Integrated analysis of mRNA and miRNA in human differentiating muscle cells
Year: 2014
Type: article
Abstract: Muscles are responsible for the movement of body and take up roughly half of the person’s body weight. Skeletal muscles are the only voluntary muscle tissue in human body, being controlled consciously. Skeletal muscle cells form when many smaller progenitor cells lump themselves together to constitute long, straight, multinucleated fibers. The proliferating muscle cells are called myoblasts. Myoblasts fuse together to form multinucleated non-proliferating cells called myotubes. The transition from proliferative to differentiated state involves a complete shift of the cell’s transcriptome based on a network of regulation at the molecular level. To uncover the intricacies of molecular activities involved in the process of muscle differentiation it is essential to have deeper and through understanding of the transcriptome in its entirety. RNA seq, which is a high through put sequencing technology, gives us the opportunity to reach into the deepest level of the transcriptome. By means of the RNA seq technology I obtained the in-depth view of the transcriptome of human muscle cells from the proliferative to the differentiated stage. The analysis was extended also to small RNAs, to have a full picture of the transcriptome. I performed the analysis with the specific objectives of identifying the expressed genes and finding out differential gene expression, of both mRNA and miRNA. I also investigated the crosstalk between mRNA and miRNA, employing two separate methods : miRNA over expression and Ago2 immunoprecipitation followed by RNA seq. The over expression experiments were carried out with 5 different miRNAs and in all cases I found more genes turned up than turned down. These results suggest that miRNA might either have a role in mRNA stabilization or it could play a part in a double negative mechanism by inhibiting some negative factor. By comparing the genes down regulated after miRNA over expression with Ago2-enriched genes we have found several candidate genes which are most likely under the down regulatory control of miRNA. Skeletal muscle has enormous plasticity and can endure a lot of stress. To study this aspect of muscle we performed mechanical stretch of differentiating muscle cells. With RNA seq we got the in-depth view of the transcriptome as a response to stretch. We performed the analysis at two time points after stretch and found that stretch triggered immune response genes soon after, but enhanced muscle structural-protein genes expression over a prolonged course of time when the immune response takes a back seat. I believe that our thorough transcriptome analysis, including miRNA and mRNA interaction studies during myogenesis, contributes towards the better understanding of the process regulating muscle development
Author: Rusha Guha
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W2275862767