Fig. 1: LDLR is an important host factor for CCHFV infection.

a Screening of LDLR and LDLR-related proteins (LRPs) that are crucial for CCHFV infection. The HEK293T cells were edited with a control or sgRNAs targeting the genes encoding LDLR and LRPs (two sgRNAs for each gene). After puromycin selection, the cell pools were infected with CCHFV (MOI = 0.05) for 24 h before RT-qPCR was performed. Data were normalized to the relative mRNA level of CCHFV S in the control sgRNA-edited cells. b Surface expression of LDLR in different cell lines. The indicated cell lines were assessed by flow cytometry using the anti-LDLR mAb (R301). c CCHFV infectivity of different cell lines. The indicated cell lines were infected with CCHFV (MOI = 0.05) for 48 h. The CCHFV Gn-positive cells were examined by flow cytometry with a customized anti-Gn monoclonal antibody (7A11). d Overexpression of LDLR enhances CCHFV infection in DLD1 cells. Control and LDLR-overexpressing DLD1 cells were infected with CCHFV (MOI = 0.05) for 24 h (left panel) or 48 h (right panel). The levels of CCHFV S mRNA and NP protein were determined by RT-qPCR (left panel) and immunoblots (right panel), respectively. e Effects of LDLR-deficiency on CCHFV infection in SW13 cells. SW13 cells were edited with a control (gNC) or three individual sgRNAs targeting different regions of LDLR coding sequence (gLDLR). The control and LDLR sgRNA-edited SW13 cell pools were infected with CCHFV (MOI = 0.05). CCHFV NP expression (left panel, 48 hpi), mRNA level of CCHFV S segment (2^nd panel, 24 hpi), percentage of Gn-positive cells (3^rd panel, 48 hpi) and cell cytopathic effects (right panel, 72 hpi) was measured by immunoblots, RT-qPCR, flow cytometry and crystal violet staining, respectively. For bar graphs, data are normalized to that of the control gRNA-edited cells. f Effects of LDLR-deficiency on production of progeny viruses. SW13 cells were edited with a control (gNC) or three individual sgRNAs targeting different regions of LDLR coding sequence (gLDLR). The sgRNA-edited SW13 cell pools were then infected with CCHFV (MOI = 0.05) for 72 h. Titers of progeny viruses in the supernatants were measured by TCID₅₀ assay. Data are normalized to that of the control gRNA-edited cells. LOD, limit of detection. g Effects of LDLR-deficiency on CCHFV infection in various cells. Huh7, Vero E6 and Hepa1-6 cells were edited with a control gRNA or the indicated numbers of gRNAs targeting LDLR gene. Cells were infected with CCHFV (MOI = 0.05) for 24 h before RT-qPCR was performed. Data are normalized to the CCHFV S mRNA level in the control gRNA-edited cells. h CCHFV infectivity in LDLR-knockout SW13 and Huh7 cells. Single clones of LDLR-knockout SW13 and Huh7 were isolated and confirmed by immunoblots (left). The control (gNC) or LDLR-deficient clone (gLDLR-C1) were infected with CCHFV (MOI = 0.05) for 24 h before RT-qPCR analysis. Data are normalized to that of each control gRNA-edited cells. i CCHFV infectivity in Ldlr^−/− primary cells. Primary hepatocytes and lung fibroblasts (MLFs) prepared from WT and Ldlr^−/− mice were incubated with CCHFV (MOI = 0.05). The mRNA level of CCHFV S segment (top, 48 hpi) and the viral genomic copies in the supernatant (bottom, 72 hpi) were measured by RT-qPCR. j Effects of LDLR-deficiency on CCHFV, RVFV, EBIV and VSV infection. The control (gNC) or LDLR-deficient clone (gLDLR-C1) were inoculated with the indicated viruses for 24 h before RT-qPCR was performed. Data are represented as mean ± SD. **P < 0.01; ***P < 0.001; ****P < 0.0001.