Year: 2013
Type: article
Abstract: Cell identity is dictated by specific programmes of gene expression driven by transcription factors that decipher inputs from signal transduction pathways to up- or downregulate their repertoire of target genes. In the melanocyte lineage, identity is initially conferred on melanoblasts exiting the neural crest by the expression of the Microphthalmia-associated transcription factor MITF that is required for melanoblast survival; MITF-null mice exhibit a complete lack of neural crest-derived pigment cells, and the retinal pigment epithelium fails to differentiate leading to the small-eye phenotype that gives the MITF gene its name. A similar phenotype is observed in zebrafish where the nacre mutation in the mitfa gene leaves fish devoid of neural crest-derived melanophores, though in contrast to mice there is no apparent eye phenotype associated with either loss of mitfa or a second mitf gene, mitfb, alone or in combination. The role of MITF in the melanocyte lineage extends well beyond promoting melanoblast survival. It controls expression of differentiation-associated genes required for melanosome biogenesis and intracellular transport (Cheli et al., 2010), as well as metabolism (Haq et al., 2013), and also suppresses melanoma senescence (Giuliano et al., 2010). Importantly, MITF plays both a positive and negative role in cell proliferation, with the pro-proliferative activity of MITF causing it to be termed a lineage-survival oncogene (Garraway et al., 2005). This apparent paradox may be explained by the rheostat model (Carreira et al., 2006) in which low levels of functional MITF characterize G1 arrested melanocyte stem cells or invasive melanoma stem-like cells, whereas MITF expression promotes proliferation, or at elevated levels a G1 arrest associated with differentiation. In terms of proliferation therefore, MITF might be regarded as a ‘Goldilocks’ factor, where proliferation is blocked if MITF activity is too high or too low, and cell division stimulated only if MITF activity is ‘just right’. The rheostat model for MITF function is useful in that it predicts that simply up- or downregulating MITF would promote switching between three phenotypic states important for both melanocyte development as well as melanoma: invasive and stem-like, proliferative, and differentiated (Hoek and Goding, 2010). However, most, though not all, experiments have been performed in melanoma cells in culture with siRNA-mediated depletion of MITF used to determine its role in promoting cell division, and overexpression its function in driving a differentiated phenotype. What has been missing is a clear demonstration that manipulating MITF levels would drive phenotype-switching in vivo. Now, work from Lister et al. describes the use of conditional inactivation of MITF on melanoma genesis and maintenance in vivo in zebrafish. At less that 26°C the hypomorphic mitfavc7 allele, characterized by a mutation in a splice donor for inton 6, gives fish with a reduced number of pigment cells, and almost no melanophores at temperatures greater than 28°C. The ability to reduce MITF activity simply by altering the water temperature gave the authors a unique opportunity to explore the consequences of changing MITF activity in vivo in fish bearing a BRAFV600E mutation that are predisposed to melanoma. As expected, at any temperature, fish with WT mitfa with or without BRAFV600E failed to develop melanoma, and at the restrictive temperature of >28°C, fish bearing the mitfavc7 allele failed to develop pigment cells in the presence or absence of BRAFV600E. Strikingly, however, at <26°C, mitfavc7 fish with the BRAFV600E mutation develop nevi of which around 27% progress to melanoma with histopathological characteristics of superficial spreading melanoma. This incidence is comparable to fish bearing an inactivating p53 mutation in combination with BRAFV600E. Unlike the melanomas found in p53 mutant/BRAFV600E fish, those arising in the BRAFV600/mitfavc7 background exhibited infiltration of heavily pigmented melanomacrophages. Interestingly, although the BRAFV600/mitfavc7 melanomas exhibited a reduced mitotic index compared with those found in p53mutant/BRAFV600E fish, expression of MITF target genes implicated in proliferation (cdk2, p16, p21, hif1a, bcl2) was not significantly different. However, expression of the pigmentation-associated genes tyr and dct was lower, and that of met higher, in the BRAFV600/mitfavc7 melanomas, indicating that not all target genes are regulated by MITF in the same way. Having established that reducing MITF activity renders the fish more susceptible to the pro-proliferative effects of BRAFV600, the authors then went on to ask whether MITF was necessary for the maintenance of pre-established melanomas. In this experiment, BRAFV600/mitfavc7 fish were reared at <26°C to enable melanomas to develop and then shifted to 32°C. Within 2 weeks of the temperature shift, 8/12 melanomas had markedly regressed with high levels of caspase activity characteristic of apoptosis, while no effect of the temperature shift was observed on melanomas in the control p53 mutant/BRAFV600E fish. By 2 months, 12/15 melanomas had regressed with six fish exhibiting apparent complete regression. These data strongly implicate MITF, not just in promoting melanoma proliferation but also in tumour maintenance. Interestingly, by shifting the BRAFV600/mitfavc7 fish exhibiting apparent complete regression back to <26°C, melanomas recurred and repopulated the original site, either because of incomplete regression or possibly because the loss of functional MITF at 32°C led not only to apoptosis and regression, but also to the generation of a population of MITF-negative stem cells that then regenerated the tumour on shifting back to 26°C. “What has been missing, is a clear demonstration that manipulating MITF levels would drive phenotype switching in vivo.”
Source: Pigment Cell & Melanoma Research
DOI: 10.1111/pcmr.12177
Author: Colin R. Goding
Language: English
Topic: Melanoma and MAPK Pathways
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: bronze
OpenAlex ID: W2060947923