Indian Himalayan natural Arabidopsis thaliana accessions with abolished miR158 levels exhibit robust miR173-initiated trans-acting cascade silencing

Plant J. 2023 May;114(4):855-874. doi: 10.1111/tpj.16175. Epub 2023 Mar 30.


Small RNAs (sRNAs) such as microRNAs (miRNAs) and small interfering RNAs (siRNAs) are short 20-24-nucleotide non-coding RNAs. They are key regulators of gene expression in plants and other organisms. Several 22-nucleotide miRNAs trigger biogenesis cascades of trans-acting secondary siRNAs, which are involved in various developmental and stress responses. Here we show that Himalayan Arabidopsis thaliana accessions having natural mutations in the miR158 locus exhibit robust cascade silencing of the pentatricopeptide repeat (PPR)-like locus. Furthermore, we show that these cascade sRNAs trigger tertiary silencing of a gene involved in transpiration and stomatal opening. The natural deletions or insertions in MIR158 led to improper processing of miR158 precursors, thereby blocking synthesis of mature miR158. Reduced miR158 levels led to increased levels of its target, a pseudo-PPR gene that is targeted by tasiRNAs generated by the miR173 cascade in other accessions. Using sRNA datasets derived from Indian Himalayan accessions, as well as overexpression and knockout lines of miR158, we show that absence of miR158 led to buildup of pseudo-PPR-derived tertiary sRNAs. These tertiary sRNAs mediated robust silencing of a gene involved in stomatal closure in Himalayan accessions lacking miR158 expression. We functionally validated the tertiary phasiRNA that targets NHX2, which encodes a Na+ -K+ /H+ antiporter protein, thereby regulating transpiration and stomatal conductance. Overall, we report the role of the miRNA-TAS-siRNA-pseudogene-tertiary phasiRNA-NHX2 pathway in plant adaptation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Arabidopsis* / genetics
  • Arabidopsis* / metabolism
  • Base Sequence
  • Gene Expression Regulation, Plant / genetics
  • MicroRNAs* / genetics
  • MicroRNAs* / metabolism
  • Nucleotides / metabolism
  • Plants / metabolism
  • RNA, Plant / genetics
  • RNA, Plant / metabolism
  • RNA, Small Interfering / genetics


  • RNA, Small Interfering
  • MicroRNAs
  • Nucleotides
  • RNA, Plant