Cold-Dependent Expression and Alternative Splicing of Arabidopsis Long Non-coding RNAs.

Cristiane P G Calixto,Csaba Hornyik,Runxuan Zhang,Allan B James,John W S Brown,Hugh G Nimmo,Wenbin Guo,Nikoleta A Tzioutziou

doi:10.3389/fpls.2019.00235

Cristiane P G Calixto, Csaba Hornyik + Show 6 more

Open Access

https://doi.org/10.3389/fpls.2019.00235

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Abstract

Plants re-program their gene expression when responding to changing environmental conditions. Besides differential gene expression, extensive alternative splicing (AS) of pre-mRNAs and changes in expression of long non-coding RNAs (lncRNAs) are associated with stress responses. RNA-sequencing of a diel time-series of the initial response of Arabidopsis thaliana rosettes to low temperature showed massive and rapid waves of both transcriptional and AS activity in protein-coding genes. We exploited the high diversity of transcript isoforms in AtRTD2 to examine regulation and post-transcriptional regulation of lncRNA gene expression in response to cold stress. We identified 135 lncRNA genes with cold-dependent differential expression (DE) and/or differential alternative splicing (DAS) of lncRNAs including natural antisense RNAs, sORF lncRNAs, and precursors of microRNAs (miRNAs) and trans-acting small-interfering RNAs (tasiRNAs). The high resolution (HR) of the time-series allowed the dynamics of changes in transcription and AS to be determined and identified early and adaptive transcriptional and AS changes in the cold response. Some lncRNA genes were regulated only at the level of AS and using plants grown at different temperatures and a HR time-course of the first 3 h of temperature reduction, we demonstrated that the AS of some lncRNAs is highly sensitive to small temperature changes suggesting tight regulation of expression. In particular, a splicing event in TAS1a which removed an intron that contained the miR173 processing and phased siRNAs generation sites was differentially alternatively spliced in response to cold. The cold-induced reduction of the spliced form of TAS1a and of the tasiRNAs suggests that splicing may enhance production of the siRNAs. Our results identify candidate lncRNAs that may contribute to the regulation of expression that determines the physiological processes essential for acclimation and freezing tolerance.

Highlights

Non-coding RNAs are a diverse set of RNAs which do not generally code for proteins
Rosettes were sampled at 3 h intervals for the last day at 20◦C, the first day at 4◦C, and the fourth day at 4◦C (Supplementary Figure S1A) and each time-point consisted of three biological replicates
We identified a total of 7302 differential expression (DE) genes and 2442 differential alternative splicing (DAS) genes whose expression was significantly differentially expressed or alternatively spliced, respectively

Summary

Introduction

Non-coding RNAs (ncRNAs) are a diverse set of RNAs which do not generally code for proteins. They include families of house-keeping ncRNAs and their precursors such as small nuclear ribonucleoprotein particle RNAs (snRNAs), small Cajal body RNAs (scaRNAs), ribosomal RNAs (rRNAs), transfer RNAs (tRNAs), and small nucleolar RNA (snoRNAs) (Shaw and Brown, 2012; Cech and Steitz, 2014). The regulatory ncRNAs include small RNAs such as microRNAs (miRNAs), short-interfering RNAs (siRNAs), and long non-coding RNAs (lncRNAs) which are expressed from intergenic regions or introns and include natural antisense transcripts (NATs). Some lncRNAs are precursors of small RNA production: primary transcripts are processed to miRNAs or siRNAs such as trans-acting siRNAs (tasiRNAs) or natural antisense siRNAs (nat-siRNAs) derived from double-stranded RNA molecules. Tens of thousands of lncRNAs are routinely detected in RNA-seq analyses demonstrating a new level of complexity of gene expression (Liu et al, 2012, 2015; Zhang and Chen, 2013; Yuan et al, 2016; Lu et al, 2017; Wang et al, 2017; Severing et al, 2018; Zhao et al, 2018)

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