Assembly of hepatic gap junctions. Topography and distribution of connexin 32 in intracellular and plasma membranes determined using sequence-specific antibodies

Rahman, S., Carlile, G. and Evans, William Howard 1993. Assembly of hepatic gap junctions. Topography and distribution of connexin 32 in intracellular and plasma membranes determined using sequence-specific antibodies. Journal of Biological Chemistry 268 (2) , pp. 1260-1265.

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Abstract

The subcellular distribution in rat liver and the topography in intracellular and plasma membranes of connexin 32, a major protein component of gap junctions, was studied using sequence-specific anti-peptide antibodies generated to extracellular and intracellular domains of the protein. The distribution of connexin 32 in liver analyzed using SDS-polyacrylamide gel electrophoresis and Western blotting showed the relative protein levels in the subcellular fractions to be: lateral plasma membranes > Golgi membranes > sinusoidal plasma membranes > lysosomes. Low amounts of connexin 32 were detected in microsomes, endosomes, and bile canalicular plasma membranes. Six highly conserved cysteine residues are located in the amino acid sequences comprising the two extracellular loops of all connexins thus far isolated, and these loops are positioned to extend the channel in the lipid bilayers across the intercellular region of the gap junction. In the present work, the intramolecular disulfide bonds linking the extracellular loops in gap junctions were shown to be present in connexins located in plasma membranes, Golgi, and a microsomal fraction, and it was concluded that the disulfide linkages were formed in the endoplasmic reticulum. In addition, immature configurations of connexin 32, probably occurring during membrane insertion, were detected in liver microsomal fractions. The results contribute to charting of the biogenetic routes followed by connexins in hepatocytes and the general mechanisms of gap junction assembly.

Item Type:	Article
Date Type:	Publication
Status:	Published
Schools:	Medicine
Subjects:	R Medicine > R Medicine (General)
Uncontrolled Keywords:	Amino Acid Sequence, Animals, Antibodies, Cell Fractionation, Cell Membrane/metabolism, Cell Membrane/ultrastructure, Connexins, Electrophoresis, Polyacrylamide Gel, Glycoside Hydrolases, Golgi Apparatus/metabolism, Golgi Apparatus/ultrastructure, Intercellular Junctions/metabolism, Intercellular Junctions/ultrastructure, Intracellular Membranes/metabolism, Intracellular Membranes/ultrastructure, Liver/metabolism, Liver/ultrastructure, Membrane Proteins/analysis, Membrane Proteins/immunology, Membrane Proteins/metabolism, Microsomes, Liver/metabolism, Microsomes, Liver/ultrastructure, Models, Biological, Molecular Sequence Data, Rats, Rats, Sprague-Dawley, Sulfhydryl Compounds/analysis
Publisher:	American Society for Biochemistry and Molecular Biology
ISSN:	0021-9258
Last Modified:	17 Mar 2021 02:58
URI:	https://orca.cardiff.ac.uk/id/eprint/66904

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