fusion with LAMP+ structures was analysed by confocal microscopy (left panel; rapamycin-treated) and quantified (right upper panel; data are mean ± s.e.m., n = 100). Arrows and insets show the LAMP1+ autophagosomes. The number of GFP-LC3-positive dots per cell was counted using a fluorescence microscope (right bottom panel; data are mean ± s.e.m., n = 60, *P 0.05; **P 0.0001). (b) Vps16 or Vps18 knockdown inhibits UVRAG-mediated autophagosome maturation. HeLa.Vec (left) and HeLa.UVRAG (right) cells were transfected with GFP-LC3 together with control siRNA, Vps16 siRNA, or Vps18 siRNA, followed by confocal microscopy. Arrows in the top panel denote the autophagosomes with LAMP1 staining. Immunoblotting of Vps16, or Vps18 and tubulin are shown in the middle panel. The percentage of LAMP1+ autophagosomes was calculated (data are mean ± s.e.m., n = 50, *P 0.05). Scale bars, 5 μm. The raw data of the immunoblots are shown in Supplementary Information, Fig. S6.
Figure 1(A) Expression patterns of Hoxa5 and Hoxc8 in MNs (demarcated by white dashed lines) along the rostrocaudal axis of the spinal cord (cervical C4 to thoracic T1) from mouse embryos from E9.5 to E12.5. Pink scale bars: 12.5 μm for E9.5; 12.5 μm for E10.5; 25 μm for E11.5; and 50 μm for E12.5. Quantification of Hoxon cells across MN domains. Data represent mean ± SD from N ≥ 3 embryos. Summary of the intrasegmental expression profiles of Hoxa5 and Hoxc8 in cervical MNs. Yellow squares represent the Hoxa5onHoxc8 on cells.
Figure 5A. NLK does not induce nuclear translocation of the S128A mutant YAP. HEK293A cells were co-transfected with Myc-NLK and Flag-YAP WT or S128A mutant. Cells were serum starved for 6 hr, and YAP localization and NLK expression were determined by Flag (green) and Myc (red) antibodies, respectively. Scale bar: 20 μm. B. YAP Ser 128 phosphorylation is required for osmotic stress-induced YAP nuclear localization. HEK293A YAP KO cells reconstituted with Flag-YAP WT, YAP S128D, or YAP S128A mutant were treated with 0.2 M sorbitol for 1 hr. YAP subcellular localization was determined by Flag immunofluorescence staining (green). Scale bar: 20 μm.
Fig.Figure 3(A) Representation of the vectors used in this study. WTFlx Ndufs3 cDNA was expressed from rAAV9-Ndufs3. An rAAV9-eGFP expressing viral preparation was used as a control. (B) Experimental design used for rAAV9 administration and analyses. (C) The KO-GFP mouse was visibly smaller and weaker than a KO-NDUFS3 mouse, at 7 months. (D) Females body weight injected with rAAV9-Ndufs3 (heretofore referred to as KO-NDUFS3) mice were similar to WTFlx mice body weight and significantly higher than the rAAV9-eGFP injected mice (KO-GFP) starting at 5 months. Statistical analysis was performed by One-way ANOVA followed by Tukey post-test. P values were calculated using Log-rank (Mantel-Cox) Test; p=0.0016 (KO-NDUFS3 vs KO-GFP); p=0.0014 (KO-GFP vs WTFlx); n-indicates the number of animals used per group. (E) Males body weight of KO-NDUFS3 mice were similar to WTFlx mice, and significantly different from KO-GFP from 4 months onwards. Statistical analysis was performed by One-way ANOVA followed by a Tukey post-test. P values were calculated using Log-rank (Mantel-Cox) Test; p=0.0016 (KO-NDUFS3 vs KO-GFP); p=0.0014 (KO-GFP vs WTFlx); n-indicates the number of animals used per group. (F) Treadmill performance was significantly improved in the KO-NDUFS3 mice when compared to the KO-GFP, from 2 months onwards. P values were calculated using Log-rank (Mantel-Cox) Test; p=0.0016 (KO-NDUFS3 vs KO-GFP); p=0.0014 (KO-GFP vs WTFlx); n-indicates the number