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Updated: Apr 16, 2026
A Reporter Assay to Analyze Intronic microRNA Maturation in Mammalian Cells
Published on: June 16, 2022
Experimental verification of a conserved intronic microRNA located in the human TrkC gene with a cell type-dependent
Sadat Dokanehiifard1, Bahram M Soltani, Sepideh Parsi
1Department of Molecular Genetics, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran.
Abstract:
Tropomyosin receptor kinase C (TrkC) is involved in cell survival, apoptosis induction and tumorigenesis. We hypothesized that, similar to p75(NTR) receptor, some of the diverse functions of TrkC could be mediated by a microRNA (miRNA) embedded within the gene. Here, we experimentally verified the expression and processing of two bioinformatically predicted miRNAs named TrkC-miR1-5p and TrkC-miR1-3p. Transfecting a DNA fragment corresponding to the TrkC-premir1 sequence in HEK293t cells caused ~300-fold elevation in the level of mature TrkC-miR1 and also a significant downregulation of its predicted target genes. Furthermore, endogenous TrkC-miR1 was detected in several cell lines and brain tumors confirming its endogenous generation. Furthermore, its orthologous miRNA was detected in developing rat brain. Accordingly, TrkC-miR1 expression was increased during the course of neural differentiation of NT2 cell, whereas its suppression attenuated NT2 differentiation. Consistent with opposite functions of TrkC, TrkC-miR1 overexpression promoted survival and apoptosis in U87 and HEK293t cell lines, respectively. In conclusion, our data report the discovery of a new miRNA with overlapping function to TrkC.
Insights
Researchers discovered a new microRNA (miRNA), TrkC-miR1, embedded within the TrkC gene. This miRNA influences cell survival, apoptosis, and neural differentiation, revealing novel functions for TrkC signaling pathways.
Area of Science:
- Molecular Biology
- Genetics
- Neuroscience
Background:
- Tropomyosin receptor kinase C (TrkC) plays critical roles in cell survival, apoptosis, and tumorigenesis.
- The p75 neurotrophin receptor (p75(NTR)) has known functions mediated by embedded microRNAs (miRNAs).
Purpose of the Study:
- To investigate the existence and function of miRNAs embedded within the TrkC gene.
- To experimentally verify the expression and processing of predicted TrkC-associated miRNAs.
Main Methods:
- Bioinformatic prediction of miRNAs within the TrkC gene.
- Experimental verification of miRNA expression and processing in HEK293t cells.
- Detection of endogenous TrkC-miR1 in cell lines, brain tumors, and developing rat brain.
- Assessment of TrkC-miR1's role in neural differentiation and cell fate determination.
Main Results:
- Two miRNAs, TrkC-miR1-5p and TrkC-miR1-3p, were experimentally validated.
- Transfection of TrkC-premir1 led to a significant increase in mature TrkC-miR1 and downregulation of target genes.
- Endogenous TrkC-miR1 was detected in various human and rat tissues, including developing brain.
- TrkC-miR1 expression correlated with neural differentiation, promoting survival or apoptosis depending on the cell line.
Conclusions:
- Discovery of a novel miRNA, TrkC-miR1, originating from the TrkC gene.
- TrkC-miR1 exhibits functions overlapping with TrkC, impacting cell survival, apoptosis, and neural differentiation.
- This finding provides new insights into the complex regulatory mechanisms of TrkC signaling.
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