Mechanisms of posttranscriptional regulation of SAM homeostasis

项目总结/摘要 S-腺苷甲硫氨酸（SAM）是几乎所有细胞甲基化反应的甲基供体。鉴于 由于甲基化在核酸、蛋白质和脂质活性中的重要性， 是所有细胞过程的基础。尽管如此，细胞维持SAM的机制 体内平衡仍然不完全清楚。最近的数据表明，在一个模型中，转录后 反馈回路调节SAM水平。该模型提出，细胞通过以下方式对细胞内SAM水平做出反应： 控制MAT 2A的内含子保留，MAT 2A是编码主要SAM合成酶的RNA。一个保守的发夹， 称为hp 1，在MAT 2A 3 ′ UTR中是应答中的关键顺式作用决定子， 表征的RNA甲基转移酶胃L16也是该途径的中心。在高SAM水平下， 胃L16甲基化MAT 2A hp 1，有利于MAT 2A转录物的内含子保留和核降解。 在SAM耗尽时，胃L16结合hp 1但保持结合，推测是由于在低浓度下酶周转不良。 Sam.当与RNA结合时，胃L16作为剪接增强子发挥作用，促进MAT 2A mRNA 生产，从而增加SAM合成酶和SAM水平。尽管有大量的经验支持这一点 模型，它只是开始定义的机制和生物学意义的胃L16和转录后 SAM稳态的调节。本提案的三个目标旨在进一步了解 控制SAM体内平衡和RNA加工。在目标1中，顺式和反式作用因子 为了阐明胃L16诱导剪接的机制，将定义剪接所需的蛋白质。在目标2中， 检验了SAM体内平衡和胃L16对MAT 2A调节之间的联系。最后，在目标3中， 描述了无偏筛选，其已经鉴定了细胞中SAM稳态的潜在新调节剂。 由于SAM是许多细胞功能的核心，因此SAM水平的精确控制的丧失具有潜在的严重性。 对人类健康的后果。这里描述的工作将定义细胞用于维持SAM的机制 体内平衡

PROJECT SUMMARY/ABSTRACT S-adenosylmethionine (SAM) is the methyl donor for nearly all cellular methylation reactions. Given the importance of methylation in the activities of nucleic acids, proteins, and lipids, proper control of SAM is fundamental to a wide array all of cellular processes. Nonetheless, the mechanisms cells use to maintain SAM homeostasis remains incompletely understood. Recent data suggest a model in which a posttranscriptional feedback loop regulates SAM levels. The model proposes that cells respond to intracellular SAM levels by controlling intron retention of MAT2A, the RNA that encodes the major SAM synthetase. A conserved hairpin, called hp1, in the MAT2A 3´ UTR is a key cis-acting determinant in the response and the recently characterized RNA methyltransferase METTL16 is also central to the pathway. Under high SAM levels, METTL16 methylates MAT2A hp1 favoring intron retention and nuclear degradation of the MAT2A transcript. Upon SAM depletion, METTL16 binds hp1 but stays bound, presumably due to poor enzymatic turnover in low SAM. When bound to the RNA, METTL16 functions as a splicing enhancer to promote MAT2A mRNA production thereby increasing SAM synthetase and SAM levels. Despite significant empirical support for this model, it only begins to define the mechanisms and biological significance of METTL16 and posttranscriptional regulation of SAM homeostasis. The three aims in this proposal seek to further understand the biological roles of METTL16 controlling SAM homeostasis and RNA processing. In Aim 1, the cis- and trans-acting factors required for splicing will be defined to elucidate the mechanism of METTL16-induced splicing. In Aim 2, the proposed links between SAM homeostasis and MAT2A regulation by METTL16 are tested. Finally, in Aim 3, an unbiased screen is described that has identified potential new regulators of the SAM homeostasis in cells. Because SAM is central to many cellular functions, loss of precise control of SAM levels has potentially serious consequences for human health. The work described here will define mechanisms cells use to maintain SAM homeostasis.