Transcriptional Regulation of the Melanoma Prognostic Marker Melastatin (TRPM1) by MITF in Melanocytes and Melanoma

Abstract

Determining the metastatic potential of intermediate thickness lesions remains a major challenge in the management of melanoma. Clinical studies have demonstrated that expression of melastatin/TRPM1 strongly predicts nonmetastatic propensity and correlates with improved outcome, leading to a national cooperative prospective study, which is ongoing currently. Similarly, the melanocytic markers MLANA/MART1 and MITF also have been shown to lose relative expression during melanoma progression. Recent studies have revealed that MITF, an essential transcription factor for melanocyte development, directly regulates expression of MLANA. This prompted examination of whether MITF also might transcriptionally regulate TRPM1 expression. The TRPM1 promoter contains multiple MITF consensus binding elements that were seen by chromatin immunoprecipitation to be occupied by endogenous MITF within melanoma cells. Endogenous TRPM1 expression responded strongly to MITF up-or down-regulation, as did TRPM1 promoter-driven reporters. In addition, MITF and TRPM1 mRNA levels were correlated tightly across a series of human melanoma cell lines. Mice homozygously mutated in MITF showed a dramatic decrease in TRPM1 expression. Finally, the slope of TRPM1 induction by MITF was particularly steep compared with other MITF target genes, suggesting it is a sensitive indicator of MITF expression and correspondingly of melanocytic differentiation. These studies identify MITF as a major transcriptional regulator of TRPM1 and suggest that its prognostic value may be linked to MITF-mediated regulation of cellular differentiation.

TRPM1 (Melastatin-1/MLSN1) mRNA expression in Spitz nevi and nodular melanomas

“…Ulceration was identified in 1 of 95 Spitz nevi and in 14 of the 33 nodular melanomas. Mitotic activity was identified in 22 of 95 of the [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17]. Of the 33 patients with nodular malignant melanoma, 17 did not experience metastasis, 5 patients were alive but had suffered metastases, and 11 patients died from complications related to metastatic melanoma.…”

Regulation of MITF stability by the USP13 deubiquitinase

“…HUBC9 was initially suggested to be the E3 ligase responsible for MITF ubiquitination 18 . However, based on subsequent evidence that MITF is subject to SUMO modification, an activity strongly associated with HUBC9, it is more likely involved in that modification 19,22 . Further studies on the precise amino-acid targets of ubiquitination will also permit further mutational analyses to test stability influences on MITF activity.…”

“…In its role as a nuclear transcriptional factor, MITF is involved in multiple processes via its regulation of different transcriptional target genes 19,31,32 . The expression of the MITF target gene TRPM1 (ref.…”

“…To this end, we isolated total RNA from 23 melanoma cell lines and 4 primary melanocyte lines from different donors, and performed real-time qRT-PCR to detect the mRNA levels of usp13 and Trpm1 (a known MITF target gene 19 ), using Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as internal reference. As shown in Figure 5, expression levels of usp13 and Trpm1 correlated (r = 0.999 for melanocyte group, r = 0.997 for melanoma cell group).…”

“…luciferase reporter that was previously shown to be regulated by MITF 19 together with shUSP13 (313) or control shRNA. Twenty-four hours after transfection, cells were treated with DMSO or Forskolin, a cAMP agonist that upregulates expression of MITF and mimics the action of melanocyte stimulating hormone 7 .…”