HORMONAL REGULATION OF CELLULAR METABOLISM

蛋白质的磷酸化-去磷酸化是最重要的 调节细胞功能的机制。蛋白激酶C (PKC)已成为监管的关键监管要素 许多细胞功能。多个PKC亚种已被鉴定为 分子克隆和各种同工酶具有明显的差异 组织、细胞和亚细胞的分布，并有差异 在发展过程中表达。这些激酶在体内的功能作用 用N1E115对细胞分化的调控进行了检验 神经母细胞瘤细胞。PKC激活佛波酯可中和cAMP- 选择性易位诱导这些细胞突起生长 激活PKC Beta1，但不激活PKcheeta。激活PKC测试版结果 在M/R=90K蛋白的磷酸化中，而cAMP的激活- 依赖的蛋白激酶减弱其磷酸化。一种方法，该方法 已经设计了一组PKC同工酶的特异性底物的制备 基于阳离子多肽与蛋白质相互作用的研究 Ca~(2+)/磷脂/洗涤剂混合胶束与 独特的表面结构。神经颗粒素(Ng)、神经调素(Nm)和Marcks 是大脑中最突出的PKC底物。Ng和Nm绑定 钙调素(CaM)在无钙条件下具有高亲和力，而Marcks 在有钙离子存在的情况下这样做。PKC对这些蛋白的磷酸化作用 降低它们对凸轮的亲和力，从而释放凸轮的凸轮- 依赖的酶。除了磷酸化诱导的效应外， 被过氧化氢或一氧化氮氧化的Ng也会降低亲和力 这种蛋白质是给卡姆的。Ng和Nm被蛋白质去磷酸化 磷酸酶1，2A和钙调神经磷酸酶和Marcks通过 蛋白磷酸酶1与磷酸酶2A和钙调神经磷酸酶相比。这些 后两种磷酸酶选择性地识别Marcks中的一组位点， 而磷酸酶1使所有的位点去磷酸化。约束： Marcks的磷脂酰肌醇抑制该蛋白的磷酸化。 磷酸化的Marcks对这些分子的亲和力降低 磷脂，并可能有助于该蛋白从 在PKC的刺激下，细胞膜转变为胞浆。一种新型的M/R=28K pKC/酪蛋白 鉴定了KII底物，并克隆了KII的基因。这种蛋白质 已被证明是N18神经母细胞瘤中酪蛋白激酶II的底物 细胞。中枢神经系统特异性PKC-γ和Ng的基因组结构为 分析的目的是为了定义组织特异性和发育- 调控这两个基因的表达。Ng的启动子活性 佛波酯或与PKC共转染法均可激活基因 CDNA。A M/R=20K蛋白与两个PKCγ的AT富集区结合 而Ng基因的启动子已经被鉴定出来，并被磷酸化了 蛋白激酶C的蛋白减弱了它与含有ATTA的DNA元件的结合， Ataa和AATA图案。

Phosphorylation-dephosphorylation of proteins is one of the most important mechanisms for the regulation of cellular functions. Protein kinase C (PKC) has emerged as a pivotal regulatory element for the regulation of many cellular functions. Multiple PKC subspecies have been identified by molecular cloning and the various isozymes were found to have distinct tissue, cellular, and subcellular distributions and were differentially expressed during development. The functional roles of these kinases in the control of cellular differentiation were tested with N1E115 neuroblastoma cells. PKC-activating phorbol ester counteracts the cAMP- induced neurite outgrowth of these cells by selective translocation and activation of PKC betaI but not PKCzeta. Activation of PKC betaI results in the phosphorylation of a M/r=90k protein, whereas activation of cAMP- dependent protein kinase attenuates its phosphorylation. A method for preparing substrates specific for a group of PKC isozymes has been devised based on onteraction of cationic polypeptide with Ca2+/phospholipid/detergent mixed micelles to form aggregates with a unique surface structure. Neurogranin (Ng), neuromodulin (Nm), and MARCKS are the most prominent PKC substrates in the brain. Ng and Nm bind calmodulin (CaM) with high affinity in the absence of Ca2+, whereas MARCKS does so in the presence of Ca2+. Phosphorylation of these proteins by PKC reduces their affinities toward CaM and thus release CaM for the CaM- dependent enzymes. In addition to the phosphorylation-induced effects, oxidation of Ng by peroxide or nitric oxide also reduces the affinity of this protein for CaM. Ng and Nm are dephosphorylated by protein phosphatase 1, 2A, and calcineurin and MARCKS is more effectively by protein phosphatase 1 compared to phosphatase 2A and calcineurin. These latter two phosphatases selectively recognize a set of sites in MARCKS, whereas phosphatase 1 dephosphorylates all the sites. Binding of phosphoinositides to MARCKS inhibits the phosphorylation of this protein. The phosphorylated MARCKS has a reduced affinity toward these phospholipids and may contribute to the translocation of this protein from membrane to cytosol upon stimulation of PKC. A novel M/r=28k PKC/casein kinase II substrate has been identified and its cDNA cloned. This protein has been shown to be a substrate of casein kinase II in N18 neuroblastoma cells. The genomic structures of the CNS-specific PKC gamma and Ng were analyzed for the purpose of defining tissue-specific and development- regulated expression of these two genes. The promoter activity of the Ng gene can be stimulated by phorbol ester or by co-transfection with PKC cDNAs. A M/r=20k protein binding to the AT-rich regions of both PKC gamma and Ng gene promoters has been identified and phosphorylation of this protein by PKC attenuates its binding to DNA elements containing ATTA, ATAA and AATA motifs.