Recent studies show that the human parathyroid calcium sensing receptor (CaSR) is expressed in human colon epithelium and functions to regulate epithelial proliferation and differentiation. In this study, we show that the cells of the colon crypt acquire CaSR expression as they differentiate and migrate towards the apex of the crypt. CaSR expression was weak in colon carcinomas with a more-differentiated histologic pattern, whereas CaSR expression was undetectable in less-differentiated tumors. We found that Ca(2+) and/or 1,25(OH)(2)D(3) stimulated CaSR promoter activity and CaSR protein expression in the human colon carcinoma CBS cells, which possessed a functional CaSR. Both agents concomitantly induced a series of changes in the CBS cells that influence proliferation and differentiation, but cellular responses to the two agents were not identical. Ca(2+) strongly induced E-cadherin expression and inhibited the expression of the nuclear transcription factor, TCF4. 1,25(OH)(2)D(3) was weaker in its effect on E-cadherin and was not able to inhibit TCF4 expression. 1,25(OH)(2)D(3) was as strong or stronger than Ca(2+) in its induction of the cyclin-dependent kinase inhibitors, P21 and p27. It is concluded that CaSR may function in the colon to regulate epithelial differentiation and that loss of CaSR expression may be associated with abnormal differentiation and/or malignant progression. Extracellular Ca(2+) and 1,25(OH)(2)D(3) are potential candidates involved in regulating CaSR expression in the colon and the chemopreventive actions of Ca(2+) and 1,25(OH)(2)D(3) in colon cancer may be mediated, in part, through the CaSR.