Transcriptional regulation of ACE2 and the adaption of SARS-CoV-2

ABSTRACT Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of the coronavirus disease 2019 (COVID-19), has caused a global health crisis. Currently effective vaccines are available to prevent SARS- CoV-2 infection, but our options for the treatment of SARS-CoV-2 infection remain limited. To inform the development of innovative interventions designed to counteract SARS-CoV-2 infection, we need a better understanding of the molecular details of SARS-CoV-2 infection, especially the host dependency factors that are required for virus infection. We have performed a genome-wide CRISPR knockout screen and identified PCBD1, TRAF3, and RAD54L2 as novel host factors required for SARS-CoV-2 infection. Our preliminary data showed that PCBD1, TRAF3, and RAD54L2 are required for the transcription of angiotensin converting enzyme 2 (ACE2). ACE2 is the primary receptor for the cellular entry of SARS-CoV-2, and plays key roles in virus infection and pathogenesis. ACE2 is differentially expressed in a wide variety of human tissues. However, the molecular basis for the transcriptional regulation of ACE2 in different tissues remains unexplored, which constitutes a clear knowledge gap in SARS-CoV-2 research. In Aim 1, we will determine the function of PCBD1, TRAF3, and RAD54L2 in transcriptional regulation of ACE2 in four cell lines derived from different tissues: Vero E6 (monkey kidney), Calu-3 (human lung), C2BBe1 (human large intestine), and HK-2 (human kidney). SARS-CoV-2 continuously evolves by genetic mutations for the adaption to the host. We identified a number of mutations in Spike (S) and Nucleocapsid (N) that may enhance the replication of SARS-CoV-2 in cells expressed low levels of ACE2. Some mutation (such as N S194L) was prevalent in India and Mexico during early circulation, suggesting these variants may contribute to the viral adaption to the host. In Aim 2, we will analyze the function of Spike (N81S, L242P, E484D, and P1079T) and Nucleocapsid (S194L) variants in virion production and cell entry of SARS-CoV-2. Our proposed studies will provide new information regarding the transcriptional regulation of ACE2 and the adaption of SARS-CoV-2. In the long term, these studies will provide new targets and strategies for the development of antiviral drugs.