Activation of the osmo-sensitive chloride conductance involves P21rho and is accompanied by a transient reorganization of the F-actin cytoskeleton.
Hypo-osmotic stimulation of human Intestine 407 cells rapidly activated compensatory CL- and K+ conductances that limited excessive cell swelling and, finally, restored the original cell volume. Osmotic cell swelling was accompanied by a rapid and transient reorganization of the F-actin cytoskeleton, affecting both stress fibers as well as apical ruffles. In addition, an increase in total cellular F-actin was observed. Pretreatment of the cells with recombinant Clostridium botulinum C3 exoenzyme, but not with mutant enzyme (C3-E173Q) devoid of ADP-ribosyltransferase activity, greatly reduced the activation of the osmo-sensitive anion efflux, suggesting a role for the ras-related GTPase p21rho. In contrast, introducing dominant negative N17-p21rac into the cells did not affect the volume-sensitive efflux. Cell swelling-induced reorganization of F-actin coincided with a transient, C3 exoenzyme-sensitive tyrosine phosphorylation of p125 focal adhesion kinase (p125FAK) as well as with an increase in phosphatidylinositol-3-kinase (PtdIns-3-kinase) activity. Pretreatment of the cells with wortmannin, a specific inhibitor of PtdIns-3-kinase, largely inhibited the volume-sensitive ion efflux. Taken together, our results indicate the involvement of a p21rho signaling cascade and actin filaments in the activation of volume-sensitive chloride channels.
Tilly
B C
BC
Department of Biochemistry, Cardiovascular Research Institute COEUR, Medical Faculty, Erasmus University, Rotterdam, The Netherlands.
Edixhoven
M J
MJ
Tertoolen
L G
LG
Morii
N
N
Saitoh
Y
Y
Narumiya
S
S
de Jonge
H R
HR
D036002:ADP Ribose Transferases / Q000494:pharmacology; D000199:Actins / Q000502:physiology* / Q000648:ultrastructure; D000838:Anions / Q000378:metabolism; D001905:Botulinum Toxins; D015815:Cell Adhesion Molecules / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D018118:Chloride Channels / Q000187:drug effects / Q000502:physiology*; D003599:Cytoskeleton / Q000502:physiology*; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D019204:GTP-Binding Proteins / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007038:Hypotonic Solutions / Q000494:pharmacology; D007422:Intestines / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D009997:Osmotic Pressure; D019869:Phosphatidylinositol 3-Kinases; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000187:drug effects / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000187:drug effects / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism; D020741:rho GTP-Binding Proteins
D000199:Actins; D000838:Anions; D015815:Cell Adhesion Molecules; D018118:Chloride Channels; D007038:Hypotonic Solutions; D014443:Tyrosine; D036002:ADP Ribose Transferases; C063007:exoenzyme C3, Clostridium botulinum; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human; D001905:Botulinum Toxins; D019204:GTP-Binding Proteins; D020741:rho GTP-Binding Proteins