SLC3A2 promotes early-stage of Zika virus infection in human placental cells and astrocytes.

Zika virus (ZIKV) has been associated with neonatal microcephaly or atypical Guillain-Barré syndrome in adults since 2016. However, despite considerable progress in understanding the biology and pathogenesis of ZIKV infection, little is known about its entry factors. This study aimed to identify host-cell proteins essential for ZIKV entry enriched by labeled viral envelope particles. The identification of solute carrier family 3 member 2 (SLC3A2) (CD98 heavy chain) through this approach highlights its potential as a novel target for therapeutic intervention against ZIKV. Gene editing via CRISPR-Cas9 and classical virological experiments were performed to identify SLC3A2 functions in ZIKV entry and infection processes. Immunofluorescence, reverse transcription‑quantitative polymerase chain reaction, and western blotting technologies were further used to detect viral proteins and genomes. Besides, coimmunoprecipitation and protein/antibody blocking assay were also conducted to identify direct interactions between SLC3A2 and the ZIKV envelope protein. Several human membrane proteins were overexpressed in HEK293 T cells, but only SLC3A2 could significantly promote ZIKV entry into host cells by two- to three- fold compared with the control. During authentic ZIKV infection, the genetic ablation of SLC3A2 in SLC3A2-KO1 JEG-3 cells reduced the viral infection rate to 59.00% ± 5.10% of the wild-type level (100.00% ± 4.97%) (P < 0.001), but its overexpression increased the susceptibility of human placenta- and brain-derived cell lines to the virus. SLC3A2 overexpression increased attached ZIKV levels to 3.4-fold of the control (3.423 ± 0.715 vs. 1.000 ± 0.897, P < 0.001) and internalized ZIKV levels to 1.7-fold of the control (3.782 ± 0.512 vs. 2.293 ± 0.272, P = 0.013) due to direct interaction with the ZIKV envelope protein in U-251MG cells. Our data further showed the close association of SLC3A2 with the ZIKV envelope protein via its extracellular domain during authentic viral infection. Notably, SLC3A2 ectodomain decoys and blocking antibodies markedly reduced ZIKV infection rather than affecting influenza A virus infection. SLC3A2 promotes ZIKV infection in placenta- or brain-derived cells by directly interacting with the ZIKV envelope protein and improving virus entry. SLC3A2 may be implicated in early-phase ZIKV infection as its entry factor, thereby identifying a potential therapeutic target and a basis for formulating antiviral strategies against ZIKV.