J Integr Plant Biol. ›› 2021, Vol. 63 ›› Issue (7): 1294-1308.DOI: 10.1111/jipb.13081

• Functional Omics and Systems Biology • Previous Articles     Next Articles

Genome-wide profiling of circular RNAs, alternative splicing, and R-loops in stem-differentiating xylem of Populus trichocarpa

Xuqing Liu1†, Yubang Gao2†, Jiakai Liao2, Miao Miao1, Kai Chen2, Feihu Xi2, Wentao Wei1, Huihui Wang1, Yongsheng Wang2, Xi Xu2, Anireddy S.N. Reddy3 and Lianfeng Gu1*   

  1. 1 Basic Forestry and Proteomics Research Center, College of Forestry, Fujian Provincial Key Laboratory of Haixia Applied Plant Systems Biology, Fujian Agriculture and Forestry University, Fuzhou 350002, China
    2 Basic Forestry and Proteomics Research Center, College of Life Science, Fujian Agriculture and Forestry University, Fuzhou 350002, China
    3 Department of Biology and Program in Cell and Molecular Biology, Colorado State University, Fort Collins, Colorado 80523, USA

    These authors contributed equally to this work.
    *Correspondence: Lianfeng Gu (lfgu@fafu.edu.cn)
  • Received:2020-11-21 Accepted:2021-02-08 Online:2021-02-11 Published:2021-07-01

Abstract: Circular RNAs (circRNAs) are a recently discovered type of non-coding RNA derived from pre-mRNAs. R-loops consist of a DNA:RNA hybrid and the associated single-stranded DNA. In Arabidopsis thaliana, circRNA:DNA R-loops regulate alternative splicing (AS) of SEPALLATA3 (SEP3). However, the occurrence and functions of circRNAs and R-loops in Populus trichocarpa are largely unexplored. Here, we performed circRNA-enriched sequencing in the stem-differentiating xylem (SDX) of P. trichocarpa and identified 2,742 distinct circRNAs, including circ-CESA4, circ-IRX7, and circ-GUX1, which are generated from genes involved in cellulose, and hemicellulose biosynthesis, respectively. To investigate the roles of circRNAs in modulating alternative splicing (AS), we detected 7,836 AS events using PacBio Iso-Seq and identified 634 circRNAs that overlapped with 699 AS events. Furthermore, using DNA:RNA hybrid immunoprecipitation followed by sequencing (DRIP-seq), we identified 8,932 R-loop peaks that overlapped with 181 circRNAs and 672 AS events. Notably, several SDX-related circRNAs overlapped with R-loop peaks, pointing to their possible roles in modulating AS in SDX. Indeed, overexpressing circ-IRX7 increased the levels of R-loop structures and decreased the frequency of intron retention in linear IRX7 transcripts. This study provides a valuable R-loop atlas resource and uncovers the interplay between circRNAs and AS in SDX of P. trichocarpa.

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