A nontranscriptional role for Oct4 in the regulation of mitotic entry

Abstract

Rapid progression through the cell cycle and a very short G1 phase are defining characteristics of embryonic stem cells. This distinct cell cycle is driven by a positive feedback loop involving Rb inactivation and reduced oscillations of cyclins and cyclin-dependent kinase (Cdk) activity. In this setting, we inquired how ES cells avoid the potentially deleterious consequences of premature mitotic entry. We found that the pluripotency transcription factor Oct4 (octamer-binding transcription factor 4) plays an unappreciated role in the ES cell cycle by forming a complex with cyclin-Cdk1 and inhibiting Cdk1 activation. Ectopic expression of Oct4 or a mutant lacking transcriptional activity recapitulated delayed mitotic entry in HeLa cells. Reduction of Oct4 levels in ES cells accelerated G2 progression, which led to increased chromosomal missegregation and apoptosis. Our data demonstrate an unexpected nontranscriptional function of Oct4 in the regulation of mitotic entry. Cell cycle progression is driven by the activation and inactivation of cyclin-dependent kinases (Cdks). The G1-to-S phase transition is regulated by cyclins E and A, both of which activate Cdk2 (5-8). The retinoblastoma protein (Rb) maintains the G1 phase by inhibiting the E2F transcription factors. In somatic cells, increased Cdk activity at the end of G1 leads to phosphorylation and inactivation of Rb and increased expression of E2F targets, including cyclins E and A2 (hereafter A) (9, 10). This feedback, in which rising cyclin levels inactivate Rb to promote further increases in cyclins E and A, mediates the transition to S phase. The G2 to mitosis (M) phase transition is governed by activation of Cdk1, primarily through cyclins A and B1 (hereafter B). Cells synthesize cyclin B in interphase, but cyclin B-bound Cdk1 remains inactive due to phosphorylation on residues T14 and Y15 by Myt1/Wee1 kinases (11-13). Once cyclin B reaches a threshold concentration, the Myt1/Wee1 kinases are inhibited, and the CDC25 phosphatases are activated (14), which leads to cyclin B-bound Cdk1 activation and mitotic entry. Cyclin A promotes the early stage of mitosis in particular, and a critical part of this role is activating cyclin B-Cdk1 by weakening the Wee1-mediated inhibition of cyclin B-Cdk1 and promoting the nuclear localization of cyclin B (7,(15)(16)(17)(18)(19)(20).Cell cycle regulation is noticeably altered in ES cells to allow for more rapid cell proliferation. In contrast to somatic cells, Rb is inactive throughout the ES cell cycle due to constitutive phosphorylation (3), which allows for increased expression of cyclins E and A and extended cyclin E/A-Cdk2 activity (4, 21, 22). Moreover, cyclin-dependent kinase inhibitors (CKIs) such as p16, p21, and p27 are absent or expressed at low abundance in ES cells, which further unleash Cdk activity (4, 22-24). The dampened oscillation of Cdk activity is at least partially responsible for the rapid cell cycle progression by reducing the time needed for Cdk activation.Although the features producing a shorte...

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