Developmentally regulated inhibition of cell cycle progression by glucocorticoids through repression of cyclin A transcription in primary osteoblast cultures.
Synthetic glucocorticoids (GCs) like dexamethasone (DEX) are effective immunosuppressants indicated for autoimmune and inflammatory diseases. However, they often promote osteoporosis and bone fractures. Here, we investigated the anti-mitogenic effect of GCs in primary osteoblast cultures. DEX did not affect cell cycle progression in confluent (day 2), or early post-confluent cultures. Starting on day 5, however, DEX strongly inhibited the G1/S cell cycle transition. Day 5 also marked the beginning of a ∼2-day commitment stage, during which cultures acquired resistance to the inhibitory effect of DEX on mineralization. Considering the importance of Wnt signaling in osteoblast development, we analyzed the effect of DEX on the Wnt pathway. DEX treatment did not inhibit the TOPGAL Wnt reporter before, but only during and after the commitment stage. However, this inhibition was not associated with repression of neither cyclin D1 nor c-Myc mRNA, well-established Wnt targets regulating cell cycle progression. On the other hand, acute (24-h) and chronic (7 days) DEX treatment significantly reduced both the mRNA and protein levels of cyclin A, another cell cycle regulator. Moreover, cyclin A repression by DEX was not observed before, but only during and after the commitment stage. Using gel shift and reporter assays, we identified an ATF/CREB-binding site critical for the DEX-mediated repression of cyclin A transcription. Furthermore, and similar to cyclin A, Atf4 expression was repressed by DEX only during and after commitment. Our data suggest that GCs attenuate cell cycle progression in osteoblasts in a developmental stage-specific manner by repressing Atf4-dependent cyclin A gene expression.
Gabet
Yankel
Y
Department of Biochemistry & Molecular Biology, Keck School of Medicine, University of Southern California, Los Angeles, California 90033, USA.
Noh
Tommy
T
Lee
Christopher
C
Frenkel
Baruch
B
D051701:Activating Transcription Factor 4 / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn; D002453:Cell Cycle / Q000187:drug effects*; D019070:Cell Lineage / Q000187:drug effects; D002478:Cells, Cultured; D019925:Cyclin A / Q000235:genetics* / Q000378:metabolism; D019938:Cyclin D1 / Q000235:genetics / Q000378:metabolism; D003907:Dexamethasone / Q000494:pharmacology*; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D005938:Glucocorticoids / Q000494:pharmacology*; D051379:Mice; D010006:Osteoblasts / Q000187:drug effects* / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D014158:Transcription, Genetic / Q000187:drug effects*; D051153:Wnt Proteins / Q000378:metabolism
C081452:Atf4 protein, mouse; D019925:Cyclin A; D005938:Glucocorticoids; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; D051153:Wnt Proteins; D019938:Cyclin D1; D051701:Activating Transcription Factor 4; D003907:Dexamethasone