of animals used per group. (G) Four limb strength was evaluated by the grid hang test. KO-NDUFS3 mice performed significantly better starting at 3 months, when compared to the KO-GFP mice. F) and G) Statistical analysis was performed by One-way ANOVA followed by Bonferroni post-test. P values were calculated using Log-rank (Mantel-Cox) Test; p=0.0016 (KO-NDUFS3 vs KO-GFP); p=0.0014 (KO-GFP vs WTFlx); n-indicates the number of animals used per group. (H) Quadriceps and gastrocnemius wet weight of 6 months old mice was similar between WTFlx and KO-NDUFS3 mice. Quadriceps weight was significantly increased in KO-NDUFS3 mice when compared to KO-GFP injected. Statistical analysis was performed by One-way ANOVA followed by Bonferroni post-test. ns- not significant. P values were calculated using Log-rank (Mantel-Cox) Test; p=0.0016 (KO-NDUFS3 vs KO-GFP); p=0.0014 (KO-GFP vs WTFlx); n-indicates the number of animals used per group. (I) Survival curve of Ndufs3 smKO mice injected with rAVV9-Ndufs3 (blue line) and with rAVV9-eGFP (green line). WTFlx mice are depicted in black. rAAV9-Ndufs3 injections extended the survival of the smKO mice.
Figure 5A-C Immunoblot analysis the effects of MPK3/6 on salt stress-induced ARR1/10/12 protein degradation. Seven-day-old seedlings of 35S:ARR1:MYC, 35S:ARR10:YFP, and 35S:ARR12:YFP in Col-0 and MPK3SR backgrounds were treated with or without 200 mM NaCl for indicated time points. Actin was used as the internal reference. The relative intensity of band detected by anti-MYC or anti-GFP antibody to that by anti-Actin antibody without treatment was set to 1.0. D-I Quantification analysis of MPK3/6 effect on salt stress-induced ARR1/10/12 protein degradation in (A-C).
Figure 3(A) Relative mRNA levels of Yap1, Pou5f1, and Gata6 during time-course embryoid body (EB) differentiation. Gene expression data were obtained from GSE3749. Pou5f1 and Gata6 serve as representative ES cell marker and lineage-specific marker, respectively. (B) Relative Yap1 mRNA levels in ES cells (ESC) and differentiating ES cells (dESC) (LIF withdrawal for 4 days) and data are represented as mean SD. To differentiate ES cells, cells were incubated in LIF-withdrawn medium for 4 days. Both ESC and dESC were passaged every 2 days. (C) Protein levels of Yap1 and Pou5f1 during time-course differentiation upon LIF withdrawal. (D) Phospho-Yap1 levels during time-course differentiation. Samples were normalized by total Yap1 level. (E-G) Immunofluorescence (IF) images depicting localization of Yap1 in J1 (D), CJ7 (E), and E14 (F) mouse ESC (top) and dESC (bottom). The white arrow indicates nucleolus. Bottom panels represent higher magnification of the above panels. Dashed circle indicates nucleus border. (H) A schematic diagram depicting a Yap1 responsive luciferase reporter (8xGTIIC) construct.(I) Luciferase reporter assay using Yap1 responsive luciferase reporter (8xGTIIC) upon transient overexpression (OE) Yap1 in ES cells. P-values were calculated using Student's t-test. Two asterisks indicate P<0.01. 'Control' indicates ES cells infected with control virus not expressing any specific shRNA sequence and all RT-qPCR data are represented as mean SD (I-K).(J) Relative activity of Yap1 responsive luciferase reporter gene in ESC and dESC. P-values were calculated using Student's t-test. (K) Relative Yap1 mRNA levels in Control and Pou5f1 KD ES cells. (L) IF images depicting localization of Yap1 in Control and Pou5f1 KD ES cells. (M) Relative activity of Yap1 responsive luciferase reporter gene upon Pou5f1 KD in ES cells. P-values were calculated using Student's t-test.
