Conformational signals in the C-terminal domain of methionine adenosyltransferase I/III determine its nucleocytoplasmic distribution (original) (raw)

The methyl donor S-adenosylmethionine is synthesized in mammalian cytosol by three isoenzymes. Methionine adenosyltransferase II is ubiquitously expressed, whereas isoenzymes I (homotetramer) and III (homodimer) are considered the hepatic enzymes. In this work, we identified methionine adenosyltransferase I/III in most rat tissues both in the cytoplasm and the nucleus. Nuclear localization was the preferred distribution observed in extrahepatic tissues, where the protein colocalizes with nuclear matrix markers. A battery of mutants used in several cell lines to decipher the determinants involved in methionine adenosyltransferase subcellular localization demonstrated, by confocal microscopy and subcellular fractionation, the presence of two partially overlapping areas at the C-terminal end of the protein involved both in cytoplasmic retention and nuclear localization. Immunoprecipitation of coexpressed FLAG and EGFP fusions and gel filtration chromatography allowed detection of tetramers and monomers in nuclear fractions that also exhibited S-adenosylmethionine synthesis. Neither nuclear localization nor matrix binding required activity, as demonstrated with the inactive F251D mutant. Nuclear accumulation of the active enzyme only correlated with histone H3K27 trimethylation among the epigenetic modifications evaluated, therefore pointing to the necessity of methionine adenosyltransferase I/III to guarantee the supply of S-adenosylmethionine for specific methylations. However, nuclear monomers may exhibit additional roles. Key words: S-adenosylmethionine synthetase, epigenetic modifications, structural determinants of localization, tissular expression, nuclear methylations. Control of gene expression and synthesis of adrenaline or phospholipids such as phosphatidylcholine are some examples of processes involving S-adenosylmethionine (AdoMet)dependent methylations . AdoMet also participates in other important reactions such as those catalyzed by SAM radical proteins (i.e. biotin synthesis) or, upon decarboxylation, in spermidine and spermine synthesis (2, 3). In fact, all the radicals surrounding the AdoMet sulfur atom can be donated, as well as the carboxyl and amino groups of the methionine and the ribosyl moiety (4, 5). Thus, the number of processes involving AdoMet has been calculated to be as large as that of reactions using ATP. This small positively charged compound is synthesized from methionine and ATP in a singular reaction catalyzed by methionine adenosyltransferases (MATs)(6). These enzymes are present in the cytoplasm of eukaryotic cells , though reactions using this compound exhibit a wider subcellular distribution. Transfer of AdoMet to other cell compartments is therefore needed and carried out by specific transporters (7). Changes in AdoMet levels have been detected in several diseases, ranging from Parkinson (8) to alcohol liver cirrhosis (9). However, a direct correlation between AdoMet levels and pathology was only obtained by generating MAT1A knockout mice, which spontaneously develop hepatocellular carcinoma (HCC)(10).