Post-transcriptional Control of Immunoglobulin Expression

在B细胞发育过程中，免疫球蛋白（Ig）重链基因的转录终止和RNA加工将被检查。这项研究的长期目标是了解控制这些事件的一般机制，并确定B细胞成熟过程中发生的发育调节变化的基础。将导入已建立的B细胞和浆细胞瘤细胞系的正常和改变的Ig u-o基因的转录终止，将使用标准的核运行试验或拟议的基于pcr的试验进行分析。通过删除聚(a)位点、改变聚(a)位点强度和突变终止区序列，分析功能性聚(a)位点和下游终止区对u-o基因终止的贡献。为了确定依赖于两种相互竞争的RNA加工反应的效率的us和um RNA的调节产生是u基因特异性事件还是由于加工效率的全局变化，可以通过剪接和聚腺苷酸化替代加工的非ig基因将在B细胞和浆细胞瘤细胞中表达。u基因将在非b细胞中表达，以识别不受调节的加工途径，并由此推断受调节的途径。这项研究的结果可能会对其他系统产生重大影响，其中交替处理的前mrna被调用，并增加我们对免疫球蛋白成熟转换过程的了解

Regulated transcriptional termination and RNA processing of the immunoglobulin (Ig) heavy chain gene during B cell development will be examined. The long-term goal of this research is to understand the general mechanisms governing these events as well as to determine the basis for the developmentally regulated changes that occur during B cell maturation. Transcriptional termination of normal and altered Ig u-o genes, introduced into established B and plasmacytoma cell lines, will be analyzed using the standard nuclear run-on assay or a proposed PCR-based assay. The contribution a functional poly (A) site and the downstream termination region make to termination within the u-o gene will be analyzed by deleting the poly (A) sites, altering the poly (A) site strength and mutating the termination region sequences. To determine whether the regulated production of us and um RNA, which is dependent on the efficiencies of two competing RNA processing reactions, is a u gene-specific event or is instead due to global changes in processing efficiencies, a non-Ig gene that can be alternatively processed by splicing and polyadenylation will be expressed in B and plasmacytoma cells. The u gene will be expressed in non-B cells to identify the non-regulated processing pathway and, by inference, the regulated pathway. The results from this study could have significant impact in other systems where alternatively processed pre-mRNAs are invoked as well as increasing our insight into the switching process in immunoglobin maturation.***//