Abstract
MicroRNAs (miRNAs) are a class of small non-coding RNAs that regulate gene or protein expression by targeting mRNAs and triggering either translational repression or mRNA degradation. Distinct expression levels of miRNAs, including miR-29b, have been detected in various biological fluids and tissues from a large variety of disease models. However, how miRNAs “react” and function in different cellular environments is still largely unknown. In this study, the regulation patterns of miR-29b between human and mouse cell lines were compared for the first time. CRISPR/Cas9 gene editing was used to stably knockdown miR-29b in human cancer HeLa cells and mouse fibroblast NIH/3T3 cells with minimum off-targets. Genome editing revealed mir-29b-1, other than mir-29b-2, to be the main source of generating mature miR-29b. The editing of miR-29b decreased expression levels of its family members miR-29a/c via changing the tertiary structures of surrounding nucleotides. Comparing transcriptome profiles of human and mouse cell lines, miR-29b displayed common regulation pathways involving distinct downstream targets in macromolecular complex assembly, cell cycle regulation, and Wnt and PI3K-Akt signalling pathways; miR-29b also demonstrated specific functions reflecting cell characteristics, including fibrosis and neuronal regulations in NIH/3T3 cells and tumorigenesis and cellular senescence in HeLa cells.
Highlights
MicroRNAs are a class of small non-coding RNAs that regulate gene or protein expression by targeting messenger RNAs (mRNAs) and triggering either translational repression or mRNA degradation
MicroRNAs are a class of 18–24 nucleotides long small non-coding RNAs that affect cellular gene and protein expression by modulating the stability and translational efficiency of their target messenger RNAs1. miRNAs have been discovered in various organisms, and exhibited their critical roles in diverse pathological processes and as potential therapeutic targets for treating diseases[2,3]
The passenger strand of the duplex usually forms miRNA* which usually goes through rapid degradation, whereas the guide strand steers miRNA induced silencing complex to target mRNA transcripts[6]
Summary
MicroRNAs (miRNAs) are a class of small non-coding RNAs that regulate gene or protein expression by targeting mRNAs and triggering either translational repression or mRNA degradation. Comparing transcriptome profiles of human and mouse cell lines, miR-29b displayed common regulation pathways involving distinct downstream targets in macromolecular complex assembly, cell cycle regulation, and Wnt and PI3K-Akt signalling pathways; miR-29b demonstrated specific functions reflecting cell characteristics, including fibrosis and neuronal regulations in NIH/3T3 cells and tumorigenesis and cellular senescence in HeLa cells. In this study we systematically designed and revealed details of the specificity and consistency in miR-29b regulations using the same editing method and experimental approaches Using this system, we investigated gene regulation via miRNA clusters, mature miRNA generation, and differential gene expression (DEG) profiles induced by miR-29b stable knockdown between two cell lines. This study provides a comprehensive analysis into understanding the regulatory patterns of miR-29b in different cellular environments and species
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