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Meg8-DMR as the Secondary Regulatory Region Regulates the Expression of MicroRNAs While It Does Not Affect Embryonic
Liang Zhang1, Zhengbin Han1, Hongjuan He1
1School of Life Science and Technology, Harbin Institute of Technology, Harbin 150001, China.
Abstract:
Meg8-DMR is the first maternal methylated DMR to be discovered in the imprinted Dlk1-Dio3 domain. The deletion of Meg8-DMR enhances the migration and invasion of MLTC-1 depending on the CTCF binding sites. However, the biological function of Meg8-DMR during mouse development remains unknown. In this study, a CRISPR/Cas9 system was used to generate 434 bp genomic deletions of Meg8-DMR in mice. High-throughput and bioinformatics profiling revealed that Meg8-DMR is involved in the regulation of microRNA: when the deletion was inherited from the mother (Mat-KO), the expression of microRNA was unchanged. However, when the deletion occurred from the father (Pat-KO) and homozygous (Homo-KO), the expression was upregulated. Then, differentially expressed microRNAs (DEGs) were identified between WT with Pat-KO, Mat-KO, and Homo-KO, respectively. Subsequently, these DEGs were subjected to the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology (GO) term enrichment analysis to explore the functional roles of these genes. In total, 502, 128, and 165 DEGs were determined. GO analysis showed that these DEGs were mainly enriched in axonogenesis in Pat-KO and Home-KO, while forebrain development was enriched in Mat-KO. Finally, the methylation levels of IG-DMR, Gtl2-DMR, and Meg8-DMR, and the imprinting status of Dlk1, Gtl2, and Rian were not affected. These findings suggest that Meg8-DMR, as a secondary regulatory region, could regulate the expression of microRNAs while not affecting the normal embryonic development of mice.
Insights
Meg8-DMR regulates microRNA expression differently depending on parental inheritance in mice. This imprinted region influences embryonic development pathways without altering overall imprinting status.
Area of Science:
- Genetics
- Developmental Biology
- Epigenetics
Background:
- Meg8-DMR is the first identified maternal methylated differentially methylated region (DMR) in the imprinted Dlk1-Dio3 domain.
- Previous studies showed Meg8-DMR deletion affects cell migration and invasion, but its role in mouse development was unknown.
Purpose of the Study:
- To investigate the biological function of Meg8-DMR during mouse development.
- To determine the impact of Meg8-DMR deletion on microRNA expression and downstream developmental pathways.
Main Methods:
- Generated Meg8-DMR genomic deletions in mice using CRISPR/Cas9.
- Utilized high-throughput and bioinformatics profiling to analyze microRNA expression.
- Performed Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology (GO) term enrichment analysis on differentially expressed genes (DEGs).
Main Results:
- Meg8-DMR deletion inherited paternally (Pat-KO) or homozygously (Homo-KO) upregulated microRNA expression, while maternal inheritance (Mat-KO) did not alter expression.
- Identified 502, 128, and 165 DEGs in Pat-KO, Mat-KO, and Homo-KO groups, respectively.
- GO analysis revealed enrichment in axonogenesis for Pat-KO/Homo-KO and forebrain development for Mat-KO, with no impact on methylation levels or imprinting status of key genes.
Conclusions:
- Meg8-DMR acts as a secondary regulatory region controlling microRNA expression.
- The effects of Meg8-DMR deletion on microRNA expression are parent-of-origin dependent.
- Meg8-DMR is crucial for regulating specific developmental pathways, particularly axonogenesis and forebrain development, without disrupting normal embryonic development.