Red1 zinc‐finger motif is required for meiotic RNA elimination.Figure 6(A) Red1 possesses a putative zinc‐finger domain. A BLAST search reveals homology between Red1 and a conserved domain termed pfam10650, a CCCH‐type zinc‐finger that has the CX8CX4C3H motif as a consensus sequence. The arrow indicates the conserved histidine residue, which is substituted for isoleucine. (B) The histidine637 to isoleucine mutation (H637I) does not affect the protein level of Red1. Protein extracts from Red1‐GFP and Red1H637I‐GFP strains were examined by western blotting using anti‐GFP or anti‐Cdc2 antibody. Cdc2 was probed as a loading control. (C) DIC images of vegetative cells expressing Red1‐GFP and Red1H637I‐GFP strains. Bars, 5 μm. (D) Fluorescent microscopy indicates that both Red1‐GFP and Red1H637I‐GFP form nuclear dots. Bars, 2 μm. (E) The H637I mutation leads to the accumulation of meiotic RNAs. Total RNAs isolated from Red1‐GFP, Red1H637I‐GFP, and red1Δ strains were subjected to RT-PCR using primers specific for four meiotic mRNAs, SPBPB2B2.03c, mcp5+, ssm4+, and rec8+. (F) RNA‐immunoprecipitation of Red1‐GFP and Red1H637I‐GFP. Cell lysates from untagged, Red1‐GFP, and Red1H637I‐GFP strains were subjected to immunoprecipitation using anti‐GFP, and precipitated RNAs were analysed by RT-PCR. The values of relative enrichment are shown.
Figure 4A-B The peritoneal recruitment of (A) CD45+/CD115-/Ly6G+ neutrophils and (B) CD45+/CD115+/Ly6Chi classical monocytes in C57BL/6J mice was assessed 4 hours after intraperitoneal (IP) injection of 500 nM CXCL12 alone (black) and in the presence of 50 nM Gal-3 (dark green; A-B: n = 7 mice). C The peritoneal recruitment of classical monocytes after IP injection of PBS (n = 6) or TG in C57BL/6J WT (dark green, n = 10 mice) and Gal-3-/- (black, n = 5 mice) mice was assessed after 18 h. Where indicated, the mice received an IP injection of CXCR4 antagonist AMD 3465 12 hours prior to the experiment. D The concentration of CXCL12 concentration was determined by ELISA on the peritoneal lavage normalized with levels from the mesenterium (n = 4 mice). E CXCR4 expression levels on Ly6Chi monocytes of the blood and the peritoneal lavage after 18 h of TG stimulation were determined by flow cytometry and indicated as mean fluorescence intensity (MFI) (n = 4 mice).
