Altered p27^Kip1 Phosphorylation, Localization, and Function in Human Epithelial Cells Resistant to Transforming Growth Factor β-Mediated G₁ Arrest

Abstract

p27Kip1 is an important effector of G1 arrest by transforming growth factor β (TGF-β). Investigations in a human mammary epithelial cell (HMEC) model, including cells that are sensitive (184S) and resistant (184A1L5R) to G1 arrest by TGF-β, revealed aberrant p27 regulation in the resistant cells. Cyclin E1-cyclin-dependent kinase 2 (cdk2) and cyclin A-cdk2 activities were increased, and p27-associated kinase activity was detected in 184A1L5R cells. p27 from 184A1L5R cells was localized to both nucleus and cytoplasm, showed an altered profile of phosphoisoforms, and had a reduced ability to bind and inhibit cyclin E1-cdk2 in vitro when compared to p27 from the sensitive 184S cells. In proliferating 184A1L5R cells, more p27 was associated with cyclin D1-cdk4 complexes than in 184S. While TGF-β inhibited the formation of cyclin D1-cdk4-p27 complexes in 184S cells, it did not inhibit the assembly of cyclin D1-cdk4-p27 complexes in the resistant 184A1L5R cells. p27 phosphorylation changed during cell cycle progression, with cyclin E1-bound p27 in G0 showing a different phosphorylation pattern from that of cyclin D1-bound p27 in mid-G1. These data suggest a model in which TGF-β modulates p27 phosphorylation from its cyclin D1-bound assembly phosphoform to an alternate form that binds tightly to inhibit cyclin E1-cdk2. Altered phosphorylation of p27 in the resistant 184A1L5R cells may favor the binding of p27 to cyclin D1-cdk4 and prevent its accumulation in cyclin E1-cdk2 in response to TGF-β.

Non-malignant and Tumor-derived Cells Differ in Their Requirement for p27Kip1 in Transforming Growth Factor-β-mediated G1 Arrest

“…Our previous work showed that the acquisition of TGF-␤ resistance in human mammary epithelial cells was associated with altered phosphorylation, altered Cdk inhibitory activity, and cytoplasmic mislocalization of the p27 protein (15). Although p27 gene mutations are rare in human tumors, increased proteasomal degradation of p27 is observed in a number of cancers, including breast, colon, and prostate, and the reduced p27 levels are associated with poor patient prognosis (reviewed in Refs.…”

The Akt/FoxO1/p27 pathway mediates the proliferative action of liraglutide in β cells

“…The first role is the inhibition of cyclin E-cdk2. The second is that p27 facilitates assembly, activation and nuclear localization of cyclin D-cdk complexes in early G1 phase (5,6). Therefore, inactivation of p27 results in cell proliferation.…”

Regulation of p27Kip1by mitogen-induced tyrosine phosphorylation

“…6 Rather complex patterns of phosphorylation were revealed in quiescent and proliferating cells by isoelectric focusing using two-dimensional electrophoresis. [35][36][37] The pattern of p27 phosphorylation differs between cell types and changes during cell cycle progression, where a significant increase in p27 phosphorylation correlates with cell cycle entry. 35,37 The β-catenin-TCF.…”