Valeria Spadotto

MicroRNAs (miRNAs) are short, non-coding RNA molecules that fine tune gene expression at the post-transcriptional level. The efficient miRNA processing is fundamental to maintain their correct levels in the cell. Alterations in microRNA biogenesis occur in different pathological conditions, including cancer. Hence, dissecting the underlying molecular mechanisms is crucial to understand how aberrant miRNA patterns are generated. The processing step operated by the Large Drosha Complex (LDC), fundamental and rate limiting for the production of miRNAs, occurs in the nucleus and consists in the cleavage of the primary (pri-) into the precursor (pre-) miRNAs. In a previous MS-based analysis of the cellular methylome, we showed that protein methylation occurs on the majority of the LDC subunits, suggesting the possibility that this post-translational modification could contribute to the regulation of miRNA biogenesis.
To investigate the role of protein methylation in the regulation of the LDC, we characterized the methylated sites on the subunits of the complex, thus generating the first comprehensive and high-confident methyl-proteome of the LDC.
By modulating the expression of PRMT1 we analyzed how methylations change at specific sites of distinct LDC subunits, thus identifying PRMT1 substrates. The alteration of the LDC-methylation state upon PRMT1 depletion correlated with an impairment of the pri- to- pre-miRNA processing, which resulted in the massive deregulation of mature miRNA expression. Although the molecular mechanism remains to be elucidated, the results described in this thesis uncover a key role of arginine methylation in the regulation of the LDC activity and, consequently, in miRNA biogenesis.