Extended Data Fig. 1: PCDH1 is required for entry of New World hantaviruses into HAP1 and U2OS cells. | Nature

Extended Data Fig. 1: PCDH1 is required for entry of New World hantaviruses into HAP1 and U2OS cells.

From: Protocadherin-1 is essential for cell entry by New World hantaviruses

Extended Data Fig. 1

a, Genes enriched for gene-trap insertions in the rVSV-ANDV Gn/Gc-selected population versus unselected control cells. The size of each bubble reflects the number of independent gene-trap insertions observed. Candidate genes are associated with cholesterol metabolism (yellow), the endoplasmic reticulum membrane complex network (green), and transcription (blue). PCDH1 (red) is a singleton hit. b, Single-cell HAP1 clones deficient for PCDH1 were generated by CRISPR–Cas9 genome engineering. The sequences of PCDH1-knockout alleles in clones 1 and 22 are shown. The sgRNA target sequence is highlighted in red. i1, insertion of a single nucleotide in PCDH1. c, WT and HAP1 PCDH1-KO cell lines were exposed to rVSVs bearing the indicated viral glycoproteins at a MOI of 0.02 IU per cell. ‘+cDNA’ indicates complementation of PCDH1-KO cells with PCDH1. Cells were scored for infection at 20 hpi. One hundred per cent relative infection corresponds to 20–30% infected cells. Averages ± s.d. are shown: HAP1-WT, three experiments, n = 6; PCDH1-KO#1-cDNA, two experiments, n = 5; PCDH1-KO#1+cDNA, three experiments, n = 8; PCDH1-KO#22-cDNA, three experiments, n = 8, except for HTNV Gn/Gc, for which two experiments, n = 4; PCDH1-KO#1+cDNA, three experiments, n = 8; PCDH1-KO#22+cDNA, three experiments, n = 8. d, Single-cell U2OS clones deficient for PCDH1 were generated by CRISPR–Cas9 genome engineering. The sequences of PCDH1-knockout alleles in clone 5 are shown. The sgRNA target sequence is highlighted in red. i1, insertion of a single nucleotide in PCDH1. e, f, WT and U2OS PCDH1-KO cell lines were exposed to the indicated rVSVs at a MOI of 0.02 IU per cell. Cells were scored for infection at 20 hpi. eGFP-positive infected cells (pseudocoloured green) were detected by fluorescence microscopy (e) and enumerated by automated counting (f). One hundred per cent relative infection corresponds to 10–20% infected cells. Averages ± s.e.m. are shown, three experiments, n = 13. In c, f, wild type versus PCDH1-KO by two-way ANOVA with Tukey’s (c) or Sidak’s (f) tests: NS, P > 0.05; ****, P < 0.0001.