pCLCA1 lacks inherent chloride channel activity in an epithelial colon carcinoma cell line.
The effects of CLCA protein expression on the regulation of Cl(-) conductance by intracellular Ca(2+) and cAMP have been studied previously in nonepithelial cell lines chosen for low backgrounds of endogenous Cl(-) conductance. However, CLCA proteins have been cloned from, and normally function in, differentiated epithelial cells. In this study, we examine the effects of differentiation of the Caco-2 epithelial colon carcinoma cell line on modulation of Cl(-) conductance by pCLCA1 protein expression. Cl(-) transport was measured as (36)Cl(-) efflux, as transepithelial short-circuit currents, and as whole cell patch-clamp current-voltage relations. The rate of (36)Cl(-) efflux and amplitude of currents in patch-clamp studies after the addition of the Ca(2+) ionophore A-23187 were increased significantly by pCLCA1 expression in freshly passaged Caco-2 cells. However, neither endogenous nor pCLCA1-dependent Ca(2+)-sensitive Cl(-) conductance could be detected in 14-day-postpassage cells. In contrast to Ca(2+)-sensitive Cl(-) conductance, endogenous cAMP-dependent Cl(-) conductance does not disappear on Caco-2 differentiation. cAMP-dependent Cl(-) conductance was modulated by pCLCA1 expression in Caco-2 cells, and this modulation was observed in freshly passaged and in mature 14-day-postpassage Caco-2 cultures. pCLCA1 mRNA expression, antigenic pCLCA1 protein epitope expression, and pCLCA1 function as a modulator of cAMP-dependent Cl(-) conductance were retained through differentiation in Caco-2 cells, whereas Ca(2+)-dependent Cl(-) conductance disappeared. We conclude that pCLCA1 expression may increase the sensitivity of preexisting endogenous Cl(-) channels to Ca(2+) and cAMP agonists but apparently lacks inherent Cl(-) channel activity under growth conditions where endogenous channels are not expressed.
Loewen
Matthew E
ME
Department of Veterinary Biomedical Sciences, University of Saskatchewan, 52 Campus Drive, Saskatoon, Saskatchewan, Canada S7N 5B4.
Bekar
Lane K
LK
Walz
Wolfgang
W
Forsyth
George W
GW
Gabriel
Sherif E
SE
D000818:Animals; D018938:Caco-2 Cells; D002118:Calcium / Q000378:metabolism; D002277:Carcinoma / Q000378:metabolism* / Q000503:physiopathology; D002454:Cell Differentiation; D016764:Cell Polarity; D016922:Cellular Senescence; D018118:Chloride Channels / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism; D003110:Colonic Neoplasms / Q000378:metabolism* / Q000503:physiopathology; D000242:Cyclic AMP / Q000378:metabolism; D004553:Electric Conductivity; D006801:Humans; D013552:Swine; D013997:Time Factors
C492109:CLCA1 protein, human; D018118:Chloride Channels; D002712:Chlorides; D000242:Cyclic AMP; D002118:Calcium