This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.Melanoma is a common human malignancy of increasing incidence and mortality in people overthe age of 45. It is also increasing rapidly in the population under age 35. Overexpression andmislocalization of B-catenin, a cell adhesion and signaling molecule, has been shown to correlatewith severity of melanoma and other malignancies. This occurs through down-regulation of itsadhesive function and an increase in its transcriptional activation function, leading to unregulatedproliferation. Retinoic acid (RA) is known to inhibit the growth rate and invasiveness ofmelanoma cell lines. Retinoids have been shown to regulate B-catenin signaling duringdevelopment and can alter its function in breast cancer cell lines to produce a less malignantphenotype. The interaction of RA and B-catenin in melanoma has not been investigated. Ourhypothesis is that B-catenin transcriptional activity is enhanced in melanoma and that RA willinhibit this activity. The hypothesis will be tested by characterizing 13-catenin expression andfunction in mouse B 16 melanoma cells, nonmalignant mouse melanocytes and in humanmelanoma cell lines derived from progressively more malignant tumors. The effect of RA on Bcateninexpression/function will be determined, including 13-catenin RNA and protein expression,cytoskeletal attachment, intracellular localization and reporter gene transactivation. It is expectedthat even if retinoic acid does not directly affect these measures, there will be important basicinformation derived on the status of g-catenin in melanoma compared to non-malignantmelanocytes. To examine a functional link between 13-catenin action and the melanomaphenotype, constructs expressing constitutively active or dominant negative 13-catenin signalingproteins will b e used. Non-malignant cells will be transfected with active 13-catenin and tested formalignant transformation. Melanoma cells will be transfected with 13-catenin inhibitory proteinsand examined for loss of the malignant phenotype. If it is determined t h at gene transactivation by13-catenin drives a malignant phenotype in melanocytes, analysis by eDNA microarrays will beused to determine the population and pattern of genes induced. The physical interaction of 13-catenin and retinoic acid receptors w i ll be examined. These studies will provide the basis forfuture investigations of mechanisms by which 13-catenin function affects the development andprogression of melanoma.dd%
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