Brk/Protein Tyrosine Kinase 6 Phosphorylates p27^KIP1, Regulating the Activity of Cyclin D–Cyclin-Dependent Kinase 4

Abstract

Cyclin D and cyclin-dependent kinase 4 (cdk4) are overexpressed in a variety of tumors, but their levels are not accurate indicators of oncogenic activity because an accessory factor such as p27^Kip1 is required to assemble this unstable dimer. Additionally, tyrosine (Y) phosphorylation of p27 (pY88) is required to activate cdk4, acting as an “on/off switch.” We identified two SH3 recruitment domains within p27 that modulate pY88, thereby modulating cdk4 activity. Via an SH3-PXXP interaction screen, we identified Brk (breast tumor-related kinase) as a high-affinity p27 kinase. Modulation of Brk in breast cancer cells modulates pY88 and increases resistance to the cdk4 inhibitor PD 0332991. An alternatively spliced form of Brk (Alt Brk) which contains its SH3 domain blocks pY88 and acts as an endogenous cdk4 inhibitor, identifying a potentially targetable regulatory region within p27. Brk is overexpressed in 60% of breast carcinomas, suggesting that this facilitates cell cycle progression by modulating cdk4 through p27 Y phosphorylation. p27 has been considered a tumor suppressor, but our data strengthen the idea that it should also be considered an oncoprotein, responsible for cyclin D-cdk4 activity.

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Supplemental material for this article may be found at http://dx.doi.org/10.1128/MCB.01206-14.

ACKNOWLEDGMENTS

We thank Danielle Joseph, Susan Gottesman, Sabina Hrabetova, and Christopher Roman for critically reading the manuscript. Statistics were verified by the Downstate Computational Core.

This work was supported by grants from the American Cancer Society and the Car-Con Research Fund to S.W.B. and NIH grant DK44525 to A.L.T.