BRCA1-mediated G2/M cell cycle arrest requires ERK1/2 kinase activation

Abstract

Germline mutations in the BRCA1 gene are associated with an increased susceptibility to the development of breast and ovarian cancers. Evidence suggests that BRCA1 protein plays a key role in mediating DNA damage-induced checkpoint responses. Several studies have shown that ectopic expression of BRCA1 in human cells can trigger cellular responses similar to those induced by DNA damage, including G2/M cell cycle arrest and apoptosis. While the effects of ectopic BRCA1 expression on the G2/M transition and apoptosis have been extensively studied, the factors that dictate the balance between these two responses remain poorly understood. We have recently shown that ectopic expression of BRCA1 in MCF-7 human breast cancer cells resulted in activation of extracellular signal-regulated protein kinase 1 and 2 (ERK1/2) and G2/M cell cycle arrest. Furthermore, inhibition of BRCA1-induced ERK1/2 activation using mitogen-activated protein kinase kinase 1 and 2 (MEK1/2)-specific inhibitors resulted in increased apoptosis, suggesting a potential role of ERK1/2 kinases in BRCA1-mediated G2/M checkpoint response. In this study, we assessed the role of ERK1/2 kinases in the regulation of BRCA1-mediated G2/M cell cycle arrest. Results indicate that BRCA1-induced G2/M cell cycle arrest and ERK1/2 activation correlate with changes in the level and/or activity of several key regulators of the G2/M checkpoint, including activation of Chk1 and Wee1 kinases, induction of 14-3-3, and down-regulation of Cdc25C. Furthermore, inhibition of ERK1/2 kinases using MEK1/2-specific inhibitors results in a marked attenuation of the BRCA1-induced G2/M arrest. Biochemical studies established that ERK1/2 inhibition abolished the effects of BRCA1 on components of the G2/M checkpoint, including regulation of Cdc25C expression and activation of Wee1 and Chk1 kinases. These results implicate a critical role of ERK1/2 signaling in the regulation of BRCA1 function on controlling the G2/M checkpoint responses.

p14ARF Triggers G2 Arrest Through ERK-Mediated Cdc25C Phosphorylation, Ubiquitination and Proteasomal Degradation

“…In keeping with these results, it was recently shown that BRCA1 destabilized Cdc25C by a mechanism requiring ERK1/2 activation and proteasomal function. 34 Furthermore, our data are reminiscent of a previously described model in which the stability of Cdc25A, another member of the Cdc25 family, is regulated through phosphorylation by CHK1. 41,42 It is presently well-known that phosphorylation of Cdc25C at serine 216 induces 14-3-3 binding, cytosolic retention and further G 2 /M checkpoint in response to DNA damage.…”

“…Downregulation of Cdc25C has been previously reported in response to UVB exposure, 38 γ radiations, 39 BRCA1 expression 34 or infection by adenovirus 40 and related either with a decrease of transcription 39 or a proteasomal degradation. 22,34,40 Interestingly, Singh and colleagues recently proposed that serine 216 could control Cdc25C stability. 22 Consistent with such idea, we demonstrate that phosphorylation at serine 216 by active ERK1/2 targets Cdc25C for ubiquitination and further proteasomal degradation in response to p14 ARF .…”

“…Interestingly, such involvement of the ERK1/2 signaling pathway in the control of G 2 /M checkpoint was recently reported in response to the tumor suppressor BRCA1. 34 The phosphatase Cdc25C is primarily involved in the regulation of mitotic entry through activation of the cyclin-dependent kinase Cdc2/Cyclin B1. 35 Cdc25C is tightly regulated at transcriptional as well as post-transcriptional levels, the latter involving a complex set of activatory/ inhibitory phosphorylations.…”

MAPK Pathway Activation Delays G2/M Progression by Destabilizing Cdc25B

“…There is contradictory evidence of a role for MAPK signaling in G 2 and mitosis in mammalian cells (5,6,8), but recent evidence suggests that some of this maybe attributed to cell lineage-dependent differences (44). MAPK signaling has also been implicated in the G 2 arrest associated with overexpression of BRCA1 and p14ARF, although these mechanisms involved Chk1, differentiating them from the present study (45,46).…”

AKT-independent Phosphorylation of TSC2 and Activation of mTOR and Ribosomal Protein S6 Kinase Signaling by Prostaglandin F2α

“…In the Ad.dnMEK1 vector, the MEK1 cDNA has been altered and two crucial serine residues located in the catalytic domain (Ser 217 and Ser 221 ) were replaced by alanines. The resulting MEK1 mutant has dominant negative activity and can be used to block the activation of ERK1/2 by wild-type MEK1 as described (30).…”