Background: Tyrosinase-Related Protein 2 (TRP2) is an enzyme involved in melanogenesis, that also exerts proliferative, anti-apoptotic and immunogenic functions in melanoma cells. TRP2 transcription is regulated by the melanocytic master transcription factor MITF. GLI2, a transcription factor that acts downstream of Hedgehog signaling, is also a direct transcriptional target of the TGF-β/SMAD pathway that contributes to melanoma progression and exerts transcriptional antagonistic activities against MITF.
Objectives: To characterize the molecular events responsible for TGF-β and GLI2 repression of TRP2 expression.
Methods: In silico promoter analysis, transient cell transfection experiments with 5'-end TRP2 promoter deletion constructs, chromatin immuno-precipitation, and site-directed promoter mutagenesis were used to dissect the molecular mechanisms of TRP2 gene regulation by TGF-β and GLI2.
Results: We demonstrate that TGF-β and GLI2-specific TRP2 repression involves direct mechanisms that occur in addition to MITF downregulation by TGF-β and GLI2. We identify two functional GLI2 binding sites within the TRP2 promoter that are critical for TGF-β and GLI2 responsiveness, one of them overlapping a CREB binding site. GLI2 and CREB competing for the same cis-element is associated with opposite transcriptional outcome.
Conclusion: Our results further refine the understanding of how TGF-β and GLI2 control the phenotypic plasticity of melanoma cells. In particular, we identify critical GLI2-binding cis-elements within the TRP2 promoter region that allow for its transcriptional repression independently from MITF concomitant downregulation.
Keywords: CREB; Kruppel-like transcription factor GLI2; Melanoma; Microphtalmia transcription factor; TGF-β; Transcriptional control; Tyrosine-related protein 2.
Copyright © 2019 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.