Mechanism of v-Src- and mitogen-activated protein kinase-induced reduction of gap junction communication

Abstract

Connexin (Cx)43 gap junction channels are phosphorylated by numerous protein kinases, with the net effect typically being a reduction in gap junction communication (GJC). This reduction must result from a decrease in channel open probability, unitary conductance, or permselectivity, because previous results suggest that channel number is unaffected. Coexpression of v-Src with wild-type Cx43 (Cx43-wt) but not Cx43 with tyrosine to phenylalanine substitutions at 247 and 265 (Cx43-Y247,265F) resulted in reduced electrical and dye coupling but no change in single-channel amplitudes. EGF treatment of cells expressing Cx43-wt but not Cx43 with serine to alanine substitutions at 255, 279, and 282 (Cx43-S255,279,282A) resulted in reduced GJC, also with no change in single-channel amplitude. Dye coupling was reduced to a far greater extent than electrical coupling, suggesting that channel selectivity was also altered but with minimal effect on unitary conductance. The absence of Src- and MAPK-induced reductions in single-channel amplitude suggests that the decreases in GJC induced by these kinases result from reduced channel open probability and possibly altered selectivity.

Gap junction channel gating modulated through protein phosphorylation

“…Cells expressing Cx43wt exhibited single channel γ j main state values of approximately 94 pS, in addition to substate conductances of 47 pS (Cottrell et al, 2003). Cells co-expressing Cx43wt and v-Src, showed unitary conductances of 87 and 115 pS with substate values at 48 and 66 pS.…”

“…In these studies (Cottrell et al, 2003), the effects of v-Src on the macroscopic electrical conductances (g j ) and single channel unitary conductances (γ j ) of connexin43 channels were examined. Connexin43 knockout mouse cell lines expressing exogenous wild type connexin43 (Cx43wt) or connexin43 tyrosine site mutants (Cx43-Y247F, Cx43-Y265F, or Cx43-Y247F,Y265F) in the absence or presence of v-Src were employed in these studies (Cottrell et al, 2003;Lin et al, 2001). G j was significantly diminished in cell lines co-expressing Cx43wt and v-Src, compared to the control Cx43wt alone value (Cottrell et al, 2003).…”

“…Connexin43 knockout mouse cell lines expressing exogenous wild type connexin43 (Cx43wt) or connexin43 tyrosine site mutants (Cx43-Y247F, Cx43-Y265F, or Cx43-Y247F,Y265F) in the absence or presence of v-Src were employed in these studies (Cottrell et al, 2003;Lin et al, 2001). G j was significantly diminished in cell lines co-expressing Cx43wt and v-Src, compared to the control Cx43wt alone value (Cottrell et al, 2003). Cells coexpressing v-Src and the Cx43-Y247F,Y265F double mutant, showed no significant change in g j compared to Cx43wt.…”

“…Changes in cell-to-cell communication have been observed after activation of protein kinases (Cottrell et al, 2003;Duncan and Fletcher, 2002;Shi et al, 2001) and protein phosphatases (John et al, 2003). From the initial reports (Kanno et al, 1984;Azarnia et al, 1988), it has been reasonable to speculate that phosphorylation induces channel gating.…”

Oxidized Phospholipids Alter Vascular Connexin Expression, Phosphorylation, and Heterocellular Communication

“…In particular, phosphorylation of tyrosine 265 on Cx43 has been associated with decreased cell coupling in multiple cell types through a v-src kinase signaling cascade. [33][34][35] Because there is evidence that OxPAPC induces activation of v-src, 15 it is conceivable that v-src mediates the OxPAPC-induced phosphorylation of Cx43.…”