Understanding the epigenetics of alternative splicing in Arabidopsis thaliana

Being sessile and photosynthetic necessitates that plants must have a certain degree of predictability for ambient conditions during the daily cycles to maximise efficiency and survival. However, subtle or sudden changes in weather conditions alongside different growth and developmental phases necessitate that plants must continuously monitor different environmental cues and synchronise them with their physiology and metabolism in a time, growth phase and development-stage dependent manner. Plants use complex gene regulatory mechanisms to overcome environmental challenges. Alternative splicing (AS) of pre-mRNAs, a process that generates two or more transcripts from multi-exon genes, adds another layer of complexity to gene regulatory mechanisms to modulate transcriptome diversity in a tissue- and condition-dependent manner. In mammals, mounting evidence indicates that chromatin structure can regulate co-transcriptional AS. Recent evidence supports co-transcriptional regulation of AS in plants, but how dynamic changes in the chromatin influence the AS process upon cold stress remains poorly understood and is the subject of this study. In order to answer this question, four approaches were followed in parallel; (1) Arabidopsis thaliana (Arabidopsis) plants with identical DNA sequence but differential DNA methylation and nucleosome occupancy (Ctrl and AzadC plants with wild type DNA methylation and hypomethylation, respectively) were developed to perform (2) RNA sequencing (RNA-seq) and (3) Micrococcal Nuclease sequencing (MNase-seq) for Ctrl and AzadC grown at 22oC and at 4oC as cold treatment for 24 hours, and (4) whole genome bisulphite sequencing (WGBS) for AzadC grown at 22oC and at 4oC as cold treatment for 24 hours. This strategy allowed us to understand how epigenetic variations between AzadC treatment derived lines and Ctrl plants affect AS under cold conditions, without the confounding effects of sequence variation. Interestingly, RNA-seq, MNase-seq, and WGBS show a strong reprogramming of AS patterns upon cold stress associated with changes in epigenetic features (i.e. DNA methylation and nucleosome occupancy). To my knowledge, this is the first study in Arabidopsis that demonstrates that changes in transcriptional and AS patterns coincide with genome-wide changes in nucleosome occupancy and DNA methylation patterns upon temperature shift.