PAX3 Expression in Normal Skin Melanocytes and Melanocytic Lesions (Naevi and Melanomas) Sandra Medic1, Mel Ziman1,2* 1 School of Exercise, Biomedical and Health Sciences, Edith Cowan University, Perth, Western Australia, Australia, 2 School of Pathology and Laboratory Medicine, University of Western Australia, Perth, Western Australia, Australia Abstract Background: Cutaneous Malignant Melanoma is an aggressive form of skin cancer, arising in cutaneous melanocytes. The transcription factor PAX3 regulates melanocyte specification from neural crest cells during development but expression in differentiated melanocytes is uncertain. By contrast it is frequently found in melanomas and naevi and is a marker for melanoma staging and detection. In this study we analysed the expression of PAX3 across the spectrum of melanocytic cells, from normal melanocytes to cells of benign and malignant lesions to better assess its function in these various tissues. Pax3 and PAX3 (italicized) refer to the mouse and human gene, respectively; whereas Pax3 and PAX3 (non-italicized) refer to the corresponding mouse and human protein. Methodology and Principal Findings: PAX3 expression was analysed by immunohistochemistry and qRT-PCR. Immunofluorescence was used for co-expression with differentiation, migration and survival markers. As expected PAX3 expression was observed in naevi and melanoma cells. It was also found in melanocytes of normal skin where it co- expressed with melanocyte markers, MITF and MLANA. Co-expression with its downstream target, antiapoptotic factor BCL2L1 confirms PAX3 as a cell survival regulator. PAX3 was also co-expressed with melanoma cell migration marker MCAM in dermal naevi and melanoma cell nests, but this downstream target of PAX3 was not present in normal epidermal melanocytes, suggesting differential roles for PAX3 in normal epidermal melanocytes and melanoma cells. Most interestingly, a proportion of PAX3-positive epidermal melanocytes in normal skin show HES1 and Ki67 co-expression, indicating their less differentiated proliferative phenotype. Conclusions and Significance: Our results suggest that a previously identified role for PAX3, that of regulator of an undifferentiated plastic state, may operate in melanocytes of normal skin. This role, possibly required for cellular response to environmental stimuli, may contribute to formation and development of melanocytic lesions in which PAX3 expression is prominent. Citation: Medic S, Ziman M (2010) PAX3 Expression in Normal Skin Melanocytes and Melanocytic Lesions (Naevi and Melanomas). PLoS ONE 5(4): e9977. doi:10.1371/journal.pone.0009977 Editor: H. Peter Soyer, The University of Queensland, Australia Received December 18, 2009; Accepted March 4, 2010; Published April 22, 2010 Copyright: 2010 Medic, Ziman. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This project was partially funded by the Cancer Council of Western Australia (http://www.cancerwa.asn.au/). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: m.ziman@ecu.edu.au Introduction Cutaneous Malignant Melanoma is a highly aggressive form of skin cancer, thought to be derived from cutaneous melanocytes or melanoblasts. Its aggressiveness is attributed to frequent metastasis and high drug resistance. Intensive research into the mechanisms regulating melanoma tumourigenesis has included investigation of the factors and pathways of normal melanocyte development and function. One key factor that regulates melanocyte development from neural crest derived progenitor cells is the transcriptional factor PAX3. PAX3 is very frequently expressed in melanomas and naevi [1–6]. In fact, PAX3 has been identified as a significant marker for melanoma staging [7,8] and for detection of circulating melanoma cells [7]. It has also been identified as an immunogenic protein in melanomas [9–11], with several epitopes able to induce the host’s immune response - stimulation of the immune response against PAX3- expressing tumour cells results in tumour growth suppression [9,11]. PAX3 is a member of the PAX family of transcription factors which are highly conserved throughout phylogeny. All play a crucial role in embryogenesis but are also implicated in tumourigenesis [12–18]. Although PAX3 is recognised as a key embryonic regulator of melanocyte specification and development, its expression and function in differentiated epidermal melanocytes of adult human skin is largely unexplored and its role in melanoma remains unclear [19]. Pax3 starts its expression in early embryos, in neural crest precursor cells that later give rise to melanocyte precursors, melanoblasts [19–24]. Here the main function of Pax3 besides neural crest specification is regulation of melanoblast survival and proliferation [23,25]. Together with SRY (sex determining region Y)-box 10 (Sox10), Pax3 regulates expression of key melanocyte specification factor Microphthalmia-associated transcription factor (Mitf). It has been suggested that Pax3 controls a ‘‘nodal point’’ in melanocyte differentiation; it simultaneously activates Mitf and represses Dopachrome-tautomerase (Dct) transcription thus blocking PLoS ONE | www.plosone.org 1 April 2010 | Volume 5 | Issue 4 | e9977