(6) Molecular mechanisms explaining premalignant changes in the etiology of melanoma

Signaling pathways generate an enormous variety of cellular outcomes, especially depending upon which receptor-mediated pathway becomes activated, which could be resultant from previous experiences that left lasting epigenetic marks on the responding cells. Similarly, the mitogenic lipid, lysophosphatidic acid, is increased in wound healing responses in normal blister skin and mediates proliferation among a large variety of cell types. In melanocytes, lysophosphatidic acid inhibits proliferation in cells lacking expression of the LPA3 receptor; however, we hypothesize that epigenetic up-regulation of the LPA3 receptor provides melanocytes with a significant advantage by mediating signals of enhanced viability and subsequent atypical cell proliferation in response to lysophosphatidic acid. The goal of this application is to establish that molecular signaling via the lysophosphatidic acid-autotaxinLPA3 receptor axis in cutaneous melanocytes is the fundamental, underlying regulatory mechanism that drives this cell population to progress to cancer with higher frequency. We hypothesize that the LPA3 receptor activation mediates the expression of previously absent miRNA and proteins that are fundamental to tumorigenesis due to their stem cell-promoting pathways and metabolic changes that facilitate pre-malignant and malignant processes. In the first aim of this application, we will definitively demonstrate that wild-type, functional LPA3 receptor is required for the appearance of miR-122 expression and significantly enhances protein levels of Notch 1, β-catenin and fibronectin as well as proliferation and viability. We will also determine whether the LPA3 receptor and subsequent expression and exportation of miR-122 results in a shift in metabolism, which contributes to malignant progression. In the second aim, we will perform a long-term observation of LPA3 receptor-expressing melanocytes using an in vivo orthotopic model with engineered 3D spheroid xenografts, which will also contain autotaxin, tumor-promoting fibroblasts and keratinocytes with and without continuous lysophosphatidic acid secretion. Subsequently, pathological analysis of viable, proliferative, engrafted, xenograft specimens will determine whether the tissue appearance has architecture consistent with pre-malignant lesions. The results of our investigation will establish that the lysophosphatidic acid signaling axis is the key component that initiates underlying advantageous molecular changes and proliferative mechanisms that cause subpopulations of melanocytes to progress to melanoma with higher frequency.