TCR RNA Surveillance

超出所提供的空间。细胞的存活依赖于检测基因表达错误的质量控制机制。本提案的主题是无义介导的衰变（NMD），一种识别和降解具有过早终止（无义）密码子的异常mrna的RNA监视途径。t细胞受体（TCR）和免疫球蛋白（Ig）转录本是NMD的常见底物，因为它们通常获得无义密码子，这是淋巴细胞发育过程中发生的程序性基因重排事件的结果，以增加淋巴细胞上的受体库。其他NMD底物是从随机产生的突变gen_s（无义突变和移码突变）和生物合成错误（转录和RNA剪接过程中的错误）产生的异常mrna转录而来。矛盾的是，异常TCR和Ig转录本的下调发生在细胞的核部分，然而触发NMD（无义密码子）的主要信号直到最近才被细胞质中的翻译装置识别。进一步证明细胞核在这种下调反应中起作用的证据是，无义密码子下游的内含子作为触发NMD的第二个信号。TCR转录本似乎对第二种信号有不同的反应，因为它们与所有其他已知的转录本不同，在无义密码子和引发NMD所需的下游内含子之间的最小距离上。此外，TCR转录本在一些结合无义密码子下游触发第二信号的因子上有所不同。TCR在无义密码子上游也有独特的调控元件，与所有已知的非重排基因转录本相比，其下调幅度要大得多（超过20倍）。该应用程序的具体目的是：(1)表征和理解作用于无义密码子上游的新调控元件的机制，从而导致TCR转录本的强烈下调；(2)识别和表征作用于无义密码子下游的调控元件（外显子-外显子连接或内含子）的因子，以传递TCR NMD必需的第二个信号。(3)通过生成和检测NMD基因UPF1零突变的淋巴细胞系，评估NMD的生理意义。了解NMD的生理功能和机制是很重要的，因为这种RNA监测途径在多种人类遗传疾病中被触发，并可能保护个体免受由于无意义和移码突变而产生的潜在有害的截断蛋白的影响。网站性能 ======================================== 节结束 ===========================================

EXCEED THE SPACE PROVIDED. The survival of cells depends on quality-control mechanisms that detect mistakes in gene expression. The subject of this proposal is nonsense-mediated decay (NMD), an RNA surveillance pathway that recognizes and degrades aberrant mRNAs that possess premature termination (nonsense) codons. T-cell receptor (TCR) and immunoglobutin (Ig) transcripts are frequent substrates of NMD, as they commonly acquire nonsense codons as a result of the programmed gene rearrangement events that occur during lymphocyte development to increase the repertoire of receptors on lymphocytes. Other NMD substrates are aberrant mRNAs transcribed from randomly generated mutant gen_s (with nonsense mutations and frameshift mutations) and those generated as a result of biosynthetic errors (mistakes during transcription and RNA splicing). Paradoxically, the downregulation of aberrant TCR and Ig transcripts occurs in the nuclearfraction of cells, yet the primary signal that triggers NMD (a nonsense codon) was until recently only known to be recognized by the translation apparatus in the cytoplasm. Further evidence that the nucleus has a role in this downregulatory response is that an intron downstream of a nonsense codon serves as the second signal to trigger NMD. TCR transcripts appear to have or to respond differently to this second signal, as they differ from all other known transcripts in the minimum distance between the nonsense codon and the downstream intron required to elicit NMD. In addition, TCR transcripts differ in some of the factors that bind downstream of a nonsense codon to trigger this second signal. TCR also has unique regulatory elements upstream of the nonsense codon that elicit much more robust downregulation (more than 20-fold) than all known transcripts from non-rearranging genes. The Specific Aims of this application are (1) to characterize and understand the mechanism of the novel regulatory elements that act upstream of nonsense codons to cause robust downregulation of TCR transcripts, (2) to identify and characterize factors that act on the regulatory elements downstream of nonsense codons (exon-exon junctions or introns) to deliver the second signal essential for TCR NMD, and (3) to assess the physiological significance of NMD by generating and testing lymphocyte cell lines with a null mutation in the NMD gene UPF1. Understanding the physiological functions and mechanisms of NMD is important, as this RNA surveillance pathway is triggered in a variety of human genetic diseases and may protect individuals from the potentially deleterious truncated proteins generated as a result of nonsense and frameshift mutations. PERFORMANCE SITE ========================================Section End===========================================