"Immediate Early" RNA-seq temperature time series experiment

All plant material was the Columbia (Col-0) ecotype. Plants were harvested from dusk during chilling (20, 40, 60, 90, 120, and 180 minutes after initiation of cooling) or at the same time points in the absence of chilling as a control. A single biological replicate consisted of 9-13 plants in hydroponic containers, aerial tissue harvested and pooled per condition/time point. Three biological replicates were generated for each time point in separate experiments. The same growth cabinet was used for all repeats to eliminate the potential effects of minor changes in light intensities and light quality on gene expression.Project SummarySplicing mediated reversals of the relative abundances of transcript isoforms of the same gene - 'isoform switching' - is a striking feature of plants' rapid response to cooling. We find the flip-flop dynamism of the REVEILLE2 isoform switch particularly striking; it is detectable on the minute-timescale and unusually is an 'off:on' switch with the shift to translatable transcript message evident during the transition to cool nights.The physiological relevance of this circadian clock cold-switch was explored using molecular genetic approaches centred around a newly characterised T-DNA insertion line, rve2-2, that is unable to cold-switch.We demonstrate altered cooling-associated rhythmic behaviour in plants bearing a clock promoter reporter (CCA1::LUC) in the rve2-2 background. RNA-seq experiments with rve2-2 identify potential RVE2 target genes and temporal profiling show they comprise a cascade of gene expression initiated shortly after peak RVE2 expression mid-nocturnal phase extending to the imminent dawn. Surprisingly we find a subset of chloroplast-encoded candidates with modified expression in rve2-2 and consequently we find altered photosynthetic capacity a rve2-2 characteristic.The RVE2 cold switch extends our understanding of plants' rapid response to cooling. We propose that RVE2 integrates nocturnal temperature with priming plants' photosynthetic capacity in anticipation of the break of the day.