Genome-wide loss-of-function screen using human pluripotent stem cells to study virus-host interactions for SARS-CoV-2

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019, has become a global health concern. Therefore, there is an immense need to understand the network of virus-host interactions by using human disease-relevant cells. We have thus conducted a loss-of-function genome-wide screen using haploid human embryonic stem cells (hESCs) to identify genes involved in SARS-CoV-2 infection. Although the undifferentiated hESCs are resistant to SARS-CoV-2, their differentiated definitive endoderm (DE) progenies, which express high levels of ACE2, are highly sensitive to the virus. Our genetic screening was able to identify the well-established entry receptor ACE2 as a host factor, along with additional potential novel modulators of SARS-CoV-2. Two such novel screen hits, the transcription factor MAFG and the transmembrane protein TMEM86A, were further validated as conferring resistance against SARS-CoV-2 by using CRISPR-mediated mutagenesis in hESCs, followed by differentiation of mutant lines into DE cells and infection by SARS-CoV-2. Our genome-wide genetic screening investigated SARS-CoV-2 host factors in non-cancerous human cells with endogenous ACE2 expression, providing a unique platform to identify novel modulators of SARS-CoV-2 cytopathology in human cells. [Display omitted] •HESCs are resistant to SARS-CoV-2, but definitive endoderm cells are highly infected•Establishment of an LOF screen to identify genes affecting viral infection•ACE2 and other membrane and nuclear proteins are suggested modulators of SARS-COV-2•Novel screen hits show pronounced decrease in SARS-CoV-2 load Benvenisty, Wolf, and colleagues report a loss-of-function genome-wide CRISPR-Cas9 knockout screen, using haploid human embryonic stem cells and their differentiated definitive endoderm progenies, in order to identify potential novel modulators of SARS-CoV-2. Their screen was able to identify potential novel membrane and nuclear proteins with pronounced effect on viral RNA load in the supernatant of the infected cells.