Reduction of Cdc25A contributes to cyclin E1-Cdk2 inhibition at senescence in human mammary epithelial cells

Charanjit Sandhu, Jeffrey Donovan, Nandita Bhattacharya, Martha Stampfer, Peter Worland, Joyce Slingerland

Research output: Contribution to journalArticle

44 Scopus citations

Abstract

Replicative senescence may be an important tumor suppressive mechanism for human cells. We investigated the mechanism of cell cycle arrest at senescence in human mammary epithelial cells (HMECs) that have undergone a period of 'self-selection', and as a consequence exhibit diminished p16(INK4A) levels. As HMECs approached senescence, the proportion of cells with a 2N DNA content increased and that in S phase decreased progressively. Cyclin D1-cdk4, cyclin E-cdk2 and cyclin A-cdk2 activities were not abruptly inhibited, but rather diminished steadily with increasing population age. In contrast to observations in fibroblast, p21(Cip1) was not increased at senescence in HMECs. There was no increase in p27(Kip1) levels nor in KIP association with targets cdks. While p15(INK4B) and its binding to both cdk4 and cdk6 increased with increasing passage, some cyclin D1-bound cdk4 and cdk6 persisted in senescent cells, whose inhibition could not be attributed to p15(INK4B). The inhibition of cyclin E-cdk2 in senescent HMECs was accompanied by increased inhibitory phosphorylation of cdk2, in association with a progressive loss of Cdc25A. Recombinant Cdc25A strongly reactivated cyclin E-cdk2 from senescent HMECs suggesting that reduction of Cdc25A contributes to cyclin E-cdk2 inhibition and G1 arrest at senescence. Although ectopic expression of Cdc25A failed to extend the lifespan of HMECs, the exogenous Cdc25A appeared to lack activity in these cells, since it neither shortened the G1-to-S phase interval nor activated cyclin E-cdk2. In contrast, in the breast cancer-derived MCF-7 line, Cdc25A overexpression increased both cyclin E-cdk2 activity and the S phase fraction. Thus, mechanisms leading to HMEC immortalization may involve not only the re-induction of Cdc25A expression, but also activation of this phosphatase.

Original languageEnglish (US)
Pages (from-to)5314-5323
Number of pages10
JournalOncogene
Volume19
Issue number47
DOIs
StatePublished - Nov 9 2000

Keywords

  • Cdc25A
  • Cell cycle
  • Cyclin E-cdk2
  • HMEC
  • P15(INK4B)
  • Senescence

ASJC Scopus subject areas

  • Molecular Biology
  • Cancer Research
  • Genetics

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