The multifunctional poly(A)-binding protein (PABP) 1 is subject to extensive dynamic post-translational modification, which molecular modelling suggests plays an important role in co-ordinating its activities

摘要

PABP1 [poly(A)-binding protein 1] is a central regulator of mRNA translation and stability and is required for miRNA (microRNA)-mediated regulation and nonsense-mediated decay. Numerous protein, as well as RNA, interactions underlie its multi-functional nature; however, it is unclear how its different activities are co-ordinated, since many partners interact via overlapping binding sites. In the present study, we show that human PABP1 is subject to elaborate post-translational modification, identifying 14 modifications located throughout the functional domains, all but one of which are conserved in mouse. Intriguingly, PABP1 contains glutamate and aspartate methylations, modifications of unknown function in eukaryotes, as well as lysine and arginine methylations, and lysine acetylations. The latter dramatically alter the pI of PABP1, an effect also observed during the cell cycle, suggesting that different biological processes/stimuli can regulate its modification status, although PABP1 also probably exists in differentially modified subpopulations within cells. Two lysine residues were differentially acetylated or methylated, revealing that PABP1 may be the first example of a cytoplasmic protein utilizing a ‘methylation/acetylation switch’. Modelling using available structures implicates these modifications in regulating interactions with individual PAM2 (PABP-interacting motif 2)-containing proteins, suggesting a direct link between PABP1 modification status and the formation of distinct mRNP (messenger ribonucleoprotein) complexes that regulate mRNA fate in the cytoplasm.Abbreviations: AdOX, adenosine dialdehyde; DTT, dithiothrietol; eEF, eukaryotic elongation factor; eIF, eukaryotic initiation factor; eRF, eukaryotic release factor; eRF3-N, N-terminal eRF3; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; G3BP, Ras GAP (GTPase-activating protein) SH3 (Src homology 3) domain-binding protein; HRP, horseradish peroxidase; MEF, mouse embryonic fibroblast; miRNA, microRNA; MLLE, methionine-leucine-leucine-glutamate motif-containing domain; MS/MS, tandem MS; PABC, poly(A)-binding protein 1 C-terminal domain; PABP, poly(A)-binding protein; PAIP, poly(A)-interacting protein; PAM, PABP-interacting motif; PAN, poly(A) nuclease; PCNA, proliferating cell nuclear antigen; PRMT, protein arginine N-methyltransferase; PTM, post-translational modification; RRM, RNA recognition motif; SG, stress granule; TD-NEM, transcription-dependent nuclear export motif; TOB, transducer of ERBB2; TSA, trichostatin A