The role of non-coding RNAs in epigenetic regulation of gene expression

尽管我们现在已经有了人类和其他几种生物的完整基因组序列，但我们仍然对基因实际上是如何控制的知之甚少。传统上，基因被定义为编码蛋白质的遗传单位，被认为是基因组中唯一有意义的部分。约98%的基因组DNA是非编码序列，通常被称为‘垃圾’DNA，包含无意义的‘间隔区’序列。基因被转录成信使核糖核酸分子，信使核糖核酸分子将遗传信息从基因传递到细胞质，在细胞质中遗传密码被翻译成具有酶或结构作用的功能蛋白质分子。我们通常认为蛋白质编码基因的转录是基因组的唯一目的，但事实上，最近的研究表明，基因转录只占基因组转录活动的很小一部分。事实上，绝大多数转录的基因组区域都没有编码潜力。许多这些非编码或基因间转录本非常不稳定或罕见，它们的功能正在研究中。其他非编码转录本稳定积累，似乎在调节基因组大范围内的基因表达方面发挥作用。例如，Xist RNA是从X染色体转录而来的。雌性细胞有两条X染色体，而雄性细胞有一条X染色体和一条Y染色体。女性细胞中X染色体基因数量的翻倍是一个潜在的问题，可能会导致女性细胞中数百种基因产物的数量失衡。然而，Xist RNA已经进化到了平分。Xist RNA似乎覆盖了雌性细胞中的一条X染色体，导致该X染色体上的几乎所有基因完全失活。关于Xist是如何实现这一壮举的，人们知之甚少。最近还发现了其他非编码RNA，它们似乎是功能分子。Air RNA是一种大的非编码RNA，似乎是沉默一小群印记基因所必需的。大多数印记基因都与生长控制有关，而且是不寻常的，因为与大多数基因不同，印记基因的表达取决于你从谁那里遗传来的。我们每个人的基因组中每个基因都有两个拷贝，一个来自我们的母亲，一个来自我们的父亲。正常情况下，两种基因都会表达，但印记基因要么表达，要么沉默，这取决于它们来自哪个亲本。我们的工作和其他人的工作表明，Air RNA通过在一组基因上传播并覆盖它们来使它们沉默，可能是以非常类似于Xist控制X染色体的方式。虽然Xist曾经被认为是一个奇怪的东西，但现在看来，其他功能的非编码RNA也可能以类似的方式工作。与Xist相比，Air在基因组的一个相当小的区域内工作，Xist覆盖了整个染色体的大片区域，这使得Air的功能更容易研究。我们将尝试确定Air与其控制的印记基因簇相互作用的位置。另一种似乎也具有功能并控制一组印记基因的非编码RNA是Kcnq1ot1 RNA。我们将对这种RNA和XIST进行类似的实验，以获得关于它们作用机制的证据。很有可能存在更多的功能RNA，它们在基因表达的调控中发挥着重要的作用，但仍未得到重视。这项提案中提到的三个RNA调节着它们之间的数百个基因。这些实验将为基因组的调控提供重要的见解，并对人类健康产生重大影响。

Although we now have the entire genomic sequence for humans and several other organisms we still know little about how genes are actually controlled. Traditionally genes are defined as the genetic units that encode proteins and have been considered to be the only meaningful parts of the genome. About 98% of the genomic DNA is non-coding sequence and has often been referred to as 'junk' DNA, containing meaningless 'spacer' sequences. Genes are transcribed into mRNA molecules, which transport genetic information from the gene to the cytoplasm of cells where the genetic code is translated into a functional protein molecule with enzymatic or structural roles. We normally think of transcription of protein-coding genes as the sole purpose of the genome, but in fact recent research shows that gene transcription makes up only a very small percentage of the transcriptional activity of the genome. In fact the vast majority of transcribed genomic regions do not have coding potential. Many of these non-coding or intergenic transcripts are highly unstable or rare and their function is being investigated. Other non-coding transcripts stably accumulate and appear to function in regulating gene expression over wide areas of the genome. For example the Xist RNA is transcribed from the X chromosome. Female cells have two X chromosomes whereas males have one X and one Y chromosome. This doubling in the amount of X chromosome genes in female cells is a potential problem that could lead to an imbalance in the amount of hundreds of gene products in female cells. However, the Xist RNA has evolved to even the score. The Xist RNA appears to coat one of the X chromosomes in female cells leading to the complete inactivation of nearly all genes on that X chromosome. Little is known about how Xist achieves this feat. More recently other non-coding RNAs have been discovered which appear to be functional molecules. The Air RNA is a large non-coding RNA that appears to be necessary to silence a small cluster of imprinted genes. Most imprinted genes are involved in growth control and are unusual because unlike most genes, expression of an imprinted gene is dependent on who you inherited it from. We all have two copies of every gene in our genome, one from our mother and one from our father. Normally both genes are expressed but imprinted genes are either expressed or silenced depending on which parent they came from. Our work and the work of others has suggested that the Air RNA silences a cluster of genes by spreading over them and coating them, possibly in a very similar way to Xist control of the X chromosome. Though Xist was once thought to be an oddity, it now appears that other functional non-coding RNAs may operate in a similar way. The fact that Air operates over a fairly small region of the genome compared to Xist which covers a vast area encompassing an entire chromosome, makes Air function more amenable to investigation. We will attempt to identify the sites of Air interaction with the imprinted gene cluster that it controls. Another non-coding RNA that also appears to be functional and control a cluster of imprinted genes is the Kcnq1ot1 RNA. We will perform similar experiments on this RNA and the Xist to obtain evidence on their mechanisms of action. It is highly likely that many more functional RNAs exist that play important but, still unappreciated roles in the regulation of gene expression. The three RNAs mentioned in this proposal regulate hundreds of genes between them. These experiments will provide important insights into the regulation of the genome with a significant impact on human health.