Temporal Control of Development in C.elegans

描述(由申请人提供)：这项工作的目的是了解 在后生动物中支配特定细胞命运决定的时间机制 发展。线虫的异时基因是全球性的 时间调节器，控制不同事件的顺序和时间 在胚胎后期发育期间，这项研究将调查其中一个 这些事件，外侧皮下“缝隙”细胞的终末分化， 在时间上仅限于野生动物的最后一次蜕皮。突变 在异时性基因中导致缝隙细胞末端分化发生 比平时早或晚。近12个异时性基因已经被 它们可以根据它们的属性大致分为两类 已知或建议采取行动的时间。早期作用基因指定发育 经历前三个幼虫阶段(L1-L3)，然后是晚期 作用基因接管并指导剩余的发育(L3后期-成人)。 这项研究将研究四个异时基因的分子作用， 早期作用基因LIN-42和LIN-58以及晚期作用基因LIN-57和 林-29。这项提案中描述的实验将试图理解 这些基因在组成其他基因的背景下的功能 异时基因途径。遗传学和分子上位性实验将是 并将确定这些基因的表达模式。林-42 编码一种与果蝇时期相似的蛋白质，一种参与 第二种生物计时机制，控制昼夜节律。 拟议的实验将调查该序列的功能相关性 同源性和酵母双杂交系统将被用来发现 不与42相互作用的蛋白质。LIN-58将被克隆，并收集信息 从它的身份将被用来设计其功能的分子测试。 因为只存在单一的LIN-57亚型等位基因，所以基因筛查 将进行检测以确定LIN-57(空)突变。这些等位基因将是 刻画和假设LIN.57在时间上由21 将检测NT let-7调控RNA。林-29发挥着关键作用，因为 缝隙细胞终末分化的下游调节因子。分子 将确定LIN-29活动的时间机制并确定关键序列 对于LIN-29，将定义规则。最后，基因筛查将是 被用来搜索额外的异时基因。 LIN-29功能的丧失导致成体阶段的细胞无限期 重复幼虫细胞分裂的程序，而不是离开细胞 循环和差异化。因此，林-29可以被概括地认为是 抑癌基因：细胞周期退出所必需的基因 对LIN-29的时间控制为生物体内的细胞如何 被指示在发育过程中的特定时间停止分裂 差异化。这方面的知识应该有助于我们理解这些问题 当增长控制出现问题时会发生这种情况，例如不适当的恢复 在某些癌症中发生的细胞分裂。

DESCRIPTION (provided by applicant): The aim of this work is to understand the timing mechanisms that govern specific cell fate decisions during metazoan development. The heterochronic genes of the nematode C. elegans are global temporal regulators that control the sequence and timing of diverse events during post embryonic development This research will investigate how one of these events, the terminal differentiation of lateral hypodermal "seam" cells, becomes temporally restricted to the final molt in wild-type animals. Mutations in the heterochronic genes cause seam cell terminal differentiation to occur earlier or later than normal. Nearly a dozen heterochronic genes have been identified and they can be roughly divided into two classes based upon their known or proposed times of action. The early acting genes specify developmental progression through the first three larval stages (L1-L3) and then the late acting genes take over and direct the remainder of development (late L3-Adult). This research will investigate the molecular roles of four heterochronic genes, the early acting genes, lin-42 and lin-58 and the late acting genes, lin-57 and lin-29. The experiments described in this proposal will seek to understand the functions of these genes in the context of the other genes that comprise the heterochronic gene pathway. Genetic and molecular epistasis experiments will be performed and the expression patterns of these genes will be determined. lin-42 encodes a protein with similarity to Drosophila Period, a protein involved in a second type of biological timing mechanism, control of circadian rhythms. Proposed experiments will investigate the functional relevance of this sequence homology and the yeast two-hybrid system will be employed to discover UN-42-interacting proteins. lin-58 will be cloned, and the information gleaned from its identity will be used to design molecular tests of its function. Because only a single, hypomorphic allele of lin-57 exists, genetic screens will be performed to identify lin-57(null) mutations. These alleles will be characterized and the hypothesis that lin.57 is temporally controlled by the 21 nt let-7 regulatory RNA will be tested. lin-29 plays a key role as the most downstream regulator of seam cell terminal differentiation. The molecular mechanism that times lin-29 activity will be determined and sequences critical for lin-29 regulation will be defined. Finally, genetic screens will be employed to search for addition heterochronic genes. Loss of lin-29 function causes cells of the adult stage to indefinitely reiterate the larval program of cell divisions rather than exiting the cell cycle and differentiating. Thus, lin-29 can be thought of in general terms as an anti-oncogene: a gene whose function is required for cell cycle exit Temporal control of lin-29 provides a model for how cells within an organism are instructed to cease dividing at a specific time in development and differentiate. This knowledge should aid in our understanding of the problems that occur when growth controls go awry, such as the inappropriate resumption of cell divisions that occurs in some cancers.