Figure 4Flow cytometric analyses of Liv2+ cells in the VPR, DPR and the liver buds (LB). Liv2+ cells from the VPR-L (brown arrow), Liv2- cells from the VPR-L (yellow arrow) and Liv2+ cells from the LB (black arrow) were sorted for single-cell RNA-seq. Immunofluorescence of PDX1 and Liv2 on sagittal section of E10.5 mouse embryo. The white and yellow arrows denote PDX1low/Liv2+ cells in in the liver and EHBD regions, respectively. Scale bar: 50 µm. PCA plot of single-cell transcriptomes from the indicated cells (left). Each dot represents a single-cell. All cells are classified into three groups (right). The two panels represent the same PCA coordinates. Hierarchical clustering of 694 genes correlated with the first two PCs identified three major distinct cell lineages (p < 1 × 10-6). Each column represents a single-cell and each row represents one gene. Cells were re-ordered according to the value of PC1 or PC2. Genes in clusters 'a, b' and 'c, d' were re-ordered by correlations with PC2 and PC1, respectively. The color of the cell type and cell source are indicated in (C). Lineage-specific TFs are listed on the right. The gene expression values of lineage marker genes were projected onto the PCA (E) and box (F) plots. Each point represents a cell. The y-axes of the box plots represent the expression levels (RPKM). Central line in the box represents median; box limits represent the first and third quartile; whiskers represent 1.5 × interquartile range. Unpaired Wilcoxon rank-sum test. **p-value <1 x 10-5; ***p-value <1 x 10-10. PCA plot of single-cell transcriptomes from the indicated cells. Each dot represents a single-cell. The PCA was performed using the same PC1 and PC2 in (C). Pdx1 and Sox17 expression levels in E9.5 VPR and DPR cells. Inferred ventral and dorsal pancreatic lineage relationships analyzed by minimum spanning tree.
Figure 2A Schematic representation of the four different F8 variants that were cloned into an AAV plasmid: wild-type F8; N6 containing 11 amino acids of the modified SQ amino acid linker (SQm) followed by the human N6 B domain; SQ containing the SQ amino acid linker; V3 containing the V3 peptide in the middle of the SQ linker. ITR, inverted terminal repeats; CMV, cytomegalovirus promoter; star symbol, 3xFlag tag; PolyA, short synthetic polyadenylation signal; Ntds, nucleotides; SP, signal peptide. Details on the exact amino acid differences in the B domain can be found in Fig EV1. B Western blot analysis of lysates of HEK293 cells 72 hours post-transfection with the various F8 variants. Neg, non-transfected cells. C Chromogenic assay of F8 activity in the medium of transfected cells are reported as International Units/deciliter (IU/dl). Data are presented as mean±SEM. Each dot within the same group corresponds to a biological replicate: Neg n=5; Wild-type n=8; N6 n=8; SQ n=7; V3 n=6. Significant differences between groups were assessed using The Kruskal-Wallis test followed by the posthoc analysis: Nemenyi's All-Pairs Rank Comparison Test. The Kruskal-Wallis test p value = 2.88e-05. **indicates the significant difference between the N6 and the Wild-type groups: p value ≤ 0.01.*indicates the significant difference between the V3 and the Wild-type groups: p value ≤ 0.05.
NO Reduced Phosphorylation of JNK1 and Bcl-2, Leading to Increased Bcl-2–Beclin 1 and Decreased Beclin 1–hVps34 Interactions.Figure 2(A) NO donors reduced JNK1 phosphorylation in HeLa cells, as analyzed by immunoprecipitation with anti-JNK1 agarose-conjugated beads and immunoblotting with anti-phospho-JNK antibody. (B) NO donors reduced Bcl-2 phosphorylation in HeLa cells, as analyzed by immunoblotting with anti-phospho-Bcl-2 antibody. C) Immunoblot analysis with anti-phospho-Bcl-2 antibody shows that DETA NONOate could not reduce phospho-Bcl-2 in Flag-CA JNK1-transfected HeLa cells, where phospho-Bcl-2 was higher compared to mock-transfected cells. (D and E) Immunoprecipitation with anti-Flag M2 affinity agarose gel and immunoblotting with anti-Myc antibody shows that NO donors increased the interaction of Flag-Beclin 1 with WT Myc-Bcl-2 (D), but not with AAA Myc-Bcl-2 (E), in HeLa cells transfected with WT Myc-Bcl-2 (D) or AAA Myc-Bcl-2 (E) along with either empty Flag or Flag-Beclin 1. Asterisk denotes IgG band (E). (F) Immunoprecipitation with anti-Flag M2 affinity agarose gel and immunoblotting with anti-Vps34 antibody shows that NO donors decreased Flag-Beclin 1-hVps34 interaction in HeLa cells transfected with hVps34 along with either empty Flag or Flag-Beclin 1. (G) Immunoprecipitation with anti-Flag M2 affinity agarose gel and immunoblotting with anti-Vps34 antibody shows that AAA Myc-Bcl-2 decreased Flag-Beclin 1-hVps34 interaction in the presence (right panel) or absence (left panel) of DETA NONOate in HeLa cells transfected with hVps34 along with either empty Flag or Flag-Beclin 1, and with pcDNA3.1 or AAA Myc-Bcl-2. Graphical data denote mean ± SEM.
