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Covalent capture of kinase-specific phosphopeptides reveals Cdk1-cyclin B substrates.

ABSTRACT: We describe a method for rapid identification of protein kinase substrates. Cdk1 was engineered to accept an ATP analog that allows it to uniquely label its substrates with a bio-orthogonal phosphate analog tag. A highly specific, covalent capture-and-release methodology was developed for rapid purification of tagged peptides derived from labeled substrate proteins. Application of this approach to the discovery of Cdk1-cyclin B substrates yielded identification of >70 substrates and phosphorylation sites. Many of these sites are known to be phosphorylated in vivo, but most of the proteins have not been characterized as Cdk1-cyclin B substrates. This approach has the potential to expand our understanding of kinase-substrate connections in signaling networks.

SUBMITTER: Blethrow JD

PROVIDER: S-EPMC2234163 | biostudies-literature | 2008 Feb

REPOSITORIES: biostudies-literature

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Covalent capture of kinase-specific phosphopeptides reveals Cdk1-cyclin B substrates.

Blethrow Justin D JD Glavy Joseph S JS Morgan David O DO Shokat Kevan M KM

Proceedings of the National Academy of Sciences of the United States of America 20080130 5

We describe a method for rapid identification of protein kinase substrates. Cdk1 was engineered to accept an ATP analog that allows it to uniquely label its substrates with a bio-orthogonal phosphate analog tag. A highly specific, covalent capture-and-release methodology was developed for rapid purification of tagged peptides derived from labeled substrate proteins. Application of this approach to the discovery of Cdk1-cyclin B substrates yielded identification of >70 substrates and phosphorylat ...[more]

PMID: 18234856

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Project description:Simple Summary Cyclin-dependent kinase 1 (CDK1), one of the key regulators of the G2/M checkpoint, is expressed in many cells and plays an important role in cell cycle control. However, CDK1 expression is substantially increased in many tumors of diverse origins and is associated with tumorigenesis. Targeting CDK1 shows promising results for several tumors. However, a systematic and integrative analysis of CDK1 in cancer has not been conducted. The present study aims to use pan-cancer analysis to investigate the relationship, similarities, and differences in genetic and cellular changes associated with CDK1 in various tumors and tumor microenvironments. Our findings elucidate that CDK1 expression increases in more than 20 human tumors and is highly correlated with oncogenic signature gene sets, biological pathways, immune cell infiltration, tumor mutational burden, microsatellite instability, and lower survival rate across multiple tumors. Targeting CDK1 may provide a novel and effective strategy for cancer immunotherapy. Abstract Cyclin-dependent kinase 1 (CDK1) is essential for cell division by regulating the G2/M phase and mitosis. CDK1 overexpression can also promote the development and progression of a variety of cancers. However, the significance of CDK1 in the formation, progression, and prognosis of human pan-cancer remains unclear. In the present study, we used The Cancer Genome Atlas database, Clinical Proteomic Tumor Analysis Consortium, Human Protein Atlas, Genotype-Tissue Expression, and other well-established databases to comprehensively examine CDK1 genetic alterations and gene/protein expression in various cancers and their relationships with the prognosis, immune reactivities, and clinical outcomes for 33 tumor types. Gene set enrichment analysis was also conducted to examine the potential mechanisms of CDK1 in tumorigenesis. The data showed that CDK1 mutation was frequently present in multiple tumors. CDK1 expression was significantly increased in various types of tumors as compared with normal tissues and was associated with poor overall and disease-free survival. In addition, CDK1 expression was significantly correlated with oncogenic genes, proteins, cellular components, myeloid-derived suppressor cell infiltration, ESTMATEScore, and signaling pathways associated with tumor development and progression and tumor microenvironments. These data indicate that CDK1 could serve as a promising biomarker for predicting tumor prognosis and a potential target for cancer treatment.

Dataset Information

Covalent capture of kinase-specific phosphopeptides reveals Cdk1-cyclin B substrates.

Publications

Covalent capture of kinase-specific phosphopeptides reveals Cdk1-cyclin B substrates.

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