ROLE OF THE P34CDC2 RELATED PITSLRE PROTEIN KINASES

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Description (Adapted from the investigator's abstract): Recurring alterations of chromosome 1 represent the most frequent site of structural chromosome abnormalities across all human solid tumors, including human cutaneous malignant melanoma. In melanoma, breakpoints involving chromosome 1 often accumulate at 1p36. The two genes encoding the PITSLRE protein kinases, Cdc2L1 and Cdc2L2, are localized to chromosome band region 1p36. The PITSLRE protein kinases are part of the p34cdc2 supergene family. Several lines of evidence suggests that some of the PITSLRE protein kinases isoforms are involved in apoptotic signal transduction. We have demonstrated deletion of Cdc2L alleles in melanoma. We can also show that the p110 isoform of PITSLRE is cleaved by caspases to a smaller p46 isoform. The p46 isoform retains kinase activity but appears to change its substrate specificity. There is also a difference in the sensitivity of melanoma cells (with wildtype and mutant Cdc2L alleles) to apoptotic stimuli. Thus, the principal hypothesis to be tested is that the caspase-processed PITSLRE isoform functions as a downstream effector in apoptotic signaling pathways and that disruption of PITSLRE protein kinase function plays a functional role in melanoma development by deregulating controlled cell death. The three specific aims are: (1) To study the timing of Cdc2L gene (PITSLRE) alteration during melanoma tumor progression using PCR-SSCP and direct DNA sequencing. Archival sporadic melanoma cases from various histological stages of melanoma are analyzed for mutations of specific regions within the Cdc2L1 gene. In addition familial melanoma kindreds linked to 1p are also investigated for Cdc2L1 gene alterations. (2) To isolate and characterize the substrate for the caspase processed PITSLRE isoform using yeast two hybrid methodology and (3) To elucidate the biological function of Cdc2L gene inactivation by generating Cdc2L1 deficient mice. An increased understanding of the critical gene alterations that give rise to the development and progression of melanoma may lead to genetic markers for melanoma. Defining the importance of Cdc2L1 alterations may be used to identify interesting subpopulations. Genetic changes of Cdc2L1 also may have prognostic value when considered in tandem with clinical data. Finally, increased understanding of biological role of Cdc2L may lead to new and innovative strategies to treat melanoma.