Integrated MAP Kinase Signaling In Cell Differentiation

这项提议的目的是定义控制神经元分化的信号转导通路。我们对培养的PC12嗜铬细胞瘤细胞的研究表明，细胞外信号调节激酶(ERK)和cJun氨基末端激酶(JNK)家族的丝裂原激活蛋白(MAP)映射激酶以及特定的CREB和Jun家族成员在神经元特异性基因诱导中的整合。我们最近建立了多潜能小鼠胚胎发育的体外神经元分化条件，所得到的神经元与体内生成的神经元具有很高的代表性。从小鼠来源的ES细胞产生神经元的能力，其中编码MAPK组分和转录因子靶标的基因纯合缺失，为神经元分化的信号转导提供了强大的遗传途径。我们假设，不同的MAP激酶信号通路及其特定的转录因子靶标的整合可以作为分子开关来诱导获得神经表型所需的多个基因。为了验证这一假设，我们将完成这些特定的目标：目的1.确定在PC12细胞和ES细胞神经分化过程中诱导NFLC基因的信号通路。目的2.确定神经元分化过程中的调控元件。目的3.明确调控NFLC启动子的信号通路和转录因子在PC12和ES细胞中诱导多个神经特异性基因的分子开关的作用。MAP激酶在有丝分裂过程中得到了广泛的分析，但在分化过程中没有得到广泛的分析。完成这些目标将阐明这些信号通路调节细胞分化所需基因转录的机制，这是一个相对研究较少的研究领域。

The objective of this proposal is to define signal transduction pathways that control neuronal differentiation. Our studies with cultured PC12 pheochromocytoma cells, an established model for analysis of nerve growth factor (NGF)- dependent signaling pathways that control neural differentiation, indicate the integration of the extracellular signal-regulated kinase (ERK) and cJun N- terminal kinase (JNK) families of mitogens-activated protein (MAP) MAP kinases and specific CREB and Jun family members in neuron-specific gene induction. We have recently established the conditions for in vitro neuronal differentiation of multipotent murine embryonic development and the resulting neurons are highly representative of their in vivo-generated counterparts. The ability to generate neurons for ES cells derived from mice in which genes encoding MAP kinase components and transcription factor targets are homozygously deleted provides a powerful genetic approach to signal transduction of neuronal differentiation. We hypothesize that integration of distinct MAP kinase signaling pathways and their specific transcription factor targets serve as a molecular switch to induce multiple genes required for acquisition of the neural phenotype. To test this hypothesis, we will complete these specific aims: Aim 1. Identify the signaling pathways that induce the NFLC gene during neuronal differentiation of PC12 cells and ES cells. Aim 2. Define the regulatory elements within the neuronal differentiation. Aim 3. Define the role of the signal pathways and transcription factors that regulate the NFLC promoter as a molecular switch involved in the induction of multiple neural-specific genes in PC12 and ES cells. MAP kinases have been extensively analyzed in the context of mitogenesis, but not differentiation. Completion of these aims will elucidate the mechanism by which these signal pathways can regulate the transcription of genes required for cell differentiation, a comparatively understudied research area.