The protein Nα-terminal acetyltransferase hNaa10p (hArd1) is phosphorylated in HEK293 cells

Using 2D-gel electrophoresis and hNaa10p specific antibodies, we have investigated whether hNaa10p is phosphorylated in HEK293 cells. Several differently charged forms of hNaa10p are present in HEK293 cells and treatment with Calf Intestine Alkaline Phophatase (CIAP) strongly suggests that hNaa10p is phosphorylated at multiple sites under various cell culture conditions. A direct or indirect role of GSK-3 kinase in regulating hNaa10p phosphorylation is supported by the observed effects of Wortmannin and LiCl, a GSK-3 activator and inhibitor, respectively.We demonstrate that hNaa10p protein is phosphorylated in cell culture potentially pointing at phosphorylation as a means of regulating the function of one of the major Nα-terminal acetyltransferases in human cells.Protein Nα-terminal acetylation is a very common protein modifications in eukaryotic cells, and approximately 80% of soluble human proteins is estimated to carry this modification. In humans, the hNatA complex is the major Nα-terminal acetyltransferase, acetylating Ser-, Thr-, Val-, Ala-, Gly- N-termini after the initial Met- has been removed by Methionine aminopeptidases [[1], Arnesen T and Van Damme P et al., submitted]. The recently described human NatB and NatC complexes are composed of distinct subunits and acetylate different subsets of Met-termini [[3,3], Starheim et al., submitted]. The hNatA complex composed of the catalytic subunit hNaa10p (hArd1) and the auxiliary subunit hNaa15p (hNat1) [4,5] is functionally conserved from yeast displaying almost identical substrate specificity (Arnesen T and Van Damme P et al., submitted). Naa10p, Naa11p, Naa15p and Naa16p represent the novel names of Ard1, Ard2, Nat1 and Nat2, respectively, and will be officially presented later this year when the nomenclature of this enzyme class is revised (Polevoda B, Arnesen T and Sherman F, unpublished). hNaa10p and/or hNaa15p has been demonstrated to be important for cell survival [6-8] suggesting important functions fo