Molecular cloning, functional expression, and tissue distribution of a novel human gap junction-forming protein, connexin-31.9. Interaction with zona occludens protein-1.
A novel human connexin gene (GJA11) was cloned from a genomic library. The open reading frame encoded a hypothetical protein of 294 amino acid residues with a predicted molecular mass of 31,933, hence referred to as connexin-31.9 (Cx31.9) or alpha 11 connexin. A clone in GenBank containing the Cx31.9 gene localized to chromosome 17q21.2. Northern analysis of Cx31.9 showed a major 4.4-kilobase transcript, which was expressed at varying levels in all tissues analyzed. Two monoclonal antibodies generated against different domains of Cx31.9 recognized a 30-33-kDa protein from cells overexpressing Cx31.9. Immunofluorescence of overexpressing cells indicated the presence of Cx31.9 between adjacent cells, consistent with its localization to gap junctions. Double voltage clamp analyses of Cx31.9-overexpressing cells, and of paired Xenopus oocytes injected with Cx31.9 cRNA, demonstrated junctional currents indicative of gap junction channel formation. In contrast to previously characterized connexins, Cx31.9 showed no voltage-dependent gating within a physiologically relevant range. Cx31.9 was detected in human tissues by immunoblot analysis, and immunofluorescence localized Cx31.9 expression to vascular smooth muscle cells. Furthermore, it was demonstrated that Cx31.9 interacted with ZO-1. Thus, Cx31.9 represents a novel connexin gene that in vivo generates a protein with unique voltage gating properties.
Nielsen
Peter A
PA
Department of Cell Biology, Scripps Research Institute, La Jolla, California 92037, USA. petern@scripps.edu
Beahm
Derek L
DL
Giepmans
Ben N G
BN
Baruch
Amos
A
Hall
James E
JE
Kumar
Nalin M
NM
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D017630:Connexins / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D017629:Gap Junctions / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000378:metabolism; D009865:Oocytes / Q000502:physiology; D016366:Open Reading Frames; D018408:Patch-Clamp Techniques; D010750:Phosphoproteins / Q000378:metabolism; D010802:Phylogeny; D011485:Protein Binding; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D014018:Tissue Distribution; D029867:Xenopus Proteins; D014982:Xenopus laevis; D062826:Zonula Occludens-1 Protein
D000911:Antibodies, Monoclonal; D017630:Connexins; C468137:GJD3 protein, human; D008565:Membrane Proteins; D010750:Phosphoproteins; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; C569045:TJP1 protein, Xenopus; C569042:TJP1 protein, human; D029867:Xenopus Proteins; D062826:Zonula Occludens-1 Protein