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mirMachine: A One-Stop Shop for Plant miRNA Annotation
Published on: May 1, 2021
Common and distinct patterns of terminal modifications to mirtrons and canonical microRNAs
Jakub O Westholm1, Erik Ladewig, Katsutomo Okamura
1Department of Developmental Biology, Sloan-Kettering Institute, New York, NY 10065, USA.
Abstract:
Nucleotide modifications to microRNAs or their precursors can influence their processing and/or activity. A challenge to their analysis is the lack of independent references for the termini generated by primary processing; typically, these are empirically assigned as the most abundant mapped reads. Mirtrons offer such an independent measure since these microRNA hairpins are defined by splicing. Consequently, mirtron-derived reads that deviate from splice sites reflect modification following primary processing. Analysis in Drosophila revealed multiple modification patterns, including select alterations of 5' termini, many 3' resection events, and unexpectedly abundant 3' untemplated monouridylation. Resections occur on mature AGO1-loaded species, whereas uridylation occurs on pre-miRNAs but is compatible with dicing and AGO1 loading. Strikingly, we found many mirtrons whose modified reads are more abundant than those produced by primary processing. In some cases, these abundant modified reads matched the genome owing to fortuitous uridines in downstream flanking exons, thus highlighting the value of an independent reference for the primary-processed sequence. We could further extend the principle of abundant and preferred uridylation of mirtrons, relative to canonical pre-miRNAs, to Caenorhabditis elegans, mouse, and human. Finally, we found that 3' resection occurs broadly across AGO1-loaded canonical miRNA and star species. Altogether, these findings substantially broaden the complexity of terminal modification pathways acting upon small regulatory RNAs.
Insights
Mirtrons, defined by splicing, provide a unique reference for studying microRNA terminal modifications. This research reveals widespread 3' uridylation and resection events across species, expanding our understanding of small regulatory RNA pathways.
Area of Science:
- Molecular Biology
- Genetics
- RNA Biology
Background:
- Nucleotide modifications impact microRNA (miRNA) processing and activity.
- Analyzing miRNA termini is challenging due to the lack of independent references for primary processing.
- Mirtrons, arising from spliced introns, offer a unique internal reference for miRNA processing.
Purpose of the Study:
- To investigate microRNA terminal modifications using mirtrons as an independent reference.
- To characterize novel modification patterns in Drosophila, Caenorhabditis elegans, mouse, and human small regulatory RNAs.
- To understand the impact of these modifications on miRNA processing and activity.
Main Methods:
- Analysis of mirtron-derived small RNA sequencing reads in Drosophila.
- Identification and characterization of 5' and 3' terminal modifications, including uridylation and resection.
- Comparative analysis across different species (Drosophila, C. elegans, mouse, human).
Main Results:
- Discovery of diverse modification patterns, including 3' uridylation and 5' terminal alterations in Drosophila mirtrons.
- Identification of 3' resection events on mature Argonaute 1 (AGO1)-loaded species.
- Demonstration that uridylation occurs on pre-miRNAs and is compatible with subsequent processing and loading.
- Observation that modified mirtron reads can be more abundant than primary processed reads, sometimes matching genomic sequences.
- Extension of findings on mirtron uridylation to C. elegans, mouse, and human.
- Broad occurrence of 3' resection across AGO1-loaded canonical miRNAs and star species.
Conclusions:
- Mirtrons serve as a valuable tool for precisely analyzing microRNA terminal modifications.
- Terminal uridylation and resection are prevalent modification pathways for small regulatory RNAs across multiple species.
- These modifications significantly increase the complexity of small RNA biology and processing.
- Findings highlight the importance of considering post-transcriptional modifications in miRNA function and regulation.
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