Differential expression of a chloride intracellular channel gene, CLIC4, in transforming growth factor-beta1-mediated conversion of fibroblasts to myofibroblasts.
Conversion of fibroblasts into myofibroblasts as mediated by transforming growth factor-beta1 (TGF-beta1) is the most prominent stromal reaction to a number of epithelial lesions including breast cancer. To identify genes which are regulated during this process, the mRNA profiles from primary breast fibroblasts treated with or without TGF-beta1 were analyzed by differential display. Ninety-five differentially expressed transcripts were PCR-cloned and sequenced, and 28 clones were selected for verification in a hybridization array. By use of gene-specific sequence tags, nine differentially expressed genes were identified. One of the clones, identified as CLIC4, a member of the CLIC family of chloride channels, was up-regulated more than 16 times in myofibroblasts and was therefore chosen for further analysis. Using RT-PCR, comparison with CLIC1, CLIC2, CLIC3, and CLIC5 demonstrated that CLIC4 was unique by being up-regulated by TGF-beta1 in myofibroblasts. Immunohistochemistry showed a hitherto unknown, distinctive pattern of CLIC4 expression in breast stroma. Whereas normal breast fibroblasts were devoid of CLIC4 protein expression, myofibroblasts of breast carcinomas were strongly CLIC4-positive. The functional significance of CLIC4 was analyzed in MEF/3T3 fibroblasts by conditional expression using the tetracycline-repressive gene regulation system. In a migration assay, we found that CLIC4 inhibited cell motility by 27%. These results suggest that CLIC4 is differentially regulated in fibroblasts and that its expression contributes to a collective stationary myofibroblast phenotype.
Rønnov-Jessen
Lone
L
Zoophysiological Laboratory, the August Krogh Institute, Copenhagen, Denmark. ironnov-jessen@aki.ku.dk
Villadsen
René
R
Edwards
John C
JC
Petersen
Ole W
OW
D001940:Breast / Q000166:cytology; D002454:Cell Differentiation / Q000187:drug effects / Q000235:genetics; D002465:Cell Movement / Q000235:genetics; D002478:Cells, Cultured; D018118:Chloride Channels / Q000096:biosynthesis / Q000235:genetics*; D005260:Female; D005347:Fibroblasts / Q000166:cytology* / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D016212:Transforming Growth Factor beta / Q000494:pharmacology
C464459:CLIC4 protein, human; D018118:Chloride Channels; D016212:Transforming Growth Factor beta