Baumgarten, N. E. | Search bioRxiv

bioRxiv2026

A knockout screening platform for interferon-stimulated genes in respiratory cells identifies LY6E as a dominant antiviral effector against coronaviruses

Human coronaviruses (HCoVs) cause pathogenic outcomes ranging from mild illness to severe respiratory disease. Determining how respiratory cells coordinate an early antiviral response is necessary to understand successful control of HCoV replication. The Type I Interferon (IFN) response is a major component of innate viral immunity. IFN signaling leads to the upregulation of hundreds of interferon-stimulated genes (ISGs) that can have antiviral activity, though the individual ISGs that are responsible for restricting mildly pathogenic HCoVs in the respiratory epithelium are not clear. Here, we developed a targeted CRISPR-Cas9 knockout sgRNA library (termed the respISG library), focusing on ISGs that are upregulated across a panel of respiratory cells, to screen for innate host factors that restrict the mildly pathogenic human coronavirus, HCoV-OC43. We executed cell death-based screens in immortalized human small airway epithelial cells and identified lymphocyte antigen 6E (LY6E) as a top hit. We confirmed LY6E activity in validation studies and found it regulates entry of various SARS-CoV-2 Spike-pseudotyped lentiviruses. In addition to demonstrating a key role for LY6E in blocking a variety of HCoVs in small airway epithelial cells, this study establishes a loss-of-function CRISPR-Cas9 screening platform for identification of HCoV antiviral ISGs in respiratory cells.

microbiology↗

bioRxiv2023

Host cell glycosylation selects for infection with CCR5- versus CXCR4-tropic HIV-1

HIV-1 infection involves a selection bottleneck that leads to transmission of one or a few HIV variants, which nearly always use CCR5 as the coreceptor (R5 viruses) for viral entry as opposed to CXCR4 (X4 viruses). The host properties that drive this selection are not well understood and may hold keys to factors that govern HIV susceptibility. In this report, we identified SLC35A2, a transporter of UDP-galactose, as a candidate X4-specific restriction factor in CRISPR-knockout screens in primary target CD4+ T cells. SLC35A2 inactivation in CD4+ T cells, which resulted in truncation of glycans due to the absence of galactose, not only increased X4 infection levels, but also consistently decreased infection levels of R5 HIV strains. Single cycle infections demonstrated that the effect is host cell dependent. SLC35A2 is expressed in CD4+ T cells at different tissue sites, with high levels in the genital tract - the site of most HIV infections. These data support a role for a host cell protein that regulates glycan structure on HIV infection, with enhanced R5 infection but reduced X4 infection associated with SLC35A2-mediated glycosylation. Host cell glycosylation may therefore contribute to R5 selection and host susceptibility during HIV transmission.

microbiology↗