MicroRNAs Regulating Gene Expression during Cellular Senescence and Aging

在衰老过程中，生物体的基因表达模式发生了变化，对压力和有丝分裂刺激的反应能力日益减弱。由于转录后过程关键地调节表达蛋白质集合的变化，限制性商业惯例(在其他项目中描述)和非编码RNA(特别是microRNAs和lncRNAs)的作用正在成为控制年龄相关基因表达模式的主要因素。为了研究ncRNA在衰老过程中的功能，我们采用了以下方法：ncRNA还原(通过转染反义分子)，ncRNA过表达(通过转染前体或成熟的ncRNA分子)，以及通过标记ncRNA(使用生物素或MS2标签)和通过各种方法(如微阵列、RT-PCR)识别与ncRNA相关的mRNAs。我们通过测量作为ncRNA丰度函数的目标mRNAs的稳态水平和半衰期，来研究ncRNAs是否影响靶mRNAs在衰老过程中的稳定性。我们研究了ncRNAs是否通过调节ncRNA水平影响目标mRNAs的翻译，随后研究了mRNAs与翻译多聚体的相对关联，并通过量化编码蛋白质的新生翻译率。我们还使用报告结构来进一步了解由ncRNAs调控的过程，并使用不同的衰老相关标记来检查衰老表型的变化。 在过去的资助期间，我们已经报告了与衰老相关的microRNA miR-519在自噬中发挥核心作用(Abdelmohsen等人，Mol.Cell Biol，2012)，miR-130通过降低PPAR的产生来抑制脂肪生成(Lee等人，Mol.细胞生物学，2011)，miR-146抑制脑转移(Hwang等人，分子和细胞，2012)。我们还证明了衰老上调的lncRNA(lincRNA-p21)的表达被microRNA let-7抑制(Yoon等人，Mol.细胞2012)，而microRNAs生产中的一个通用因子(DROSHA)被衰老下调蛋白AUF1抑制(Abdelmohsen等人，2012年)。我们在几篇文章(Srikantan等人，Cell Cycle，2011；Abdelmohsen和Gorospe，2011；Srikantan等人，Cell Cycle，2011)以及一本书的一章(Grammatikakis和Gorospe，in microRNAs in Medicine，2012)中综述了microRNAs在衰老和衰老中的作用。

During aging, organisms show altered gene expression patterns and have an increasingly impaired ability to respond to stress-causing and mitogenic stimuli. Since post-transcriptional processes critically regulate changes in the collections of expressed proteins, the role of RBPs (described in other projects) and noncoding RNAs (particularly microRNAs and lncRNAs) are emerging as major factors controlling age-related gene expression patterns. To investigate ncRNA function during senescence, we employ approaches such as ncRNA reduction (by transfecting an antisense molecules), ncRNA overexpression (by transfecting precursors or mature ncRNA molecules), and identification of ncRNA-associated mRNAs by tagging the ncRNAs (using biotin or MS2 tags) and identifying target mRNAs through various methods (eg, microarray, RT-PCR). We investigate whether ncRNAs affect the stability of target mRNAs during senescence by measuring the steady-state levels and half-lives of the mRNAs of interest as a function of ncRNA abundance. We investigate whether ncRNAs affect the translation of target mRNAs by modulating ncRNA levels, and subsequently studying the relative association of the mRNA with translating polysomes and by quantifying the nascent translation rates of the encoded proteins. We also employ reporter constructs to gain additional insight into the processes modulated by ncRNAs and use different senescence-associated markers to examine changes in the senescence phenotype. During the past funding period, we have reported that the senescence-associated microRNA miR-519 plays a central role in autophagy (Abdelmohsen et al., Mol. Cell Biol, 2012), that miR-130 suppresses adipogenesis by lowering PPAR production (Lee et al., Mol. Cell Biol, 2011), and that miR-146 inhibits brain metastases (Hwang et al., Molecules and Cells, 2012). We have also shown that expression of a senescence-upregulated lncRNA (lincRNA-p21) is inhibited by the microRNA let-7 (Yoon et al., Mol. Cell 2012) and that a general factor in the production of microRNAs (Drosha) is inhibited by the senescence-downregulated protein AUF1 (Abdelmohsen et al., 2012). We reviewed the contributions of microRNAs in senescence and aging in several articles (Srikantan et al., Cell Cycle, 2011; Abdelmohsen and Gorospe, 2011; Srikantan et al., Cell Cycle, 2011) as well as in one book chapter (Grammatikakis and Gorospe, in MicroRNAs in Medicine 2012).