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Published on: May 17, 2016
The bHLH/Per-Arnt-Sim transcription factor SIM2 regulates muscle transcript myomesin2 via a novel, non-canonical
Susan Woods1, Alexandra Farrall, Carl Procko
1School of Molecular & Biomedical Science (Biochemistry) and the Centre for the Molecular Genetics of Development, University of Adelaide, South Australia 5005, Australia. susan.woods@ucsf.edu
Abstract:
Despite a growing number of descriptive studies that show Single-minded 2 (Sim2) is not only essential for murine survival, but also upregulated in colon, prostate and pancreatic tumours, there is a lack of direct target genes identified for this basic helix-loop-helix/PAS transcription factor. We have performed a set of microarray experiments aimed at identifying genes that are differentially regulated by SIM2, and successfully verified that the Myomesin2 (Myom2) gene is SIM2-responsive. Although SIM2 has been reported to be a transcription repressor, we find that SIM2 induces transcription of Myom2 and activates the Myom2 promoter sequence when co-expressed with the heterodimeric partner protein, ARNT1, in human embryonic kidney cells. Truncation and mutation of the Myom2 promoter sequence, combined with chromatin immunoprecipitation studies in cells, has lead to the delineation of a non-canonical E-box sequence 5'-AACGTG-3' that is bound by SIM2/ARNT1 heterodimers. Interestingly, in immortalized human myoblasts knock down of Sim2 results in increased levels of Myom2 RNA, suggesting that SIM2 is acting as a repressor in these cells and so its activity is likely to be highly context dependent. This is the first report of a direct SIM2/ARNT1 target gene with accompanying analysis of a functional response element.
Insights
Single-minded 2 (Sim2) directly regulates the Myomesin2 (Myom2) gene, identified through microarray analysis. Sim2
Area of Science:
- Molecular Biology
- Genetics
- Cancer Research
Background:
- Single-minded 2 (Sim2) is a transcription factor crucial for survival and implicated in various cancers.
- Direct target genes of Sim2, a basic helix-loop-helix/PAS protein, remain largely unidentified.
- Understanding Sim2's regulatory network is essential for its role in development and disease.
Purpose of the Study:
- To identify direct target genes regulated by the transcription factor Sim2.
- To characterize the functional interaction between Sim2 and its target genes.
- To elucidate the mechanism of Sim2-mediated gene regulation.
Main Methods:
- Microarray experiments to identify differentially expressed genes.
- Co-expression assays in human embryonic kidney cells.
- Promoter analysis, including truncation and mutation studies.
- Chromatin immunoprecipitation (ChIP) assays.
Main Results:
- The Myomesin2 (Myom2) gene was identified as a direct SIM2-responsive gene.
- SIM2, in conjunction with ARNT1, activates the Myom2 promoter in human embryonic kidney cells.
- A novel non-canonical E-box (5'-AACGTG-3') was identified as the binding site for SIM2/ARNT1 heterodimers.
- Sim2 knockdown in human myoblasts led to increased Myom2 RNA levels, indicating context-dependent repression.
Conclusions:
- Myomesin2 is the first identified direct target gene of the SIM2/ARNT1 complex.
- SIM2's transcriptional activity is context-dependent, acting as an activator or repressor.
- The study delineates a functional response element for SIM2/ARNT1 heterodimers.
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