Who Watches the Watchmen: Roles of RNA Modifications in the RNA Interference Pathway

Abstract

RNA levels are widely thought to be predictive of RNA function. However, the existence of more than a hundred chemically distinct modifications of RNA alone is a major indication that these moieties may impart distinct functions to subgroups of RNA molecules that share a primary sequence but display distinct RNA "epigenetic" marks. RNAs can be modified on many sites, including 5' and 3' ends, the sugar phosphate backbone, or internal bases, which collectively provide many opportunities for posttranscriptional regulation through a variety of mechanisms. Here, we will focus on how modifications on messenger and microRNAs may affect the process of RNA interference in mammalian cells. We believe that taking RNA modifications into account will not only advance our understanding of this crucial pathway in disease and cancer but will also open the path to exploiting the enzymes that "write" and "erase" them as targets for therapeutic drug development.

Fig 1. Simplified schematic of (A) the miRNA biogenesis pathway and (B) the interaction between the mRNA and miRNA.

(A) The primary miRNA precursor (pri-miRNA) is synthesized by RNAP II. The pri-miRNA is first cleaved by Drosha to release a hairpin loop–shaped RNA called pre-miRNA. The loop of this pre-miRNA is further cleaved by Dicer to generate a miRNA duplex. The miRNA duplex is dissociated and the passenger strand (dashed line) is discarded while the guide strand is loaded onto the Argonaute protein to form an active RISC complex. (B) Example of miRNA–target mRNA interaction by base-pairing mainly at the seed region of miRNA (nt 2–8) but also on other downstream regions of the miRNA. In the mRNA, the coding region is represented as a double line. N.B. The asterisks indicate the m7G cap (7-methyl-guanosine with 5′, 5′-triphosphate linkage).

Fig 2. List of known messenger or miRNA modifications, with chemical structures, abbreviation, writers, erasers, and readers.

*The Drosophila dTET enzyme has recently been shown to hydroxylate 5meC on RNA [11].

Fig 3. Model for the mode of action of the BCDIN3D RNA methyltransferase on the biogenesis of specific miRNAs.

BCDIN3D’s enzymatic activity consists in the methylation of the two available oxygen moieties of the 5′ monophosphate, which removes the 5′ monophosphate charge and makes it bulkier. This methylation blocks the processing of specific miRNAs [8], possibly through perturbing the interaction of the 5′ monophosphate with its binding pocket in Dicer [8,14].

Fig 4. Model for how m6A may stimulate pri-miRNA processing.

m6A is deposited on pri-miRNA by METTL3 and is thought to stimulate the recruitment of Drosha/DGCR8 for co-transcriptional processing of pri-miRNA to pre-miRNA [27]. The question mark is to highlight that the identity of the full set of m6A readers in pri-miRNAs is unknown. In yellow is shown RNAP II on DNA surrounded by nucleosomes.