Cyclin E, a redundant cyclin in breast cancer

Abstract

Cyclin E is an important regulator of cell cycle progression that together with cyclin-dependent kinase (cdk) 2 is crucial for the G 1 ͞S transition during the mammalian cell cycle. Previously, we showed that severe overexpression of cyclin E protein in tumor cells and tissues results in the appearance of lower molecular weight isoforms of cyclin E, which together with cdk2 can form a kinase complex active throughout the cell cycle. In this study, we report that one of the substrates of this constitutively active cyclin E͞cdk2 complex is retinoblastoma susceptibility gene product (pRb) in populations of breast cancer cells and tissues that also overexpress p16. In these tumor cells and tissues, we show that the expression of p16 and pRb is not mutually exclusive. Overexpression of p16 in these cells results in sequestering of cdk4 and cdk6, rendering cyclin D1͞cdk complexes inactive. However, pRb appears to be phosphorylated throughout the cell cycle following an initial lag, revealing a time course similar to phosphorylation of glutathione S-transferase retinoblastoma by cyclin E immunoprecipitates prepared from these synchronized cells. Hence, cyclin E kinase complexes can function redundantly and replace the loss of cyclin Ddependent kinase complexes that functionally inactivate pRb. In addition, the constitutively overexpressed cyclin E is also the predominant cyclin found in p107͞E2F complexes throughout the tumor, but not the normal, cell cycle. These observations suggest that overexpression of cyclin E in tumor cells, which also overexpress p16, can bypass the cyclin D͞cdk4-cdk6͞p16͞pRb feedback loop, providing yet another mechanism by which tumors can gain a growth advantage.Progression through the eukaryotic cell cycle is mediated both positively and negatively by a variety of growth regulatory proteins (1-3). Cyclins and their catalytic cyclin-dependent kinase (cdk) partners act positively to propel a cell through the proliferative cycle (4, 5). Activation of cyclin-cdk complexes results in a cascade of protein phosphorylations that ultimately induce cell cycle progression (1, 4). Although the identity of downstream substrates and effectors of cyclin-cdks remains to be firmly established, it is commonly believed that cdk-mediated phosphorylations manifest cell cycle regulation via inhibition of growth inhibitory signals and activation of proteins necessary for each stage of the cell cycle (6). A putative, well-characterized substrate for the G1 cyclins is retinoblastoma susceptibility gene product (pRb; refs. 7 and 8). This protein is sequentially phosphorylated during the cell cycle presumably through the concerted activity of different cyclin-cdk complexes (9-11). This phosphorylation is required for cell cycle progression, and the hypophosphorylated form of pRb inhibits cell cycle progression by tethering and inactivating transcription factors of the E2F family, which are required for the transactivation of S phase-specific proteins, including dihydrofolate reductase, cyclin A, and thymidyla...

Impact of cyclins E, neutrophil elastase and proteinase 3 expression levels on clinical outcome in primary breast cancer patients

“…5,[24][25][26][27] We showed that CCNE1 and CCNE2 were both associated with RFI in a univariate analysis, whereas CCNE2 was the only significant variable in the multivariate model. However, when CCNE2 was discarded, CCNE1 became significant.…”

Lack of relationship between CDK activity and G1 cyclin expression in breast cancer cells

“…While these are as yet unidenti®ed, likely candidates with demonstrated cyclin E-Cdk2 binding ability include known and potential substrates such as the pRB-related proteins p107 and p130, E2F, adaptor proteins like the p300 transcriptional coactivator, or homologues of suc1-CKS family of CDK binding proteins (Parge et al, 1993;La Thangue, 1994;Sidle et al, 1996;Perkins et al, 1997). Indeed, p107 and E2F are complexed to the overexpressed cyclin E protein in the MDA-MB-157 cell line (Gray-Bablin et al, 1996). Furthermore, recent data from this laboratory indicate that the very high molecular weight, active cyclin ECdk2 complexes in oestrogen-stimulated MCF-7 cells contain p130 (Prall, Rogan, Musgrove, Watts and Sutherland, submitted).…”

“…non-CDK bound) cyclin D1. This cell line overexpresses p16, which likely competed with cyclin D1 for Cdk4, thus preventing cyclin D1-Cdk4 complex formation (Musgrove et al, 1995;Gray-Bablin et al, 1996). MDA-MB-134 cells displayed a very broad distribution of cyclin D1 protein across the fractions, including some of apparent molecular mass 4100 kDa indicating that some of the overexpressed protein is free in this cell line.…”

Pluripotent cell division cycles are driven by ectopic Cdk2, cyclin A/E and E2F activities

“…Precise temporal control of these activities is crucial in establishing transitions from one cell cycle phase to the next and in ensuring that cell cycle events are maintained in the correct order. Uncoupling of Cdk activity from cell cycle progression frequently results in genomic instability and aneuploidy, uncoupling of chromosome replication from segregation or uncontrolled cell proliferation (Gray-Bablin et al, 1996;Mumberg et al, 1996;Haas et al, 1997;Nielsen et al, 1997;Spruck et al, 1999;Harwell et al, 2000). In metazoans, more than one Cdk-cyclin complex is generally required for the transition from one phase to the next.…”