Identification of binding partners for the cytoplasmic loop of connexin43: a novel interaction with β-tubulin.
Connexin43 (Cx43), a component of gap junctions, has a relatively large carboxy-terminal region with multiple proteomic interactions. Proteomic interactions with its cytoplasmic loop, however, are poorly defined. The goal of this study is to examine proteomic interactions involving the cytoplasmic loop (CL) of Cx43. The authors utilized various techniques, including glutathione-S-transferase (GST) pull-down, immunoblot analysis, two-dimensional (2D) gel electrophoresis, and mass spectrometry, to elucidate binding partners for Cx43-CL. The authors identified novel interactions with Cx43-CL involving α- and β-tubulin, myelin basic protein, and Purα. Because tubulin interacts with the C-terminus of Cx43 (Cx43-CT), the authors further investigated the nature of the interaction between β-tubulin and Cx43-CL. β-Tubulin binds with the full length of Cx43-CL with approximately one-fifth the affinity of the interaction between Cx43-CT and β-tubulin. This study demonstrates novel proteomic interactions involving Cx43-CL that may lead to a more complete understanding of trafficking and gating of gap junction channels.
Kang
Eunice Y
EY
Leon H. Charney Division of Cardiology, Department of Medicine, New York University School of Medicine, New York, New York 10016, USA.
Ponzio
Marc
M
Gupta
Pritha P
PP
Liu
Fangyu
F
Butensky
Adam
A
Gutstein
David E
DE
D000818:Animals; D018031:Connexin 43 / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000032:analysis* / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D017629:Gap Junctions / Q000378:metabolism; D005982:Glutathione Transferase / Q000037:antagonists & inhibitors; D015151:Immunoblotting; D007473:Ion Channels / Q000378:metabolism; D013058:Mass Spectrometry; D051379:Mice; D004676:Myelin Basic Protein / Q000032:analysis* / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000032:analysis* / Q000378:metabolism; D011485:Protein Binding; D040901:Proteomics / Q000379:methods*; D014404:Tubulin / Q000032:analysis* / Q000378:metabolism
D018031:Connexin 43; D004268:DNA-Binding Proteins; D007473:Ion Channels; D004676:Myelin Basic Protein; D009419:Nerve Tissue Proteins; C121485:Pura protein, mouse; D014404:Tubulin; D005982:Glutathione Transferase