The Journal of Membrane Biology

, Volume 150, Issue 3, pp 243–253

Effect of Antipeptide Antibodies Directed against Three Domains of Connexin43 on the Gap Junctional Permeability of Cultured Heart Cells

  • B.  Bastide
  • T.  Jarry-Guichard
  • J.P.  Briand
  • J.  Délèze
  • D.  Gros

DOI: 10.1007/s002329900048

Cite this article as:
Bastide, B., Jarry-Guichard, T., Briand, J. et al. J. Membrane Biol. (1996) 150: 243. doi:10.1007/s002329900048

Abstract.

Cell-to-cell communication can be blocked by intracellular injections of antibodies raised against gap junction proteins, but the mechanism of channel obstruction is unknown. Binding to connexins could lead to a conformational change, interfere with regulatory domains or cause a steric hindrance. To address these questions, the effects on cell-to-cell communication of affinity purified polyclonal antibodies raised against peptides reproducing the intracellular sequences 5–17, 314–322 and 363–382 of rat connexin43 were investigated in cultured rat ventricular cells. The antibodies against sequence 363–382 were characterized by immunoblotting and immunocytochemistry. Characterization of antibodies 5–17 and 314–322 has been previously reported. In a first series of experiments, the effect on gap junctional communication was assessed by injecting a junction-permeant fluorescent dye into cells adjacent to one cell previously microinjected with antibodies. In a second series, junctional permeability was quantitatively determined on records of fluorescence recovery after the photobleaching of 6-carboxyfluorescein-loaded cells. Antibodies 5–17 marked a 43 kDa band on immunoblots, but did not immunolabel gap junctions and had no functional effect. Antibodies 314–322 recognized the 43 kDa protein and labeled the intercalated disks, but failed to interfere with junctional permeability. Antibodies to the nearby sequence 363–382, for which all immunospecific tests had been positive, caused a delayed diffusional uncoupling in 50% of the microinjected cells. It is suggested that the blocking of junctional communication by antibodies results from interference with a regulatory domain of the connexin.

Key words: Cardiac myocytes — Gap junction — Intercellular communication — Connexin43 — Antipeptide antibodies — Cell-to-cell dye transfer

Copyright information

© 1996 Springer-Verlag New York Inc.

Authors and Affiliations

  • B.  Bastide
    • 1
  • T.  Jarry-Guichard
    • 2
  • J.P.  Briand
    • 3
  • J.  Délèze
    • 1
  • D.  Gros
    • 2
  1. 1.Laboratoire de Physiologie Cellulaire, Unité de Recherche Associée au CNRS n° 1869, Faculté des Sciences, 86022 POITIERS cedex, FranceFR
  2. 2.IBDM, Laboratoire de Génétique et Physiologie du Développement, UMR C9943, Faculté des Sciences de Luminy, Case 907, 13288 MARSEILLE cedex 09, FranceFR
  3. 3.Laboratoire d'Immunologie des peptides et virus, UPR CNRS 9021, 67089 STRASBOURG cedex, FranceFR