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Title: Role of microphthalmia transcription factor (Mitf) in melanoma differentiation

Abstract

We transfected the melanocyte-specific Mitf-M isoform into the aggressive melanoma UISO-Mel-6 cell lines. Our data show that Mitf decreases cell proliferation and results in cells which grow in clusters. By analyzing the expression of the markers of differentiation, we demonstrate that Mitf favored increased expression of tyrosinase and tyrosinase-related protein-1. In addition, Mitf induces Bcl-2 expression following transfection of UISO-Mel-6 cells. We also showed that Mitf gene affects cell-cycle distribution by resting cells preferentially in G2/G1 phase, and inducing the expression of p21 and p27. Moreover, we performed in vivo studies using subcutaneous injection of UISO-Mel-6 and UISO-Mel-6-Mitf in Balb/c nude mice. Our data show that Mitf inhibits tumor growth and decreases Ki67 expression. Tumors induced by UISO-Mel-6 cells were ulcerated and resulted in metastases to liver. None of the mice injected with UISO-Mel-6{sup Mitf+} cells harbored liver metastases. Our results suggest that Mitf is involved in melanoma differentiation and leads to a less aggressive phenotype.

Authors:
 [1];  [2];  [2];  [2];  [2]
  1. Department of Surgical Oncology, University of Illinois at Chicago, 840 South Wood Street, M/C 820, Chicago, IL 60612 (United States). E-mail: fatimal@uic.edu
  2. Department of Surgical Oncology, University of Illinois at Chicago, 840 South Wood Street, M/C 820, Chicago, IL 60612 (United States)
Publication Date:
OSTI Identifier:
20979834
Resource Type:
Journal Article
Resource Relation:
Journal Name: Biochemical and Biophysical Research Communications; Journal Volume: 354; Journal Issue: 3; Other Information: DOI: 10.1016/j.bbrc.2007.01.075; PII: S0006-291X(07)00128-3; Copyright (c) 2007 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; BORON CHLORIDES; CELL CYCLE; CELL PROLIFERATION; GENES; LIVER; MELANOMAS; METASTASES; MICE; MONOCLINIC LATTICES; PHENOTYPE; SUBCUTANEOUS INJECTION; TRANSCRIPTION FACTORS; TYROSINASE

