P452: THE INTERACTOME OF CDK6 IN DIFFERENT ONCOGENIC CONTEXTS

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Background: The mammalian cell cycle is controlled by a complex signaling network that is frequently deregulated in cancer. Cyclin-dependent kinases (CDKs) are the key players in governing cell cycle progression. CDK6 and its close homologue CDK4 are responsible for driving the cell cycle from G1 to S-Phase. In complex with their regulatory subunits, the D-type cyclins, CDK4/6 phosphorylate the retinoblastoma protein family members (Rb) to induce G1 to S-Phase progression and DNA replication. CDK6, and not CDK4, plays an important role in hematopoiesis and is frequently found overexpressed in hematologic malignancies. In the recent years, CDK6 has been shown to regulate transcription by interacting with chromatin in a kinase dependent and independent manner. This function is crucial in leukemia as well as hematopoietic stem cells. Aims: To better understand the transcriptional role of CDK6 in transformed and untransformed cells we will analyze the interactome of CDK6. We hypothesize that different oncogenes lead to the expression of different CDK6-dependent transcriptional programs and to different CDK6-containing protein complexes on DNA level. We aim to identify common and leukemia subtype-specific interaction partners of CDK6 that will represent novel therapeutic vulnerabilities in combination with CDK6 inhibition. Methods: To characterize the interactome of CDK6 we transformed the murine hematopoietic progenitor cell line HPC7 with the BCR-ABL or the MLL-AF9 oncogene that give rise to myeloid leukemia. We isolated CDK6-containing protein complexes by immunoprecipitation followed by mass spectrometry. To extend our data and to get a specific set of interaction partners involved in the transcriptional role of CDK6, we performed a nuclear fractionation prior to immunoprecipitation and mass spectrometry. This enabled us to focus on interaction partners solely in the nucleus. Results: We obtained proteins interacting with CDK6 on a global cell level as well as specific for the nucleus that represent co-factors of CDK6-mediated transcriptional regulation. By intersecting the proteomics data from transformed and untransformed HPC7 cells, we detected interaction partners of CDK6 that are common for all tested leukemia entities, specific for leukemic subtypes and interaction partners which just occur in untransformed cells. As a next step, the determined proteins will be subjected to pathway analysis to reveal CDK6 dependent signaling networks. These proteins might serve as valuable targets for potential combinatorial treatments together with CDK6 inhibition. Summary/Conclusion: Combining our results with next generation sequencing data will allow us deciphering the distinct transcriptional signatures regulated by CDK6 in different leukemia entities. Our insights will provide a better understanding of malignancies with enhanced CDK6 expression and may help to develop novel treatment strategies by interfering with CDK6 interaction partners.

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