The TRAF3-DYRK1A-RAD54L2 complex maintains ACE2 expression to promote SARS-CoV-2 infection

Angiotensin converting enzyme 2 (ACE2), the host receptor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, is differentially expressed in a wide variety of tissues and cell types. The expression of ACE2 is under tight regulation, but the mechanisms regulating ACE2 expression have not yet been well defined. Through a genome-wide CRISPR knockout screen, we discovered that host factors TRAF3, DYRK1A, and RAD54L2 (TDR) form a complex to regulate the expression of ACE2. Knockout of TRAF3, DYRK1A, or RAD54L2 reduces the mRNA levels of ACE2 and inhibits the cellular entry of SARS-CoV-2. On the other hand, SARS-CoV-2 continuously evolves by genetic mutations for the adaption to the host. We have identified mutations in spike (S) (P1079T) and nucleocapsid (N) (S194L) that enhance the replication of SARS-CoV-2 in cells that express ACE2 at a low level. Our results have revealed the mechanisms for the transcriptional regulation of ACE2 and the adaption of SARS-CoV-2.IMPORTANCE The expression of ACE2 is essential for the entry of SARS-CoV-2 into host cells. We identify a new complex-the TDR complex-that acts to maintain the abundance of ACE2 in host cells. The identification and characterization of the TDR complex provide new targets for the development of therapeutics against SARS-CoV-2 infection. By analysis of SARS-CoV-2 virus replicating in cells expressing low levels of ACE2, we identified mutations in spike (P1079T) and nucleocapsid (S194L) that overcome the restriction of limited ACE2. Functional analysis of these key amino acids in S and N extends our knowledge of the impact of SARS-CoV-2 variants on virus infection and transmission. The expression of ACE2 is essential for the entry of SARS-CoV-2 into host cells. We identify a new complex-the TDR complex-that acts to maintain the abundance of ACE2 in host cells. The identification and characterization of the TDR complex provide new targets for the development of therapeutics against SARS-CoV-2 infection. By analysis of SARS-CoV-2 virus replicating in cells expressing low levels of ACE2, we identified mutations in spike (P1079T) and nucleocapsid (S194L) that overcome the restriction of limited ACE2. Functional analysis of these key amino acids in S and N extends our knowledge of the impact of SARS-CoV-2 variants on virus infection and transmission.