Citation Formats

Lekmine, Fatima, Chang, C.K., Sethakorn, Nan, Das Gupta, Tapas K., and Salti, George I. Role of microphthalmia transcription factor (Mitf) in melanoma differentiation. United States: N. p., 2007. Web. doi:10.1016/j.bbrc.2007.01.075.
Lekmine, Fatima, Chang, C.K., Sethakorn, Nan, Das Gupta, Tapas K., & Salti, George I. Role of microphthalmia transcription factor (Mitf) in melanoma differentiation. United States. doi:10.1016/j.bbrc.2007.01.075.
Lekmine, Fatima, Chang, C.K., Sethakorn, Nan, Das Gupta, Tapas K., and Salti, George I. Fri . "Role of microphthalmia transcription factor (Mitf) in melanoma differentiation". United States. doi:10.1016/j.bbrc.2007.01.075.
@article{osti_20979834,
title = {Role of microphthalmia transcription factor (Mitf) in melanoma differentiation},
author = {Lekmine, Fatima and Chang, C.K. and Sethakorn, Nan and Das Gupta, Tapas K. and Salti, George I.},
abstractNote = {We transfected the melanocyte-specific Mitf-M isoform into the aggressive melanoma UISO-Mel-6 cell lines. Our data show that Mitf decreases cell proliferation and results in cells which grow in clusters. By analyzing the expression of the markers of differentiation, we demonstrate that Mitf favored increased expression of tyrosinase and tyrosinase-related protein-1. In addition, Mitf induces Bcl-2 expression following transfection of UISO-Mel-6 cells. We also showed that Mitf gene affects cell-cycle distribution by resting cells preferentially in G2/G1 phase, and inducing the expression of p21 and p27. Moreover, we performed in vivo studies using subcutaneous injection of UISO-Mel-6 and UISO-Mel-6-Mitf in Balb/c nude mice. Our data show that Mitf inhibits tumor growth and decreases Ki67 expression. Tumors induced by UISO-Mel-6 cells were ulcerated and resulted in metastases to liver. None of the mice injected with UISO-Mel-6{sup Mitf+} cells harbored liver metastases. Our results suggest that Mitf is involved in melanoma differentiation and leads to a less aggressive phenotype.},
doi = {10.1016/j.bbrc.2007.01.075},
journal = {Biochemical and Biophysical Research Communications},
number = 3,
volume = 354,
place = {United States},
year = {Fri Mar 16 00:00:00 EDT 2007},
month = {Fri Mar 16 00:00:00 EDT 2007}
}
  • The microphthalmia-associated transcription factor (MITF) is required for melanocyte development, maintenance of the melanocyte-specific transcription, and survival of melanoma cells. MITF positively regulates expression of more than 25 genes in pigment cells. Recently, it has been demonstrated that expression of several MITF downstream targets requires the SWI/SNF chromatin remodeling complex, which contains one of the two catalytic subunits, Brm or Brg1. Here we show that the expression of MITF itself critically requires active SWI/SNF. In several Brm/Brg1-expressing melanoma cell lines, knockdown of Brg1 severely compromised MITF expression with a concomitant dowregulation of MITF targets and decreased cell proliferation. Although Brmmore » was able to substitute for Brg1 in maintaining MITF expression and melanoma cell proliferation, sequential knockdown of both Brm and Brg1 in 501mel cells abolished proliferation. In Brg1-null SK-MEL-5 melanoma cells, depletion of Brm alone was sufficient to abrogate MITF expression and cell proliferation. Chromatin immunoprecipitation confirmed the binding of Brg1 or Brm to the promoter of MITF. Together these results demonstrate the essential role of SWI/SNF for expression of MITF and suggest that SWI/SNF may be a promissing target in melanoma therapy.« less
  • Highlights: • In human melanocytes, cadmium decreases the expression of MITF-M and tyrosinase and their mRNAs. • In human melanoma cells, cadmium decreases the expression of MITF-M protein and tyrosinase mRNA. • Expression of MITF-H is less sensitive to cadmium toxicity in melanocyte-linage cells. • Cadmium does not decrease the expression of MITF-H in retinal pigment epithelial cells. • MITF-M is the molecular target of cadmium toxicity in melanocytes. - Abstract: Dietary intake of cadmium is inevitable, causing age-related increase in cadmium accumulation in many organs, including hair, choroid and retinal pigment epithelium (RPE). Cadmium has been implicated in themore » pathogenesis of hearing loss and macular degeneration. The functions of cochlea and retina are maintained by melanocytes and RPE, respectively, and the differentiation of these pigment cells is regulated by microphthalmia-associated transcription factor (MITF). In the present study, we explored the potential toxicity of cadmium in the cochlea and retina by using cultured human melanocytes and human RPE cell lines. MITF consists of multiple isoforms, including melanocyte-specific MITF-M and widely expressed MITF-H. Levels of MITF-M protein and its mRNA in human epidermal melanocytes and HMV-II melanoma cells were decreased significantly by cadmium. In parallel with the MITF reduction, mRNA levels of tyrosinase, the key enzyme of melanin biosynthesis that is regulated by MITF-M, were also decreased. In RPE cells, however, the levels of total MITF protein, constituting mainly MITF-H, were not decreased by cadmium. We thus identify MITF-M as the molecular target of cadmium toxicity in melanocytes, thereby accounting for the increased risk of disability from melanocyte malfunction, such as hearing and vision loss among people with elevated cadmium exposure.« less
