Growth Arrest in Differentiating Keratinocytes

描述（由申请人提供）：众所周知， 真核细胞G1期和S期穿越的复杂调控 受一个潜伏蛋白激酶家族控制， 通过合成和结合其积极作用的细胞周期蛋白亚基而激活。 相比之下，关于以下的势函数的了解少得多： 在哺乳动物细胞终末分化过程中的细胞周期蛋白/CDKs。像 增殖、分化是通过一系列严格控制的 事件的时间调节序列。不过，目前还不清楚， 细胞周期蛋白/CDK已经成为细胞生长中的一个既定范例， 我们已经确定了同样的负责调节分化的基因， 细胞周期蛋白G2作为一个强诱导基因在小鼠角质形成细胞被迫接受 在悬浮培养模型和基底上角质形成细胞中的分化 从皮肤上取下来的本申请旨在确定这种细胞周期蛋白的作用， 在终末期小鼠角质形成细胞中， 分化 在第一个具体目标中，拟议的研究将寻求确定 细胞周期蛋白G2与细胞周期蛋白/cdks协同发挥作用， 在生长细胞中的复制控制。这些研究将利用异位 G1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 通过平行的独立路径。第二部分中描述的研究 具体目标将侧重于确定造成 我们在细胞周期蛋白G2的转录诱导中发现， 角质形成细胞进一步的研究将确定p53的作用 同源物p63在细胞周期蛋白G2诱导中作用。最后的具体目标将针对 鉴定细胞周期蛋白G2相互作用蛋白，以鉴定催化的 细胞周期蛋白G2的伴侣。这些研究将通过使用 分离对重组细胞周期蛋白G2具有亲和力的蛋白质的生物化学方法 以及通过使用双杂交系统筛选哺乳动物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.