LAUSR

Cyclin-dependent kinase 1 (CDK1) is a serine/threonine kinase that drives the G2/M phase of the cell cycle, and increased CDK1 kinase activity is a frequent feature of several tumor types. In the current study, we have used the National Cancer Institute9s Clinical Proteomic Tumor Analysis Consortium (CPTAC) datasets to identify potential target phosphorylation sites associated with CDK1 activity in breast and lung adenocarcinoma and to experimentally study some of their biologic effects. Among the global protein phosphorylation sites detected in CPTAC, 80% and 81% of the identified phosphorylation sites in breast cancer and lung adenocarcinoma were found to correlate well with the level of an activating phosphorylation site in CDK1, t161. Positive correlations (r > 0.6) were present in most paired samples of tumor tissue and normal tissue. In addition, 42% and 40% of all phosphorylation sites in both of these tumor types were found to contain the minimum CDK1 consensus motif Ser/Thr-Pro (S/T-P). This result suggests that many of the sites with this motif are directly phosphorylated by CDK1, while most of the sites that lack this sequence are likely to be phosphorylated by other kinases. In both tumor types, increased phosphorylation of proteins in several signaling pathways, including those for glycolysis and the cell cycle, were found to be associated with phosphorylation of CDK1 t161. To study a possible direct effect of CDK1-dependent phosphorylation on a protein involved in cell cycle progression, we selected Epithelial Cell Transforming 2 (ECT2), which is a Rho family-specific GEF (guanine nucleotide exchange factor) with a role in G2/M. In agreement with the proteomic data, mass spectrometry analysis showed elevated ECT2 phosphorylation on T359, S367, T373, S376 and S861 in the G2/M phase that was sensitive to treatment with a CDK1 inhibitor. Biological experiments confirmed that phosphorylation of these sites enhanced the pro-oncogenic and GEF activity of ECT2. Additional experimental mechanistic analysis is needed to demonstrate whether specific phosphorylations are attributable to direct or indirect effects of CDK1. Citation Format: Dunrui Wang, Xiaolan Qian, Sarah Eng, Lisa Jenkins, Marian Durkin, Douglas Lowy. A high proportion of protein phosphorylation sites identified in CPTAC breast cancer and lung adenocarcinoma may result from direct phosphorylation by CDK1 [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2137.