Growth Arrest in Differentiating Keratinocytes

描述(由申请人提供)：公认的是，许多 真核细胞G1期和S期穿越过程中的复杂调控 受潜伏蛋白激酶家族的控制，这些蛋白激酶是 通过合成和结合其正向作用的细胞周期蛋白亚基而激活。 相比之下，人们对原子力的潜在作用知之甚少 哺乳动物细胞终末分化过程中的细胞周期蛋白/CDKs喜欢 增殖、分化通过一系列严格控制的 时间上有规律的事件顺序。然而，目前还不清楚同样的情况是否 细胞周期蛋白/CDK已经成为细胞生长的既定范式 我们已经确定了负责调节分化的相同基因 细胞周期蛋白G2在小鼠角质形成细胞中的强诱导基因 悬浮培养模型和超基底层角质形成细胞的分化 从皮肤中提取。此应用程序旨在确定此循环的角色 在终末期对小鼠角质形成细胞的生长抑制 差异化。 在第一个具体目标中，拟议的研究将寻求确定 Cyclin G2与Cyclin/CDK协同发挥作用 生长细胞中的复制控制。这些研究将利用异位 G1期和S期细胞周期蛋白的表达以及异位细胞周期蛋白G2的表达 确定是否存在功能关系或细胞周期蛋白G2是否起作用 通过一条平行的独立路径。第二部分描述的研究 具体目标将集中在找出造成这一事件的原因(S) 细胞周期蛋白G2的转录诱导作用 角质形成细胞。进一步的研究将确定P53的作用 细胞周期蛋白G2诱导中的同源基因p63。最终的具体目标将针对 鉴定细胞周期蛋白G2相互作用蛋白以鉴定一种催化 Cyclin G2的合伙人(S)。这些研究将通过使用 分离重组细胞周期蛋白G2亲和力蛋白的生物化学方法 也通过使用双杂交系统筛选哺乳动物的cDNA文库。

DESCRIPTION (provided by applicant): It is well established that much of the complex regulation exerted during G1 and S phase traverse in eukaryotic cells is under the control of a family of latent protein kinases, cdks, which are activated by synthesis and binding of their positively acting cyclin subunits. By contrast, substantially less is known about the potential function of cyclin/cdks during terminal differentiation in mammalian cells. Like proliferation, differentiation occurs through a series of tightly controlled temporally regulated sequence of events. However, it is unclear if the same cyclin/cdks that have become an established paradigm in cell growth are the same that are responsible for regulating differentiation We have identified cyclin G2 as a strongly induced gene in murine keratinocytes forced to undergo differentiation in a suspension culture model and in suprabasal keratinocytes harvested from skin. This application seeks to identify the role of this cyclin in enforcing a growth arrest in murine keratinocytes during terminal differentiation. In the first specific aim, the proposed studies will seek to establish whether cyclin G2 functions in collaboration with the cyclin/cdks that exert replicative control in growing cells. These studies will utilize ectopic expression of G1 and S phase cyclins as well as ectopic cyclin G2 expression to ascertain if there is a functional relationship or if cyclin G2 functions through a parallel independent pathway. The studies described in the second specific aim will focus on identifying the factor(s) responsible for the transcriptional induction of cyclin G2 that we have discovered in keratinocytes. Additional investigations will determine the role of the p53 homolog p63 in cyclin G2 induction. The final specific aim will be directed at identifying cyclin G2 interacting proteins in order to identify a catalytic partner(s) for cyclin G2. These studies will be accomplished by using biochemical methods to isolate proteins with affinity for recombinant cyclin G2 and also by screening mammalian cDNA libraries using a two hybrid system.