Functional role of the carboxyl terminal domain of human connexin 50 in gap junctional channels.
Gap junction channels formed by connexin 50 (Cx50) are critical for maintenance of lens transparency. Because the C-terminus of Cx50 can be cleaved post-translationally, we hypothesized that channels formed by the truncated Cx50 exhibit altered properties or regulation. We used the dual whole-cell patch-clamp technique to investigate the macroscopic and single-channel properties of gap junctional channels formed by wild-type human Cx50 and a truncation mutant (Cx50A294stop) after transfection of N2A cells. Our results show that wild-type Cx50 formed functional gap junctional channels. The macroscopic Gjss-Vj relationship was well described by a Boltzmann equation with A of 0.10, V0 of 43.8 mV and Gjmin of 0.23. The single-channel conductance was 212 +/- 5 pS. Multiple long-lasting substates were observed with conductances ranging between 31 and 80 pS. Wild-type Cx50 gap junctional channels were reversibly blocked when pHi was reduced to 6.3. Truncating the C-terminus at amino acid 294 caused a loss of pHi sensitivity, but there were no significant changes in single-channel current amplitude or Gjss-Vj relationship. These results suggest that the C-terminus of human Cx50 is involved in pHi sensitivity, but has little influence over single-channel conductance, voltage dependence, or gating kinetics.
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Department of Physiology and Biophysics, Finch University of Health Science/The Chicago Medical School, 3333 Green Bay Rd, North Chicago, IL 60064, USA.
Berthoud
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Beyer
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Ebihara
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D000595:Amino Acid Sequence; D000818:Animals; D017630:Connexins; D003593:Cytoplasm; D004553:Electric Conductivity; D004594:Electrophysiology; D005136:Eye Proteins / Q000235:genetics / Q000502:physiology*; D017629:Gap Junctions / Q000378:metabolism*; D015870:Gene Expression; D006801:Humans; D007473:Ion Channels / Q000235:genetics / Q000502:physiology*; D051379:Mice; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D012756:Sheep; D014407:Tumor Cells, Cultured; D000099264:Connexin 50
D017630:Connexins; D005136:Eye Proteins; D007473:Ion Channels; D000099264:Connexin 50