Autoacetylation regulates differentially the roles of ARD1 variants in tumorigenesis

Abstract

225 autoacetylation inhibited tumor angiogenesis by decreasing the stability of hypoxia-inducible factor-1α (HIF-1α). Autoacetylation stimulated the catalytic activity of mARD1 225 to acetylate Lys532 of the oxygen-dependent degradation (ODD) domain of HIF-1α, leading to the proteosomal degradation of HIF-1α. In contrast, autoacetylation of mARD1 235 and hARD1 235 contributed to cellular growth under normoxic conditions by increasing the expression of cyclin D1. Taken together, these data suggest that autoacetylation of ARD1 variants differentially regulates angiogenesis and cell proliferation in an isoform-specific manner.

The biological functions of Naa10 — From amino-terminal acetylation to human disease

“…All isoforms share an N-terminal N-Acetyltransferase superfamily domain. Transfection of mNaa10 225 into HeLa cells strongly decreased the HIF-1α protein and VEGF mRNA levels under hypoxia whereas both other variants had only minor effects (Kim et al, 2006; Seo et al, 2014). In immunoprecipitates isolated by an acetyl-lysine antibody from HeLa cells treated with the proteasome inhibitor MG132, HIF-1α could be detected when the cells were transfected with mNaa10 225 but was nearly undetectable when the cells were transfected with mNaa10 235 or hNaa10 235 (Kim et al, 2006).…”

“…In mouse, an isoform specific regulation has been described. Overexpression of mNaa10 235 increased Cyclin D1 levels as shown by RT-PCR and western blot whereas overexpression of wt mNaa10 225 or mutated mNaa10 235 that abrogate acetyl-CoA binding or autoacetylation (K82A/Y122F or K136R) had no effects (Seo et al, 2014). …”

“…Moreover, it is not clear whether Naa10 directly acetylates HIF-1α or if another acetyltransferase is involved that is stimulated by Naa10. The acetyltransferase activity of Naa10 seems to be important for its action in hypoxia since mutated variants of mNaa10 225 which are not able to undergo autoacetylation (K136R) or acetyl-CoA binding (K82A/Y122F) were unable to bind to HIF-1α and did not increase HIF-1α acetylation in HeLa cells as detected by acetyl-lysine antibodies (Seo et al, 2014). …”

“…The use of an enzymatic-dead hNaa10 mutant helped to differentiate between NAT-dependent and -independent functions. This R82A mutation is located in a R 82 -x-x-G 85 -x-A 87 consensus sequence, critical for the binding of acetyl-CoA, and has been shown to exhibit low acetyltransferase activity and inhibited autoacetylation of Naa10 (Asaumi et al, 2005; Seo et al, 2010, 2014). According to the widely accepted idea that the acetyltransferase activity of Naa10 is crucial for its function, disruption of the catalytic activity abrogated Naa10 function to regulate APP/Aβ40 endocytosis and secretion, mTOR signaling, NF-κB activation, cyclin D1 expression and HIF-1α stability (Asaumi et al, 2005; Kuo et al, 2010; Seo et al, 2010, 2014; Park et al, 2012).…”

“…Autoacetylation at an internal lysine K136 in human and mouse Naa10 has been shown to regulate its enzymatic activity (Seo et al, 2010, 2014). Particularly, in in vitro acetylation assays with recombinantly expressed and purified wt Naa10 and subsequent detection with anti-acetyl-lysine antibody revealed acetylation of mNaa10 225 , mNaa10 235 and hNaa10 235 (see below for isoforms), whereas Naa10 with K82A and/or Y122F mutations, that inhibit acetyl-CoA binding, or Naa10 K136 (a mutation of the putative acetyl-acceptor site) were not found to be acetylated (Seo et al, 2014). In strong contrast to this, LC/MS/MS analyses on human Naa10 expressed and purified from E. coli did not show acetylation at any of the internal 16 lysines after incubation with Acetyl-CoA (Murray-Rust et al, 2006).…”