DRP-1 is localized to the lumen of autophagic vesicles.Figure 10.DRP-1 is localized to the lumen of autophagic vesicles. Immunogold staining of 293 cells expressing HA-DRP-1 reveals a specific staining of DRP-1 in the lumen of double membrane autophagosomes containing cytoplasmic material (A and B) in autophagosomes containing organelles and vacuoles (C) and inside the body of autolysosomes containing digested matter (D). The gold particles do not overlap with the phagocytosed organelles in C or the lysosomes in D.
a, b, Analysis of HeLa cells stably expressing YFP fused to the indicated galectins and infected with S.f1a, b, Analysis of HeLa cells stably expressing YFP fused to the indicated galectins and infected with S. Typhimurium for 1 h. a, Percentage of bacteria coated by the indicated galectins. YFP-positive bacteria were counted by microscopy. Mean and standard deviation (s.d.) of triplicate HeLa cultures, n > 100 bacteria per coverslip. b, Confocal microscopy. Arrowheads, bacteria shown in insets. DAPI, 4′,6-diamidino-2-phenylindole. c, Kinetics of fold replication for S. Typhimurium in HeLa cells transfected with the indicated siRNAs. Bacteria were counted on the basis of their ability to form colonies on agar plates. Mean and s.d. of triplicate HeLa cultures and duplicate colony counts. siRNAs are further characterized in Supplementary Fig. 2a-c. **P  0.01, Student's t-test. Scale bar, 10 µm.
Figure 4A Representative neurons expressing myc-tagged gephyrin or G375D and pHluorin-tagged GABAAR γ2. Surface-expressed γ2-subunits were immunolabelled with GFP-antibodies in red, and the pHluorin-fluorescence (total γ2 levels) are shown in green.B Size and C fluorescence (fl.) intensity of surface-expressed γ2 clusters in myc-gephyrin and myc-G375D-expressing dendrites (n = 20 myc-gephyrin and 21 myc-G375D neurons in B and 18 myc-gephyrin and myc-G375D neurons in C)D Mean γ2-pHluorin fluorescence in myc-gephyrin and myc-G375D-expressing dendrites (n = 20 and 18 myc-gephyrin and myc-G375D expressing neurons).E Number of surface γ2 clusters on dendrites normalized to 20 µm segments (n = 18 myc-gephyrin and myc-G375D expressing neurons). At least three independent cultures were used for each experiment. Results are expressed as mean ± SEM; Data were analyzed by t-test. F Representative traces of miniature recordings on cultured hippocampal neurons transfected either with GFP-gephyrin or GFP-G375D.G mIPSC amplitudes from neurons expressing GFP-gephyrin or GFP-G375D. Numbers show analyzed cell-numbers. Data were analyzed by one-way ANOVA and post-hoc Tukey test.H Frequency of the mIPSCs from neurons expressing GFP-gephyrin or GFP-G375D. The horizontal line within the boxes indicates the median frequency for the represented condition (bottom/top boundary indicates the 25th /75th percentile). Top and bottom error bars indicate the 90th and 10th percentiles, respectively. Analysis by Kruskal-Wallis test followed by post-hoc Mann-Whitney test.
