REGULATION C. ELEGANS VITELLOGENIN GENES

规则 C. 线虫卵黄蛋白基因

• 批准号: 3278755
• 负责人: Thomas Blumenthal
• 金额: $ 23.84万
• 依托单位: TRUSTEES OF INDIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-09-01 至 1992-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3278755
• 关键词: Caenorhabditis elegans; RNA; antiserum; developmental genetics; egg proteins; endonuclease; fusion gene; gel electrophoresis; gene expression; genetic mapping; genetic promoter element; genetic regulation; genetic transcription; genetic translation; genetically modified animals; hormone regulation /control mechanism; immunodiffusion; immunoelectrophoresis; immunofluorescence technique; ion exchange chromatography; messenger RNA; molecular cloning; nucleic acid hybridization; nucleic acid sequence; regulatory gene; reporter genes; structural genes; temperature sensitive mutant; transcription factor; vitellin

The vitellogenin gene family from the nematode, Caenorhabditis elegans is being studied. The genes are expressed abundantly in the intestine of the adult hermaphrodite worm, but are not transcribed in the male, in any of the four larval stages, or inother hermaphrodite tissues. In C. elegans five closely-related genes and one distant member of the gene family encode two distinct classess of vitellogenins. The main objective of this research is to understand the nature of the developmental signals which control transcription of these genes and the detailed mechanisms by which the regulation is effected. By sequencing the promoter regions of the six vitellogenin genes plus those from a related nematode species, two highly conserved heptameric sequences which are likely to be involve in the regulation have been identified. Development of a nematode transformation system to test these hypotheses has just recently been accomplished. A vitellogenin gene promoter has been fused to the structural gene for E. coli beta-glucuronidase and several transgenic worm strains containing the fusion gene integrated in their genomes have been obtained. These lines will be tested for regulated expression. The regions of the promoter required for particular aspects of the regulation will be identified using deletions, linker-scanning mutagenesis, and synthesis of putative regulatory elements. The regulatory roles of the different heptameric element will also be tested in competition experiments: Transformants containing long tandem arrays of the elements will be tested for inappropriate expression. Sequences bound by proteins in crude extracts will be identified by footprinting assays. The proteins that bind to the heptamers will be purified and the genes that encode them will be cloned. Regulatory mutants will be selected and their effects of the binding proteins measured. Mutants that express the vitellogenins in males or that fail to express them in hermaphrodites will be analyzed. Sequence analysis has revealed the presence of potential stem- loop structures at the 5' ends of the vitellogenin mRNAs. Evolutionary evidence strongly suggests that such stems exist. Direct evidence for their existence will be obtained by use of single-and double-strand specific nuclease digestion. Possible functions for the stems will be investigated, and the importance of the stems in performance of the proposed functions studied in vivo using transgenic worms containing the stem-containing region fused to a reporter gene.

杆状线虫卵黄原基因家族