Structural Studies of Tissue Specificity and Diversity of Gap Junctions.

组织特异性和间隙连接多样性的结构研究。

基本信息

批准号：
03454117
负责人：
SHIBATA Yosaburo
金额：
$ 3.65万
依托单位：
KYUSHU UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1991
资助国家：
日本
起止时间：
1991 至 1992
项目状态：
已结题

项目摘要

Molecular architectures of Liver Gap Junctions: The localizations of gap junction proteins, Cx32 and Cx26 were examined in rat and guinea pig livers by using site-specific antibodies against synthetic oligopeptides. Double labeling immunofluorescence microscopy revealed that in guinea pig liver both proteins spread throughout the liver lobules and seemed to localize together within the same gap junctional plaque. Quick-freeze, deep-etch immunoelectron microscopy showed that immunolabeling of isolated guinea pig liver gap junctional plaques with either Cx32 or Cx26 antiserum yielded complete and dense antibody decoration of the cytoplasmic surface of the plaques. We did not observe any focal or patchy clusters of the labeling in any plaques examined. Double labeling immuno electron microscopy confirmed that both Cx32 and Cx26 are colocalized in the same gap junction plaques. These results suggest that in the hepatocytes expressing both Cx32 and Cx26, both types of gap junction proteins are not segregated but intermix randomly within the same plaques.Cytoplasmic Surface Structures of Bovine lens Fiber Gap Junctions: Bovine lens fibers studied by quick freeze-deep etch replica method revealed particulate substructures on cytoplasmic surfaces in situ. In isolated membranes, thin section electron microscopy showed that 16-17 nm pentalamellar junctions were included in this specimen. Deep-etch replica showed that gap junctional plaques also had particles on their cytoplasmic surfaces clearly distinguished from the surrounding particle-free non-gap junctional surfaces. Treatment of isolated lens membranes with endoproteinase glu-C induced cleavage of MP70 to MP38 in SDS-PAGE and loss of their antigenicity to the anti-MP70 mono-clonal antibody, whereas the particulate patterns persisted still on the cytoplasmic surfaces of gap junctional plaques, suggesting the coexistance of additional proteins other than MP70 in the lens fiber gap junction structures.

肝间隙连接的分子体系结构：通过使用针对合成寡肽的位点特异性抗体，在大鼠和豚鼠肝脏中检查了间隙连接蛋白，CX32和CX26的局部化。双重标记免疫荧光显微镜显示，在豚鼠肝脏中，这两种蛋白质均遍布整个肝小叶，似乎在同一间隙连接斑块中置于同一间隙连接斑块中。快速冻结，深蚀刻免疫电子显微镜表明，具有CX32或CX26抗血清的分离的豚鼠肝间隙间隙连接斑的免疫标记产生了斑块的细胞质表面的完整且密集的抗体装饰。我们没有在检查的任何斑块中观察到标记的任何焦点或斑点簇。双重标记免疫电子显微镜证实，CX32和CX26均在同一间隙连接斑块中共定位。 These results suggest that in the hepatocytes expressing both Cx32 and Cx26, both types of gap junction proteins are not segregated but intermix randomly within the same plaques.Cytoplasmic Surface Structures of Bovine lens Fiber Gap Junctions: Bovine lens fibers studied by quick freeze-deep etch replica method revealed particulate substructures on cytoplasmic surfaces原位。在孤立的膜中，薄截面电子显微镜表明，该标本中包括16-17 nm的Pentalamellar连接。 Deep-etch副本表明，间隙连接斑块在其细胞质表面上也有颗粒与周围无颗粒的非差异连接表面明显区别。用内蛋白酶GLU-C在SDS-PAGE中诱导MP70至MP38的分离的分离透镜膜的处理以及对抗MP70单链抗体的抗原性的损失，而颗粒模式仍然存在于其他含量7的含量上，而颗粒的其他图案则持续使用。在镜头纤维间隙连接结构中。