Acetylation of the influenza A virus polymerase subunit PA in the N‐terminal domain positively regulates its endonuclease activity

Abstract

The post-translational acetylation of lysine residues is found in many nonhistone proteins and is involved in a wide range of biological processes. Recently, we showed that the nucleoprotein of the influenza A virus is acetylated by histone acetyltransferases (HATs), a phenomenon that affects viral transcription. Here, we report that the PA subunit of influenza A virus RNA-dependent RNA polymerase is acetylated by the HATs, P300/ CREB-binding protein-associated factor (PCAF), and general control nonderepressible 5 (GCN5), resulting in accelerated endonuclease activity. Specifically, the full-length PA subunit expressed in cultured 293T cells was found to be strongly acetylated. Moreover, the partial recombinant protein of the PA N-terminal region containing the endonuclease domain was also acetylated by PCAF and GCN5 in vitro, which facilitated its endonuclease activity. Mass spectrometry analyses identified K19 as a candidate acetylation target in the PA N-terminal region. Notably, the substitution of the lysine residue at position 19 with glutamine, a mimic of the acetyl-lysine residue, enhanced its endonuclease activity in vitro; this point mutation also accelerated influenza A virus RNA-dependent RNA polymerase activity in the cell. Our findings suggest that PA acetylation is important for the regulation of the endonuclease and RNA polymerase activities of the influenza A virus.

Human N-Alpha-Acetyltransferase 60 Promotes Influenza A Virus Infection by Dampening the Interferon Alpha Signaling

“…Above acetyltransferases and deacetylases exert their proviral and antiviral function, respectively, during IAV infection through both viral and host effectors. The acetyltransferases, Gcn5 and PCAF are involved in the suppression of the expression of host type I IFN and ISGs such as STAT1, MX1 and viperin ( 15 ), p300/CBP are implicated in the regulation, both positive and negative, of the expression of multiple host proteins involved in various stages of IAV life cycle ( 29 ), whereas NAA20/25 and also Gcn5 and PCAF facilitate the positive regulation of the IAV polymerase and host shutoff activities through acetylation of viral proteins ( 16 , 31 , 32 ). In contrast, HDAC6 ( 14 ) and HDAC11 ( 13 ) are involved in the enhancement of the IAV-induced type I IFN production.…”

Molecular and functional characterization of an evolutionarily conserved CREB‐binding protein in the LymnaeaCNS

“…The partial recombinant protein of the LymCBP HAT domain (from 1240th arginine to 1641st lysine, GenBank accession number BAE02656.1) with His 6 -tag was synthesized in Escherichia coli and purified as previously described, with slight modifications. [36][37][38] LymCBP catalytic domain was amplified by RT-PCR with cDNA prepared from Lymnaea CNSs, PrimeSTAR® HS DNA Polymerase (Takara Clontech), and primers, whose nucleotide sequences were summarized in Table S1 (No. 23,24).…”

Protein Acetylation Going Viral: Implications in Antiviral Immunity and Viral Infection

“…Interestingly, viral polymerase activity increases after silencing PCAF and decreases after silencing GCN5, suggesting that the acetylation of K31 and K90 has opposite effects on viral replication [ 61 ]. Hatakeyama et al also verified that PCAF and GCN5 can acetylate K19 on the PA subunit of the IAV RNA-dependent RNA polymerase, enhancing its endonuclease and RNA-dependent RNA polymerase activity while also affecting viral transcription [ 64 ]. Ma et al reported that the acetylation of K77, K113, and K229 on NP and of K108 on NS1 of IAV enhances the activity of these proteins and induces IFN antagonism, which promotes viral replication and survival [ 62 , 63 ].…”