Identification of Novel miRNAs and Their Target Genes in the Response to Abscisic Acid in Arabidopsis.

Agnieszka Ludwików,Sivakumar Krishnamoorthy,Syed Muhammad Muntazir Mehdi,Michal Wojciech Szczesniak

doi:10.3390/ijms22137153

Agnieszka Ludwików, Sivakumar Krishnamoorthy + Show 2 more

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https://doi.org/10.3390/ijms22137153

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Abstract

miRNAs are involved in various biological processes, including adaptive responses to abiotic stress. To understand the role of miRNAs in the response to ABA, ABA-responsive miRNAs were identified by small RNA sequencing in wild-type Arabidopsis, as well as in abi1td, mkkk17, and mkkk18 mutants. We identified 10 novel miRNAs in WT after ABA treatment, while in abi1td, mkkk17, and mkkk18 mutants, three, seven, and nine known miRNAs, respectively, were differentially expressed after ABA treatment. One novel miRNA (miRn-8) was differentially expressed in the mkkk17 mutant. Potential target genes of the miRNA panel were identified using psRNATarget. Sequencing results were validated by quantitative RT-PCR of several known and novel miRNAs in all genotypes. Of the predicted targets of novel miRNAs, seven target genes of six novel miRNAs were further validated by 5′ RLM-RACE. Gene ontology analyses showed the potential target genes of ABA-responsive known and novel miRNAs to be involved in diverse cellular processes in plants, including development and stomatal movement. These outcomes suggest that a number of the identified miRNAs have crucial roles in plant responses to environmental stress, as well as in plant development, and might have common regulatory roles in the core ABA signaling pathway.

Highlights

Over the last few decades, there has been intensive research into the physiological effects of abscisic acid (ABA) [1] on plants
Our results are in accordance with earlier reports from different plants, which show that 24 nt sRNAs are a highly abundant category of sRNA reads [58,59,60]. miRNA length is important for alignment with the RNA-induced silencing complex (RISC), which results in the degradation of the target mRNA or inhibition of its translation
Using RNA-Seq, we identified miRNAs in the WT Col-0 of Arabidopsis and its abi1td, mkkk17, and mkkk18 mutant lines, which are differentially expressed in response to ABA treatment

Summary

Introduction

Over the last few decades, there has been intensive research into the physiological effects of abscisic acid (ABA) [1] on plants. ABA controls plant responses via numerous effector proteins, but the most important route for the regulation of plant growth and development is the core ABA signaling pathway This linear pathway comprises three classes of protein: the ABA receptors themselves, variously named pyrabactin resistance/pyrabactin resistance-like/regulatory component of the ABA receptor (encoded by the PYR/PYL/RCAR genes); the negative regulators of the pathway, the protein phosphatase 2C (PP2C) group A family; and the positive regulators SNF1-related protein kinases type 2 (SnRK2 genes). It has been shown that ABA can affect the expression levels of several microRNAs (miRNAs), which regulate downstream effector genes [7]

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