权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

MOLECULAR MECHANISM OF REGULATION OF HISTAMINE H2RECEPTOR FUNCTION

组胺H2受体功能调控的分子机制

基本信息

批准号：
06454259
负责人：
SUGANO Kentaro
金额：
$ 4.22万
依托单位：
UNIVERSITY OF TOKYO
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1994
资助国家：
日本
起止时间：
1994 至 1995
项目状态：
已结题

项目摘要

In order to elucidate regulatory mechanisms of histamine H2 receptor function at the molecular level, we have done the following experiments :(1) We artificially mutated thr ee potential N-glycosylation ites in the canine histamine H2 receptor gene and examined the effects of N-glycosylation mutation on the appar ent molecular weight, ligand recognition, intracellular signal transduction and cellular localization of the receptors. From the analysis on the SDS-polyacrylaminde gel electr ophoresis combined with tunicamycin treatment, we found that two of the thr ee potential glycosylation sites are actually glycosylaed. Furthermore, recognition of the ligand, signaltransduction and cellular localization of non-glycosylated mutant receptors are essentially unaffected. Therefore, we conclude that N-glycosylation is not vital for histamine H2 receptor function.(2) Activation of histamine H2 receptor brings about at least two intracellular signaling mechanism, one thr-ough cAMP-proteinkinase A pathway and another thr ough phopholipase C and C-kinase pathway. We examined the role of C-kinase activation on the histamine H2 receptor function. We found that C-kinase activation sensitizes the histamine H2 receptor. This effect was abolished by staur osporine pretreatment without affectiong the desensitizing mechanism, indicating the inter action of two signaling mechanisms plays role in modulating the receptor function.(3) In our preliminary experiments, we found that histamine H2 receptors are phosphor ylated after histamine stimulation. Detailed analysis of the enzymes involved and the phosphor ylation sites are in progress.(4) We found that the receptor internalization after histamine stimulation was compromised in the C-terminally truncated mutant H2 receptors This mutation, however, did not affect the desensitization phenomenon. Thus, C-terminal portion of the receptor appears to be important for receptor cycling with in the cells, but not for desensitization.

为了从分子水平上阐明组胺H2受体功能的调节机制，我们做了以下实验：(1)人工突变犬组胺H2受体基因的三个潜在的N-糖基化位点，检测N-糖基化突变对受体的分子质量、配体识别、细胞内信号转导和细胞定位的影响。从SDS-聚丙烯酰胺凝胶电泳法结合衣霉素处理的分析中发现，三个潜在的糖基化位点中的两个实际上是糖囊化的。此外，配体的识别、信号转导和非糖基化突变受体的细胞定位基本上不受影响。因此，我们得出结论：N-糖基化对组胺H2受体的功能不是至关重要的。(2)组胺H2受体的激活至少导致两种细胞内信号机制，一种是通过cAMP-蛋白激酶A途径，另一种是通过磷脂酶C和C-激酶途径。我们研究了C-激酶激活对组胺H2受体功能的影响。我们发现C-激酶的激活使组胺H2受体变得敏感。这种作用可被葡萄球菌素预先处理而取消，但不影响脱敏机制，表明两种信号机制的相互作用在调节受体功能中起作用。(3)在我们的初步实验中，我们发现组胺H2受体在组胺刺激后被磷酸化。对涉及的酶和磷酸化位点的详细分析正在进行中。(4)我们发现组胺刺激后的受体内化在C端截短的H2受体中受到损害，但该突变并不影响脱敏现象。因此，受体的C末端部分似乎对细胞内的受体循环很重要，但对脱敏作用并不重要。