Inhibition of F-Box protein p45(SKP2) expression and stabilization of cyclin-dependent kinase inhibitor p27(KIP1) in vitamin D analog-treated cancer cells.
Treatment of cancer cells with 1,25-dihydroxyvitamin D3 [1,25(OH)(2)D(3)] or its analogs induces growth arrest and expression of the cyclin-dependent kinase inhibitor p27(KIP1). Although 1,25(OH)(2)D(3) transiently enhances p27(kip1) gene transcription in some cells, its effects on p27(KIP1) protein levels are generally more gradual and sustained. This suggests that 1,25(OH)(2)D(3) treatment may be stabilizing p27(KIP1) protein, which is sensitive to modification by the SCF(SKP2) protein ubiquitin ligase and proteosomal degradation. Here, we show that treatment of AT-84 head and neck squamous carcinoma cells with the 1,25(OH)(2)D(3) analog EB1089 increases p27(KIP1) protein levels without significantly affecting expression of its mRNA. EB1089 treatment repressed expression of mRNAs encoding the F-box protein p45(SKP2), a marker of poor head and neck cancer prognosis, and the cyclin kinase subunit CKS1, which is essential for targeting p45(SKP2) to p27(KIP1). This coincided with a reduction of total p45(SKP2) protein, and p45(SKP2) associated with p27(KIP1). Consistent with these findings, turnover of p27(KIP1) protein was strongly inhibited in the presence of EB1089. A similar reduction in p45(SKP2) expression and stabilization of p27(KIP1) protein was observed in 1,25(OH)(2)D(3)-sensitive UF-1 promyelocytic leukemia cells, which also respond by transiently increasing p27(kip1) gene transcription. Our results reveal that 1,25(OH)(2)D(3) analogs increase levels of p27(KIP1) in different cell types by inhibiting expression of SCF(SKP2) subunits and reducing turnover of p27(KIP1) protein.
Lin
Roberto
R
Department of Physiology, McGill University, Montréal, Québec H3G 1Y6, Canada.
Wang
Tian Tian
TT
Miller
Wilson H
WH
White
John H
JH
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D015152:Blotting, Northern; D002117:Calcitriol / Q000031:analogs & derivatives* / Q000494:pharmacology; D002294:Carcinoma, Squamous Cell / Q000378:metabolism / Q000473:pathology; D018797:Cell Cycle Proteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism*; D050760:Cyclin-Dependent Kinase Inhibitor p27; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors*; D004355:Drug Stability; D004791:Enzyme Inhibitors / Q000378:metabolism*; D016193:G1 Phase / Q000187:drug effects; D015870:Gene Expression / Q000187:drug effects; D006258:Head and Neck Neoplasms / Q000378:metabolism / Q000473:pathology; D006801:Humans; D007163:Immunosorbent Techniques; D051379:Mice; D009369:Neoplasms / Q000378:metabolism* / Q000473:pathology; D012333:RNA, Messenger / Q000032:analysis; D016192:Resting Phase, Cell Cycle / Q000187:drug effects; D020133:Reverse Transcriptase Polymerase Chain Reaction; D044786:S-Phase Kinase-Associated Proteins; D014407:Tumor Cells, Cultured; D025521:Tumor Suppressor Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*
D000970:Antineoplastic Agents; C493961:Cdkn1b protein, mouse; D018797:Cell Cycle Proteins; D004791:Enzyme Inhibitors; D012333:RNA, Messenger; D044786:S-Phase Kinase-Associated Proteins; D025521:Tumor Suppressor Proteins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D018844:Cyclin-Dependent Kinases; D002117:Calcitriol; C078903:seocalcitol