Cyclin D2 activates Cdk2 in preference to Cdk4 in human breast epithelial cells

Abstract

To investigate the possibility of diering roles for cyclins D1 and D2 in breast epithelial cells, we examined the expression, cell cycle regulation and activity of these two G 1 cyclins in both 184 normal breast epithelial cells and T-47D breast cancer cells. Synchronisation studies in 184 cells demonstrated that cyclin D1 and cyclin D2 were dierentially regulated during G 1 , with cyclin D2 abundance increasing by 3.7-fold but only small changes in cyclin D1 abundance observed. The functional consequences of increased cyclin D2 expression were examined in T-47D cells, which express no detectable cyclin D2. Induced expression of cyclin D2 resulted in increases in cyclin E expression, pRB phosphorylation and the percentage of cells in S-phase, while constitutive expression resulted in a consistent trend toward reduced dependence on serum for continued proliferation. Thus, cyclin D2 is a positive regulator of G 1 progression in breast cells analogous to the well-documented eects of cyclin D1. Indeed, equimolar concentrations of inducible cyclin D1 and D2 resulted in quantitatively similar cell cycle eects. Marked divergence was found, however, in the CDKs activated by the two cyclins in breast epithelial cells. Cyclin D2 complexes contained a higher Cdk2/Cdk4 ratio than cyclin D1 complexes. The cyclin D2-associated kinase activity was largely inhibited by Cdk2-speci®c inhibitors and could phosphorylate histone H1, a substrate for Cdk2 but not for Cdk4 and Cdk6. Therefore, cyclin D2 preferentially activated Cdk2 in breast epithelial cells. In contrast, Cdk4 and Cdk6 were predominantly responsible for cyclin D1-associated kinase activity as previously reported. Thus, although cyclins D1 and D2 elicited similar eects on breast epithelial cell cycle progression they appeared to achieve this end via activation of dierent CDKs. This is the ®rst evidence of cyclin D2 activating Cdk2 in mammalian cells thus providing further evidence that D-type cyclins are not necessarily redundant.

Cellular responses to ErbB-2 overexpression in human mammary luminal epithelial cells: comparison of mRNA and protein expression

“…Although cyclins are generally considered as positive mediators of mitogen-induced cell cycle progression, it has been shown that loss of CCND2 expression occurs in primary breast tumours (Evron et al, 2001). Despite this, expression of CCND2 can reduce the dependence on serum for proliferation of breast epithelial cells, and promote cell cycle progression through activation of cdk2 (Sweeney et al, 1997). Thus, a combination of factors appears to drive ErbB-2-dependent cell cycle progression and hence tumorigenic potential in ErbB-2-overexpressing luminal epithelial cells.…”

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

“…Densitometric analysis of autoradiographs was performed using a Molecular Dynamics (Sunnyvale, CA) Personal Densitometer SL and IP LabGel analysis software (Signal Analytics, VA). Quantitation of protein levels by this method was linear over the range of intensities measured see also Sweeney et al, 1997). Relationships between expression levels were determined by simple regression analysis using the software program StatView SE+Graphics (Abacus Concept Inc.).…”

CDC25A Levels Determine the Balance of Proliferation and Checkpoint Response

“…51 Interestingly, Cyclin D2 seems to have a unique preference for CDK binding among the D-type cyclin family. 52 Unlike Cyclin D1, Cyclin D2 can preferentially bind to CDK2 under some cellular contexts. Although it has not been tested, Cyclin D2/CDK2 might be relatively insensitive to the control by CDC25A.…”