Regulation of mammalian mRNA decapping

The modulation of mRNA degradation plays a critical role for regulation of gene expression. A major mRNA decay pathway in mammals proceeding from the 5' to 3' end is initiated with shortening of 3' poly (A) tail, followed by the cleavage of the 5' cap structure (decapping) and degradation of the mRNA body by the Xrn1 exoribonuclease. Dcp2 is a well characterized mRNA decapping enzyme. It is an RNA binding protein that must bind the RNA in order to recognize the cap for hydrolysis. We identified mRNAs bound by human Dcp2 by coimmunoprecipitation and microarray analysis. Further biochemical assays identified a sequence element of 60 nucleotides at the 5' terminus of the mRNA encoding Rrp41 as a specific Dcp2 substrate which could specifically bind Dcp2 protein and enhance Dcp2 decapping. Mutational analysis of this element revealed a stable stem-loop structure (termed DBDE) contained in the first 33 nucleotides is critical for Dcp2 decapping stimulation. A bioinformatic search was carried out to identify a subset of mRNAs that have a comparable stem-loop structure at the 5? end as potential Dcp2 substrates.;We identified a second cytoplasmic mRNA decapping enzyme Nudt16 in mammalian cells and established stable MEF cells that contained reduced level of Dcp2 and/or Nudt16 protein. Using these MEF cells, we demonstrated the distinct roles for Dcp2 and Nudt16 in nonsense mediated mRNA decay (NMD), ARE-mediated decay and miRNA mediated gene silencing. Our results indicated that NMD preferentially utilizes Dcp2 rather than Nudt16; Dcp2 and Nudt16 are redundant in miRNA mediated silencing; and Dcp2 and Nudt16 are differentially utilized for ARE-mRNA decay.;At last, we demonstrated that anti-viral immune response was regulated by Dcp2 in mouse embryonic fibroblast. In MEF cells lacking Dcp2 protein, interferon-mediated anti-viral response was significantly elevated. We identified IRF7, a crucial transcription factor in anti-viral immunity, as the direct target of Dcp2. Knockdown of Dcp2 led to stabilization of IRF7 mRNA and increased IRF7 protein level. Therefore, Dcp2 functions as a negative regulator of interferon-mediated anti-viral immunity. Interestingly, Dcp2 expression is also induced in virus infection, suggesting a negative feedback loop in the anti-viral immune response.