Large-scale expression analysis of natural antisense transcripts

描述(申请人提供)：最近的研究已经确定了大量的自然反义转录本，这些转录本是从注释良好的基因的基因组位置转录而来的，但来自相反的DNA链。目前还不清楚大多数反义转录本是有功能的，还是仅仅是转录噪音。我们假设，如果反义转录与某些生理功能有关，它们将在不同类型的组织中表现出不同水平的表达，并且这种模式将趋于进化保守。通过采用商业高密度寡核苷酸微阵列，我们开发了一种经济高效的方法，可以监测哺乳动物基因组中所有外显子基因的反义表达。基于这种方法，我们将对人类、小鼠和大鼠的各种正常组织中的反义表达进行系统的分析。再加上对同一样本中正义转录本的表达分析，这将在外显子水平上定义一个“双链”的表达谱。这些数据将用于比较分析，以确定是否存在组织依赖反义表达的保守模式。我们将比较正性反义转录子对的表达模式的相关性与置换对的表达模式的相关性。我们还将确定高表达的反义转录本对是否具有较小的DNA序列差异，以及具有较小DNA序列差异的反义转录本对是否倾向于(1)其在不同组织中的表达谱具有更高的相关性，(2)物种间表达水平的较小变化，以及(3)表达广度的较小变化，类似于在蛋白质编码基因中观察到的。这可以为反义转录的选择性净化压力提供实质性证据，也可以支持“中性漂移”模式。综合数据集还将用于识别新的反义转录本和组织特异性反义转录本，以供进一步研究。如此大规模的反义表达分析将关键地评估反义转录是否是一个高度调控的过程。 与公共卫生相关：本提案代表了一项尝试，即获得关于可能干扰普通蛋白质编码基因的特定类型的新基因表达的全面数据。我们将研究它们在人、小鼠和大鼠各种正常组织中的表达模式，以期深入了解它们潜在的生理功能。

DESCRIPTION (provided by applicant): Recent studies have identified a large number of natural antisense transcripts that are transcribed from the genomic loci of well annotated genes, but from the opposite DNA strands. It is still unclear whether the majority of antisense transcripts are functional or merely transcriptional noise. We hypothesized that if antisense transcripts are related to certain physiological functions, they will exhibit variable levels of expression in different types of tissues and the pattern would tend to be evolutionarily conserved. By adapting commercial high-density oligonucleotide microarrays, we developed a cost-efficient approach that can monitor antisense expression across all exonic loci in mammalian genomes. Based on this approach, we will perform systematic profiling of antisense expression in various normal tissues in human, mouse and rat. Coupled with expression analysis of sense transcripts in the same samples, this will define a "double stranded" expression profile at the exon level. The data will be used for comparative analyses to determine if there are conserved patterns of tissue-dependent antisense expression. We will compare the correlations of expression patterns in orthologous pairs of antisense transcripts with the correlation of the expression pattern of permutated pairs. We will also determine whether highly expressed pairs of orthologous antisense transcripts have smaller DNA sequence divergence and whether orthologous pairs of antisense transcripts with smaller DNA sequence divergence tend to have (1) higher correlation in their expression profiles across various tissues, (2) smaller changes in expression level between species, and (3) smaller change in breadth of expression, similar to what have been observed in protein-coding genes. This could either provide substantial evidence for a selective purifying pressure on antisense transcription or favor a "neutral drift" model. The comprehensive datasets will also be used to identify novel antisense transcripts and tissue-specific antisense transcripts for further investigation. Such large-scale analysis of antisense expression will critically evaluate whether antisense transcription is a highly regulated process. PUBLIC HEALTH RELEVANCE: The present proposal represents an attempt to obtain comprehensive data on the expression of a certain type of novel genes that could interfere with ordinary protein- coding genes. We will study their expression patterns in various normal tissues of human, mouse and rat so that we could gain insight into their potential physiological function.