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Journal of Cell Science, Vol 108, Issue 9 3079-3090, Copyright © 1995 by Company of Biologists
JOURNAL ARTICLES |
MM Atkinson, PD Lampe, HH Lin, R Kollander, XR Li and DT Kiang
Department of Medicine, University of Minnesota, Minneapolis 55455, USA.
Direct communication between cells via gap junctions is thought to be an important component of homeostasis and coordinated cellular responses to external signals. We investigated how the second messenger cAMP exerts its effects on junctional communication in a mouse mammary tumor cell line, MMT22. Junctional permeance was quantitatively assessed using dye microinjection and video microscopy. An increase of permeance was found after exposure to 8-bromo-cAMP, being detectable after 30 minutes of treatment and attaining a fourfold higher level of permeance by 24 hours. This elevated level was maintained with continuous exposure to 8-bromo-cAMP for seven days. The permeability change was accompanied by an increase in gap junctions as shown by freeze-fracture electron microscopy and by confocal microscopy using antibodies directed against the gap junction protein, connexin43. The amount of detergent-insoluble connexin43 also increased with 8-bromo-cAMP treatment, and most of the increase could be attributed to an increase of slower migrating (i.e. phosphorylated) species of connexin43. However, connexin43 mRNA and the total cellular content of connexin43 did not change over this period of exposure to 8-bromo-cAMP, as shown by densitometric analyses of northern and western blots. We conclude that 8-bromo-cAMP affects the distribution of connexin43 such that a greater proportion of the protein is utilized for channel formation. Since these changes were relatively slow to develop and persisted with prolonged exposure to 8-bromo-cAMP, it is possible that the junctional permeability of these mammary tumor cells is linked to the 'basal' level of cAMP, i.e. levels maintained by the cells in accordance with a particular cell state.
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