Bose, Mainak (2017) MECHANISM OF miRNA ACTIVITY REGULATION IN MAMMALIAN CELLS. PhD thesis, University of Calcutta.


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    Bhattacharyya, Suvendra Nath


    Cells must be able to respond to changes in their microenvironment in order to survive. Genes encode proteins and proteins dictate cell function. Information flows from DNA to RNA to protein, according to the central dogma of molecular biology and each of the steps are under stringent regulatory control to ensure cell fate and function. Cells can control which genes get transcribed and which transcripts get translated. Regulation of the two major steps — transcription and translation — is critical to its adaptability. Regulation of transcription and translation occurs in both prokaryotes and eukaryotes, but it is far more complex in eukaryotes. MicroRNAs (miRNAs) are a class of small non-coding RNAs that posttranscriptionally regulate gene expression via translational repression and/or mRNA degradation. These tiny regulators search for cognate targets by base pairing with the 3’UTR of the target mRNAs. Over the years, since its discovery the role of miRNAs has become apparent in regulating developmental timing, host-pathogen interactions as well as cell differentiation, proliferation, apoptosis, tumorigenesis, etc. Just like any other regulatory element, the biogenesis, activity and turnover of miRNAs themselves are under strict regulatory control. Extensive research has established how miRNAs regulate target mRNAs by translation repression. However, information regarding the effect of target mRNA on biogenesis and stability of corresponding miRNAs is limited. In this study, we have reported increased biogenesis of cognate miRNAs in presence of abundant amounts of target mRNA in both cells as well as cell-free in vitro system. These miRNAs get loaded onto AGO2 to form functionally competent miRISCs. This target-driven miRNA increase is proportional to the concentration of target mRNA and is affected by the translatability of the target message. While investigating the molecular mechanism of the phenomenon, we identified that increased pre-miRNA processing by AGO2-associated DICER1 in presence of target mRNA, contributes to this increased miRNP formation. Compartmentalization of biological processes provides a mechanism of regulation of the processes with exquisite spatial and temporal control. We have observed that miRNA activity is compartmentalized on the rough Endoplasmic Reticulum (rER) membranes in human cells and have explored the effects of this compartmentalization on miRNA function. Probing further into the phenomenon of target driven miRNA biogenesis, we identified the rER membranes as the site of target mRNA-governed miRNA assembly in human cells. It is likely that target driven miRNA biogenesis operates in addition to the conventional process of repression by preformed miRNPs. Rather than transcriptionally upregulating miRNA synthesis, modulating the final step of biogenesis will serve as an immediate means to increase miRNA production. This in turn will help the cell respond to specific and urgent cellular needs like rapid target driven miR-122 biogenesis during starvation stress reversal in human hepatic cells. Thus, we have identified an additional layer of post-transcriptional regulation of gene expression that helps the cell to maintain requisite levels of mature forms of respective miRNAs by modulating its synthesis depending on target availability.

    Item Type: Thesis (PhD)
    Subjects: Molecular & Human Genetics
    Divisions: UNSPECIFIED
    Depositing User: Dr N C Ghosh
    Date Deposited: 18 Jun 2018 15:27
    Last Modified: 18 Jun 2018 15:28

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