Electron microscopic immunocytochemistry of protein kinase C using subspecies-specific anti-peptide antibody

使用亚种特异性抗肽抗体对蛋白激酶 C 进行电子显微镜免疫细胞化学分析

基本信息

批准号：
63870011
负责人：
TANAKA Chikako
金额：
$ 9.47万
依托单位：
Kobe University School of Medicine
依托单位国家：
日本
项目类别：
Grant-in-Aid for Developmental Scientific Research
财政年份：
1988
资助国家：
日本
起止时间：
1988 至 1989
项目状态：
已结题

项目摘要

Protein kinase C (PKC) is a key enzyme involved in the signal transduction mechanism of many physiological processes. The enzyme is now known to exist as a family of several subtly different, but closely related subspecies. Thus far several laboratories have isolated cDNA clones encoding distinct forms of PKC, namely, alpha, betaI, betaII, gamma, delta, epsilon, and zeta. PKC subspecies exhibit subtle difference in their enzymological characteristics and cellular locations, suggesting they have distinct physiological roles. We synthesized subspecies-specific oligopeptides for alpha, betaI, betaII, and gamma-PKC and obtained subspecies-specific antibodies by immunizing rabbits with the oligopeptides. We aimed in this project to elucidate the functional difference between the subspecies of PKC in nervous tissue using electron microscopic immunocytochemistry. The distribution of each PKC subspecies was distinctly different and the distributions did not completely correspond with those of any neurotransmitters and receptors. Under electron microscopy, gamma-PKC was present in the perikaryon, plasma membrane, nucleus, dendrite, and axon. betaI-PKC was just adjacent to plasma membrane, while betaII-PKC was located around Golgi complex. In the hippocampus, gamma-PKC was distributed diffusely throughout the cytoplasm of pyramidal cells from the perikarya to dendritic spine. In contrast, betaII-PKC was concentrated around Golgi complex and present diffusely in distal dendrites, except for the dendritic spines. Neither PKC subspecies could not be detected in the synaptic terminals. These results suggest that each PKC subspecies has a specific function in the neurons and that gamma-PKC and betaII-PKC are involved in long term potentiation postsynaptically in the hippocampus.

蛋白激酶C（PKC）是许多生理过程的信号转导机制涉及的关键酶。该酶现在已知是一个有几个微妙但密切相关的亚种的家族。迄今为止，几个实验室已经隔离了编码不同形式的PKC的cDNA克隆，即Alpha，Betai，Betaii，Gamma，Gamma，Delta，Epsilon和Zeta。 PKC亚种在其酶学特征和细胞位置表现出细微的差异，这表明它们具有不同的生理作用。我们合成了α，betai，betaii和gamma-pkc的亚种特异性寡肽，并通过用寡肽免疫兔子来获得亚种特异性抗体。我们旨在使用电子显微镜免疫细胞化学阐明神经组织中PKC亚种之间的功能差异。每个PKC亚种的分布截然不同，并且分布与任何神经递质和受体的分布都不完全对应。在电子显微镜下，γ-PKC存在于周围，质膜，核，树突和轴突中。 BETAI-PKC刚刚与质膜相邻，而Betaii-PKC位于Golgi Complex附近。在海马中，γ-PKC分布在整个锥体细胞的细胞质中，从周围的脊柱到树突状脊柱。相比之下，除树突状棘以外，betaii-pkc集中在高尔基体综合物周围，并在远端树突中扩散。在突触终端都无法检测到PKC亚种。这些结果表明，每个PKC亚种在神经元中具有特定的功能，并且γ-PKC和BETAII-PKC在海马中涉及突触后的长期增强。