Terminal Differentiation in C. elegans

线虫的终末分化

• 批准号: 9305208
• 负责人: Ann Rougvie
• 金额: $ 41.18万
• 依托单位: University of Minnesota-Twin Cities
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-07-15 至 1997-12-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9305208&HistoricalAwards=false
• 关键词: Terminal; Differentiation; C.; elegans

The proper form of a multicellular organism requires precise control of terminal differentiation events. This research will address how a single terminal differentiation event is executed during the development of a simple model organism, the nematode C. elegans. The analysis will focus on the terminal differentiation of lateral hypodermal "seam" cells, which terminally differentiate during the final (4th) molt. Several distinct cellular processes are coordinately controlled during this terminal differentiation event, including cell cycle, cell fusion, and the stage-specific regulation of cuticle gene expression. A gene called lin-29 has been identified as a pivotal component in programming the execution of seam cell terminal differentiation. Loss of lin-29 function causes hypodermal cells of the adult stage to reiterate indefinitely the larval program of cell divisions instead of exiting the cell cycle and differentiating. lin-29 encodes a putative transcription factor of the (Cys)2-(His)2 class, and, thus likely programs these events by regulating the transcription of other genes. The major objective of this work is to identify genes controlled by the lin-29 protein. Three approaches will be employed to identify lin-29-regulated genes. ı1! Interactions of lin-29 protein with predicted target genes: The stage-specific expression of the cuticle collagen genes, col-17 and col-19, will be analyzed. col-17 expression is transcriptionally repressed during the larval/adult switch, whereas col-19 expression is activated. The role of lin-29 in the alternative repression and activation of these genes will be tested. ı2! Differential hybridization screens to identify genes whose expression is influenced by the presence or absence of lin-29 protein: Genes isolated in this fashion will be examined for direct regulation by lin-29, and those directly regulated will be characterized further. ı3! Development of a protein-DNA UV crosslinking method to isolate DNA sequences bound by a specific protein in vivo: In the future, this method will be used to identify genes bound by lin-29 protein in vivo, and should also be applicable to the isolation of genes controlled by other C. elegans gene-regulatory proteins. Genes involved in cell cycle exit and cell fusion, as well as additional stage-specifically expressed cuticle genes, should be identified by these methods. Analysis of the types of genes regulated by lin-29 in vivo will lead to an understanding of how a complex terminal differentiation program is orchestrated at a specific time during development.

多细胞组织的正确形式需要精确控制终端分化事件。这项研究将介绍在简单模型组织（线虫C. exekemans）开发过程中如何执行单个终端分化事件。该分析将集中于侧向皮下注射“接缝”细胞的末端分化，该细胞在最终（第四）蜕皮中终极分化。在此末端分化事件中，对几个不同的细胞过程进行了协调控制，包括细胞周期，细胞融合和角质层基因表达的特定阶段调控。一个称为LIN-29的基因已被确定为编程接缝细胞末端分化的执行时的关键成分。 LIN-29功能的丧失导致成人阶段的皮下注射细胞无限期地重复细胞分裂的幼虫程序，而不是退出细胞周期和区分。 LIN-29编码（CYS）2-（His）2类的推定转录因子，因此很可能通过控制其他基因的转录来编程这些事件。这项工作的主要目的是确定由LIN-29蛋白控制的基因。将采用三种方法来识别LIN-29调节的基因。 ı1！ LIN-29蛋白与预测靶基因的相互作用：角质层胶原基因COL-17和COL-19的阶段特异性表达将进行分析。 COL-17表达在幼虫/成人开关期间转录反射，而Col-19表达被激活。 LIN-29在这些基因的替代表达和激活中的作用将进行测试。 ı2！差异杂交筛选以鉴定其表达受到LIN-29蛋白的存在或不存在的基因：以这种方式分离的基因将通过LIN-29进行直接调节，而直接调节的基因将进一步表征。 ı3！开发一种蛋白-DNA紫外线交联方法来分离由特定蛋白质在体内结合的DNA序列的开发：将来，该方法将用于鉴定由Lin-29蛋白在体内绑定的基因，也应适用于由其他C. exelelans Gener-Gener-Gener-Gener-Gener-Gener-gen-Gener-gener-gener-gener-gener-gener-gener-gener-gener-Gener-gener-gener-gener-gener-genem-Gener-Gener-gene-gener-gene-gener-Gener-gener-gene-gene-gener-Gener-基因蛋白的隔离。这些方法应通过这些方法鉴定参与细胞周期出口和细胞融合以及其他阶段特异性表达的角质层基因的基因。对由LIN-29在体内调节的基因类型的分析将导致对在开发过程中特定时间进行编排的复杂终端分化程序的理解。