Peripheral RAB13 RNA translation is required for RAB13 protein activation but not steady state distribution or membrane association.Figure 6A. Widefield images of RAB13 immunofluorescence in MDA-MB-231 cells treated with control or RAB13 (191+230) PMOs and ratios of peripheral/perinuclear intensity. Scale bars: 10 μm. n=45-50 cells. Bars: mean ± s.e.m.. Similar results were observed in two additional independent experiments. B. Fluorescence images (projections of confocal slices throughout the cell height) of cells expressing GFP-RAB13 with the indicated UTRs. Note that in both cases the protein assumes indistinguishable distribution. Scale bars: 10 μm. C. Soluble/particulate fractionation of the indicated cell lines followed by Western blot to detect the indicated proteins. RhoGDI and TfRc serve as soluble and particulate markers, respectively. Graph shows quantitation from n=3 independent experiments. Bars: mean ± s.e.m.. Active RAB13 (RAB13-GTP) was pulled-down using MICAL-L1 RBD-GST from the indicated PMO-treated cells The amount of endogenous RAB13 was measured by quantitative Western and relative levels of active RAB13 are plotted. n=8 (D), n=4 (E). Bars: mean ± s.e.m.. E. Active RAB13 (RAB13-GTP) was pulled-down using MICAL-L1 RBD-GST from GFP-RAB13 expressing lines The amount of endogenous or exogenous RAB13 was measured by quantitative Western and relative levels of active RAB13 are plotted. n=8 (D), n=4 (E). Bars: mean ± s.e.m..
Measurements of SARS-CoV-2 and SARS-CoV S1 binding to ACE2.Figure 2.Serial diluted recombinant S1 proteins of SARS-CoV-2, SARS-CoV and MERS-CoV were coated on 96 well plates, incubated with the recombinant Fc-tagged ACE2 (ACE2-Fc) for binding evaluation. Recombinant S1 proteins of SARS-CoV-2 and SARS-CoV were incubated with 293T-ACE2 cells and subjected to FACS evaluation for binding.
Figure 6(A-J) Confocal z-projections of brains expressing MiMIC mediated Dpr12GFSTF (Dpr12-GFP) and DIP-δGFSTF (DIP-δ-GFP) fusion proteins of the indicated genotypes and time points. (A-D) Dpr12-GFP expression is diffuse in DIP-δT2A-Gal4 homozygotes mutant brains at L3 (A; n=20/20), 24hr APF (B; n=14/14), 48hr APF (C; n=28/28) and adult (D; n=26/26). (E-H) DIP-δ-GFP expression in dpr12∆50-81 homozygotes mutant brains remains localized to the distal part of the γ-lobe at L3 (E; n=16/16), 24hr APF (F; n=16/16), and 48hr APF (G; n=10/10) but cannot be identified in adult brains (H; n=16/16). (I-J) Dpr12-GFP expression in WT animals (I, n=8/8) or in those ectopically expressing DIP-δ in PAM-DANs that innervate the γ3 zone (J, n=14/14) driven by MB441B-Gal4 (PAM-DAN-γ3-Gal4). DIP-δ expression in PAM-DAN-γ3 resulted in Dpr12-GFP localization within the γ3 zone (arrow), in addition to its normal γ4/γ5 localization.
Figure 4The overall complex structures of hACE2 bound to the SARS-CoV-2 RBD, GD/1/2019 RBD and GX/P2V/2017 RBD. The binding between the RBDs and hACE2 is mainly composed of two patches of interactions, and patch 1 and patch 2 are indicated in black and red dashed boxes, respectively. The N-glycans are shown as spheres. hACE2, SARS-CoV-2 RBD, GD/1/2019 RBD and GX/P2V/2017 RBD are colored in cyan, orange, lightpink and palegreen, respectively. Detailed interaction of hACE2 with the SARS-CoV-2 RBD, GD/1/2019 RBD and GX/P2V/2017 RBD in patch 1 and patch 2. Residues involved in the interaction are labeled, H-bonds are shown as red dotted lines with a cutoff of 3.5 Å.