1alpha,25-Dihydroxyvitamin D3-induced down-regulation of the checkpoint proteins, Chk1 and Claspin, is mediated by the pocket proteins p107 and p130.
A previous cDNA microarray analysis in murine MC3T3-E1 osteoblasts revealed a cluster of genes involved in cell cycle progression that was significantly down-regulated after a single treatment with 1alpha,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] [L. Verlinden, G. Eelen, I. Beullens, M. Van Camp, P. Van Hummelen, K. Engelen, R. Van Hellemont, K. Marchal, B. De Moor, F. Foijer, H. Te Riele, M. Beullens, M. Bollen, C. Mathieu, R. Bouillon, A. Verstuyf, Characterization of the condensin component Cnap1 and protein kinase Melk as novel E2F target genes down-regulated by 1,25-dihydroxyvitamin D3, J. Biol. Chem. 280 (45) (2005) 37319-37330]. Among those genes were the DNA replication and DNA damage checkpoint proteins, Chk1 and Claspin, of which the human homologues were recently shown to be E2F-responsive. Quantitative real-time PCR experiments in 1,25(OH)(2)D(3)-treated MC3T3-E1 cells confirmed the down-regulation observed in the microarray experiment. Moreover, Chk1 and Claspin promoter activities were also reduced after incubation with 1,25(OH)(2)D(3), and this reduction was mediated through the E2F recognition motifs within their promoters because mutation of these motifs almost completely abolished the repressive effect of 1,25(OH)(2)D(3). The antiproliferative effect of 1,25(OH)(2)D(3) as well as its potential to down-regulate the expression of Chk1 and Claspin depended on the pocket proteins p107 and p130 because 1,25(OH)(2)D(3) lost its antiproliferative action and failed to repress these E2F-target genes in p107(-/-);p130(-/-)-cells, but not in pRb(-/-)-cells.
Verlinden
Lieve
L
Laboratorium voor Experimentele Geneeskunde en Endocrinologie, Katholieke Universiteit Leuven, Herestraat 49, 3000 Leuven, Belgium.
Eelen
Guy
G
Van Hellemont
Ruth
R
Engelen
Kristof
K
Beullens
Ine
I
Van Camp
Mark
M
Marchal
Kathleen
K
Mathieu
Chantal
C
Bouillon
Roger
R
Verstuyf
Annemieke
A
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000818:Animals; D002117:Calcitriol / Q000494:pharmacology*; D002460:Cell Line; D049109:Cell Proliferation; D000071877:Checkpoint Kinase 1; D015536:Down-Regulation / Q000187:drug effects*; D051379:Mice; D018345:Mice, Knockout; D011401:Promoter Regions, Genetic / Q000235:genetics; D011494:Protein Kinases / Q000378:metabolism*; D050720:Retinoblastoma-Like Protein p107 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D050717:Retinoblastoma-Like Protein p130 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D050720:Retinoblastoma-Like Protein p107; D050717:Retinoblastoma-Like Protein p130; C488160:claspin protein, mouse; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; C000605869:Chek1 protein, mouse; D002117:Calcitriol