Dominant negative effect of connexin33 on gap junctional communication is mediated by connexin43 sequestration.
Gap junctional intercellular communication is involved in the control of cell proliferation and differentiation. Connexin33, a member of the multi-gene family of gap junction proteins, exerts an inhibitory effect on intercellular communication when injected into Xenopus oocytes. However, the molecular mechanisms involved remain to be elucidated. Our results show that connexin33 was only expressed within the seminiferous tubules in the testis. In contrast to the majority of connexins, connexin33 was unphosphorylated. Immunoprecipitation experiments revealed that connexin33 physically interacted with connexin43, mainly with the phosphorylated P1 isoform of connexin43 but not with connexin26 and connexin32, two other connexins expressed in the tubular compartment. In Sertoli cells and COS-7 cells, connexin43 was located at the plasma membrane, whereas in connexin33 transfected cells, the specific association of connexin33/43 was sequestered in the intracellular compartment. High-resolution fluorescent deconvolution microscopy indicated that the connexin33/43 complex was mainly found within early endosomes. Sequestration of connexin33/43 complex was associated with a complete inhibition of the gap junctional coupling between adjacent cells. These findings provide the first evidence of a new mechanistic model by which a native connexin, exerting a dominant negative effect, can inhibit gap junctional intercellular communication. In the testis, connexin33 could exert a specific role on germ cell proliferation by suppressing the regulatory effect of connexin43.
INSERM EMI 00-09, IFR 50, Faculté de Médecine, Avenue de Valombrose, 06107 Nice CEDEX 02, France.
Céline
C
Fiorini
Baharia
B
Mograbi
Laurent
L
Cronier
Isabelle
I
Bourget
Xavier
X
Decrouy
Marielle
M
Nebout
Bernard
B
Ferrua
André
A
Malassine
Michel
M
Samson
Patrick
P
Fénichel
Dominique
D
Segretain
Georges
G
Pointis
D000818:Animals; D019556:COS Cells; D002450:Cell Communication / Q000502:physiology*; D002522:Chlorocebus aethiops; D018031:Connexin 43 / Q000235:genetics / Q000378:metabolism*; D017630:Connexins / Q000235:genetics / Q000378:metabolism*; D011992:Endosomes / Q000737:chemistry / Q000378:metabolism; D017629:Gap Junctions / Q000378:metabolism*; D008297:Male; D010766:Phosphorylation; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012708:Sertoli Cells / Q000166:cytology / Q000502:physiology; D014018:Tissue Distribution
D018031:Connexin 43; D017630:Connexins; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; C099106:Gja6 protein, rat