  • We analyzed alterations in transcription factor binding to specific, known promoter DNA consensus sequences between irradiated and unirradiated radioresistant human melanoma (U1-Mel) cells. The goal of this study was to begin to investigate which transcription factors and DNA-binding sites are responsible for the induction of specific transcripts and proteins after ionizing radiation. Transcription factor binding was observed using DNA band-shift assays and oligonucleotide competition analyses. Confluence-arrested U1-Mel cells were irradiated (4.5 Gy) and harvested at 4 h. Double-stranded oligonucleotides containing known DNA-binding consensus sites for specific transcription factors were used. Increased DNA binding activity after ionizing radiation was noted withmore » oligonucleotides containing the CREB, NF-kB and Sp1 consensus sites. No changes in protein binding to AP-1, AP-2, AP-3, or CTF/NF1, GRE or Oct-1 consensus sequences were noted. X-ray activation of select transcription factors, which bind certain consensus sites in promoters, may cause specific induction or repression of gene transcription. 22 refs., 2 figs.« less
  • Highlights: {yields} Elafin expression is epigenetically silenced in human melanoma cells. {yields} Foxa2 expression in melanoma cells is silenced by promoter hypermethylation. {yields} Foxa2 directs activation of the elafin promoter in vivo. {yields} Foxa2 expression induces apoptosis of melanoma cells via elafin re-expression. -- Abstract: Elafin, a serine protease inhibitor, induces the intrinsic apoptotic pathway in human melanoma cells, where its expression is transcriptionally silenced. However, it remains unknown how the elafin gene is repressed in melanoma cells. We here demonstrate that elafin expression is modulated via epigenetically regulated expression of the transcription factor Foxa2. Treatment of melanoma cells withmore » a DNA methyltransferase inhibitor induced elafin expression, which was specifically responsible for reduced proliferation and increased apoptosis. Suppression of Foxa2 transcription, mediated by DNA hypermethylation in its promoter region, was released in melanoma cells upon treatment with the demethylating agent. Luciferase reporter assays indicated that the Foxa2 binding site in the elafin promoter was critical for the activation of the promoter. Chromatin immunoprecipitation assays further showed that Foxa2 bound to the elafin promoter in vivo. Analyses of melanoma cells with varied levels of Foxa2 revealed a correlated expression between Foxa2 and elafin and the ability of Foxa2 to induce apoptosis. Our results collectively suggest that, in melanoma cells, Foxa2 expression is silenced and therefore elafin is maintained unexpressed to facilitate cell proliferation in the disease melanoma.« less
  • Activation of the aryl hydrocarbon receptor (AhR) transcriptionally induces phase I (cytochrome P450 (CYP) 1A1) and phase II (NAD(P)H quinone oxidoreductase 1 (NQO1) detoxifying enzymes. The effects of the classical and nonclassical AhR ligands on phase I and II enzymes are well studied in human hepatocytes. Additionally, we observed that the proton pump inhibitor, omeprazole (OM), transcriptionally induces CYP1A1 in the human adenocarcinoma cell line, H441 cells via AhR. Whether OM activates AhR and induces the phase II enzyme, NAD(P)H quinone oxidoreductase 1 (NQO1), in fetal primary human pulmonary microvascular endothelial cells (HPMEC) is unknown. Therefore, we tested the hypothesis thatmore » OM will induce NQO1 in HPMEC via the AhR. The concentrations of OM used in our experiments did not result in cytotoxicity. OM activated AhR as evident by increased CYP1A1 mRNA expression. However, contrary to our hypothesis, OM increased NQO1 mRNA and protein via an AhR-independent mechanism as AhR knockdown failed to abrogate OM-mediated increase in NQO1 expression. Interestingly, OM activated Nrf2 as evident by increased phosphoNrf2 (S40) expression in OM-treated compared to vehicle-treated cells. Furthermore, Nrf2 knockdown abrogated OM-mediated increase in NQO1 expression. In conclusion, we provide evidence that OM induces NQO1 via AhR-independent, but Nrf2-dependent mechanisms. - Highlights: • We investigated whether omeprazole induces NQO1 in human fetal lung cells. • Omeprazole induces the phase II enzyme, NQO1, in human fetal lung cells. • AhR deficiency fails to abrogate omeprazole-mediated induction of NQO1. • Omeprazole increases phosphoNrf2 (S40) protein expression in human fetal lung cells. • Nrf2 knockdown abrogates the induction of NQO1 by omeprazole in human lung cells.« less