R-loops at microRNA encoding loci promote co-transcriptional processing of pri-miRNAs in plants

• Published in: Nature plants Vol. 8; no. 4; pp. 402 - 418
• Main Authors: Gonzalo, Lucia, Tossolini, Ileana, Gulanicz, Tomasz, Cambiagno, Damian A., Kasprowicz-Maluski, Anna, Smolinski, Dariusz Jan, Mammarella, María Florencia, Ariel, Federico D., Marquardt, Sebastian, Szweykowska-Kulinska, Zofia, Jarmolowski, Artur, Manavella, Pablo A.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 01.04.2022; Nature Publishing Group
• Subjects: 631/208/505; 631/449/1659; Animals; Biomedical and Life Sciences; Biosynthesis; DNA-directed RNA polymerase; Hybrids; Imaging techniques; Life Sciences; Loci; MicroRNAs; MicroRNAs - genetics; MicroRNAs - metabolism; miRNA; Plant Sciences; Plants - genetics; Post-transcription; R-Loop Structures; R-loops; Ribonucleic acid; RNA; RNA polymerase; RNA polymerase II; RNA Polymerase II - genetics; RNA Processing, Post-Transcriptional
• ISSN: 2055-0278; 2055-0278
• DOI: 10.1038/s41477-022-01125-x

In most organisms, the maturation of nascent RNAs is coupled to transcription. Unlike in animals, the RNA polymerase II (RNAPII) transcribes microRNA genes ( MIRNA s) as long and structurally variable pri-miRNAs in plants. Current evidence suggests that the miRNA biogenesis complex assembly initiates early during the transcription of pri-miRNAs in plants. However, it is unknown whether miRNA processing occurs co-transcriptionally. Here, we used native elongating transcript sequencing data and imaging techniques to demonstrate that plant miRNA biogenesis occurs coupled to transcription. We found that the entire biogenesis occurs co-transcriptionally for pri-miRNAs processed from the loop of the hairpin but requires a second nucleoplasmic step for those processed from the base. Furthermore, we found that co- and post-transcriptional miRNA processing mechanisms co-exist for most miRNAs in a dynamic balance. Notably, we discovered that R-loops, formed near the transcription start site region of MIRNA s, promote co-transcriptional pri-miRNA processing. Furthermore, our results suggest the neofunctionalization of co-transcriptionally processed miRNAs, boosting countless regulatory scenarios. This study demonstrates that the processing of miRNAs in plants can occur coupled to transcription, post-transcriptionally or as a mixture of both in a process promoted by DNA–RNA hybrids upstream from the transcription start site of miRNA loci.