Slow Transcription of the 99a/let-7c/125b-2 Cluster Results in Differential MiRNA Expression and Promotes Melanoma Phenotypic Plasticity

Danna Sheinboim, Shivang Parikh, Roma Parikh, Amitai Menuchin, Guy Shapira, Oxana Kapitansky, Nadav Elkoshi, Shmuel Ruppo, Lital Shaham, Tamar Golan, Sharona Elgavish, Yuval Nevo, Rachel E. Bell, Hagar Malcov-Brog, Noam Shomron, Jeffrey W. Taub, Shai Izraeli, Carmit Levy*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

Almost half of the human microRNAs (miRNAs) are encoded in clusters. Although transcribed as a single unit, the levels of individual mature miRNAs often differ. The mechanisms underlying differential biogenesis of clustered miRNAs and the resulting physiological implications are mostly unknown. In this study, we report that the melanoma master transcription regulator MITF regulates the differential expression of the 99a/let-7c/125b-2 cluster by altering the distribution of RNA polymerase II along the cluster. We discovered that MITF interacts with TRIM28, a known inhibitor of RNA polymerase II transcription elongation, at the mIR-let-7c region, resulting in the pausing of RNA polymerase II activity and causing an elevation in mIR-let-7c expression; low levels of RNA polymerase II occupation over miR-99a and miR-125b-2 regions decreases their biogenesis. Furthermore, we showed that this differential expression affects the phenotypic state of melanoma cells. RNA-sequencing analysis of proliferative melanoma cells that express miR-99a and miR-125b mimics revealed a transcriptomic shift toward an invasive phenotype. Conversely, expression of a mIR-let-7c mimic in invasive melanoma cells induced a shift to a more proliferative state. We confirmed direct target genes of these miRNAs, including FGFR3, BAP1, Bcl2, TGFBR1, and CDKN1A. Our study demonstrates an MITF-governed biogenesis mechanism that results in differential expression of clustered 99a/let-7c/125b-2 miRNAs that control melanoma progression.

Original languageEnglish
Pages (from-to)2944-2956.e6
JournalJournal of Investigative Dermatology
Volume141
Issue number12
DOIs
StatePublished - Dec 2021

Bibliographical note

Funding Information:
The authors gratefully acknowledge Meenhard Herlyn and Levi Garraway for supplying melanoma cultures; Karen B. Avraham, Anya Rudnicki, and Ofer Isakov for supplying mouse ear miRNA profiling data; and the Smoler Proteomics Center at the Technion for the mass spectrometry analysis. We gratefully thank Mehdi Khaled for MITF?wild-type, MITF-mutated (MITF-E87R), and FUGW-mCherry-Luciferase plasmids. We thank David Fisher (Massachusetts General Hospital, Boston, MA) for the MITF-human influenza hemagglutinin expression vector, pGL3b-hTRPM1-luciferase plasmid constructs, and C5 mouse anti-MITF mAb. We thank Yanan Yang (Mayo Clinic, Rochester, MN) and Avner Yayon (Weizmann Institute of Science, Rehovot, Israel) for sharing reagents. CL gratefully acknowledges grant support from the European Research Council under the European Union's Horizon 2020 research and innovation program (grant agreement no. 726225), I-CORE, Melanoma Research Alliance (grant no. 402792), David Brown through the Israel Cancer Association, Cancer Biology Research Center of Tel Aviv University, Israel Cancer Research Fund, Fritz Thyssen Stiftung, Marie Curie Career Integration Grants, Dalya Gridinger Fund, Fingerhot Carol and Lionara Fund, and the Shtacher Family award. SI and JWT were supported by grants from the National Cancer Institute of the National Institutes of Health (U10CA098543 and U10CA180886). Conceptualization: CL; Formal Analysis: DS, SP, RP, AM, GS, NS, OK, TG, SR, HM-B, LS, NE, SI, JWT, SE, YN, REB, CL; Funding Acquisition: CL; Investigation: DS, SP, RP, CL; Methodology: DS, AM, CL; Project Administration: CL; Resources: CL; Software: GS, NS, SR, SE, YN, NE, REB; Supervision: CL; Validation: DS, SP, RP, AM, OK, HM-B, LS, SI, JWT; Visualization: DS, SP, RP, CL; Writing - Original Draft Preparation: DS, AM, CL; Writing - Review and Editing: DS, CL

Funding Information:
The authors gratefully acknowledge Meenhard Herlyn and Levi Garraway for supplying melanoma cultures; Karen B. Avraham, Anya Rudnicki, and Ofer Isakov for supplying mouse ear miRNA profiling data; and the Smoler Proteomics Center at the Technion for the mass spectrometry analysis. We gratefully thank Mehdi Khaled for MITF–wild-type, MITF-mutated (MITF-E87R), and FUGW-mCherry-Luciferase plasmids. We thank David Fisher (Massachusetts General Hospital, Boston, MA) for the MITF-human influenza hemagglutinin expression vector, pGL3b-hTRPM1-luciferase plasmid constructs, and C5 mouse anti-MITF mAb. We thank Yanan Yang (Mayo Clinic, Rochester, MN) and Avner Yayon (Weizmann Institute of Science, Rehovot, Israel) for sharing reagents. CL gratefully acknowledges grant support from the European Research Council under the European Union’s Horizon 2020 research and innovation program (grant agreement no. 726225), I-CORE, Melanoma Research Alliance (grant no. 402792), David Brown through the Israel Cancer Association , Cancer Biology Research Center of Tel Aviv University , Israel Cancer Research Fund, Fritz Thyssen Stiftung , Marie Curie Career Integration Grants, Dalya Gridinger Fund, Fingerhot Carol and Lionara Fund, and the Shtacher Family award. SI and JWT were supported by grants from the National Cancer Institute of the National Institutes of Health (U10CA098543 and U10CA180886).

Publisher Copyright:
© 2021 The Authors

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