Mammalian target of rapamycin complex 1 signaling opposes the effects of anchorage loss, leading to activation of Cdk4 and Cdc6 stabilization

AbstractWhen deprived of an anchorage to the extracellular matrix, fibroblasts arrest in the G1 phase with inactivation of Cdk4/6 and Cdk2 and destruction of Cdc6, the assembler of prereplicative complexes essential for S phase onset. How cellular anchorages control these kinases and Cdc6 stability is poorly understood. Here, we report that in rat embryonic fibroblasts, activation of mammalian target of rapamycin complex 1 by a Tsc2 mutation or overexpression of a constitutively active mutant Rheb overrides the absence of the anchorage and stabilizes Cdc6 at least partly via activating Cdk4/6 that induces Emi1, an APC/CCdh1 ubiquitin ligase inhibitor.Structured summaryMINT-7890626: cdc27 (uniprotkb:Q4V8A2) physically interacts (MI:0915) with Cyclin-A (uniprotkb:Q6AY13) by anti bait coimmunoprecipitation (MI:0006

Bone Morphogenetic Protein 2-Induced Osteoblast Differentiation Requires Smad-Mediated Down-Regulation of Cdk6

Because a temporal arrest in the G(1) phase of the cell cycle is thought to be a prerequisite for cell differentiation, we investigated cell cycle factors that critically influence the differentiation of mouse osteoblastic MC3T3-E1 cells induced by bone morphogenetic protein 2 (BMP-2), a potent inducer of osteoblast differentiation. Of the G(1) cell cycle factors examined, the expression of cyclin-dependent kinase 6 (Cdk6) was found to be strongly down-regulated by BMP-2/Smads signaling, mainly via transcriptional repression. The enforced expression of Cdk6 blocked BMP-2-induced osteoblast differentiation to various degrees, depending on the level of its overexpression. However, neither BMP-2 treatment nor Cdk6 overexpression significantly affected cell proliferation, suggesting that the inhibitory effect of Cdk6 on cell differentiation was exerted by a mechanism that is largely independent of its cell cycle regulation. These results indicate that Cdk6 is a critical regulator of BMP-2-induced osteoblast differentiation and that its Smads-mediated down-regulation is essential for efficient osteoblast differentiation