Scientific article
Open access

A tyrosine-based sorting signal is involved in connexin43 stability and gap junction turnover

Published inJournal of cell science, vol. 116, no. Pt 11, p. 2213-2222
Publication date2003

The gap junction protein connexin43 is known to have a rapid turnover, involving degradation by both the proteasomal and lysosomal systems, but the structural features of connexin43 that govern these actions are not known. The connexin43 C-terminal sequence contains a proline-rich region corresponding to the consensus of a protein-protein interaction PY-motif (xPPxY), and an overlapping putative tyrosine-based sorting signal (Yxxphi; =hydrophobic), known to play a role in the intracellular trafficking of many membrane proteins. As both motifs may control turnover of connexin43, we used a combination of metabolic radiolabelling, immuno-precipitation and functional assays to determine the possible role of these motifs in controlling degradation of human connexin43 expressed in SKHep1 cells. Mutation V289D in the tyrosine-based sorting motif increased the steady-state pool of connexin43 by approximately 3.5-fold, while mutation P283L in the PY-motif produced a comparatively modest augmentation (1.7-fold). No additive effect was observed when the overlapping tyrosine was mutated. In pulse-chase experiments, the Y286A substitution increased the half-life of connexin43 from 2 to 6 hours, indicating that the increased steady-state levels reflected reduced protein degradation. Moreover, expression at the junctional membrane, as well as gap junction-mediated intercellular communication (GJC), were nearly abolished by lysosomal inhibitors and Brefeldin A in cells expressing wild-type connexin43, but were unaffected in the tyrosine mutant. These results provide strong evidence that the tyrosine-based motif of human connexin43 is a prime determinant controlling connexin43 stability, and consequently GJC, by targeting connexin43 for degradation in the endocytic/lysosomal compartment.

  • Carcinoma, Hepatocellular
  • Cell Line, Tumor
  • Connexin 43/genetics/metabolism
  • Cysteine Endopeptidases/metabolism
  • Endosomes/metabolism
  • Gap Junctions/metabolism
  • Humans
  • Liver Neoplasms
  • Lysosomes/metabolism
  • Multienzyme Complexes/metabolism
  • Mutagenesis/physiology
  • Proteasome Endopeptidase Complex
  • Protein Sorting Signals/physiology
  • Protein Transport/physiology
  • Tyrosine/metabolism
Citation (ISO format)
THOMAS, Marc et al. A tyrosine-based sorting signal is involved in connexin43 stability and gap junction turnover. In: Journal of cell science, 2003, vol. 116, n° Pt 11, p. 2213–2222. doi: 10.1242/jcs.00440
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Article (Published version)
ISSN of the journal0021-9533

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