Prohibitin is a novel target gene of vitamin D involved in its antiproliferative action in breast cancer cells.
Previously, we showed that N-methyl-N-nitrosourea-transformed MCF12F breast epithelial cells exhibited differential expression of several genes, including up-regulation of prohibitin and elevated sensitivity to a relatively noncalcemic vitamin D analogue, 1alpha-hydroxyvitamin D5 [1alpha(OH)D5]. In this report, we evaluated the functional significance of prohibitin in relation to the cellular response to vitamin D. The in silico screening for putative transcription factor binding sites identified two vitamin D receptor (VDR)/retinoid X receptor binding sites in the 1-kb promoter region of prohibitin. Prohibitin up-regulation by 1alpha(OH)D5 treatment at both transcriptional and translational levels was confirmed by real-time reverse transcription-PCR and Western blot analysis in breast cancer cells, identifying prohibitin as a vitamin D target gene. Confocal microscopic analysis showed that prohibitin was localized in the nuclei of MCF-7 cells and a portion of prohibitin was colocalized with VDR, but direct physical interaction between VDR and prohibitin in cell lysates was not detectable. In MCF-7 cells expressing tetracycline-inducible prohibitin (Tet-On model), the overexpression of prohibitin inhibited cell proliferation and enhanced vitamin D-induced antiproliferative activity. Knockdown of prohibitin was accompanied by increased number of cells incorporating bromodeoxyuridine in the whole population and increased cell distribution in the S phase of cell cycle. In addition, prohibitin level had no significant effect on the vitamin D-induced transactivation of CYP24, a VDR target gene. This is the first report to suggest that prohibitin serves as a novel vitamin D target gene, which is involved in the antiproliferative action of vitamin D without affecting CYP24 transactivation in breast cancer cells.
Peng
Xinjian
X
Illinois Institute of Technology Research Institute, Chicago, Illinois 60616, USA.
Mehta
Rajeshwari
R
Wang
Sheng
S
Chellappan
Srikumar
S
Mehta
Rajendra G
RG
D001483:Base Sequence; D001943:Breast Neoplasms / Q000188:drug therapy* / Q000235:genetics* / Q000378:metabolism; D001973:Bromodeoxyuridine / Q000378:metabolism; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D006801:Humans; D006887:Hydroxycholecalciferols / Q000494:pharmacology*; D008969:Molecular Sequence Data; D000091402:Prohibitins; D011401:Promoter Regions, Genetic; D034622:RNA Interference; D012097:Repressor Proteins / Q000096:biosynthesis / Q000235:genetics*; D016196:S Phase / Q000235:genetics; D013250:Steroid Hydroxylases / Q000096:biosynthesis / Q000235:genetics; D014158:Transcription, Genetic; D015533:Transcriptional Activation / Q000187:drug effects; D065668:Vitamin D3 24-Hydroxylase
C103958:1-hydroxyvitamin D5; D006887:Hydroxycholecalciferols; D000091402:Prohibitins; D012097:Repressor Proteins; D013250:Steroid Hydroxylases; D065668:Vitamin D3 24-Hydroxylase; D001973:Bromodeoxyuridine