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Time of day prioritizes the pool of translating mRNAs in response to heat stress

ABSTRACT: The circadian clock helps organisms to anticipate and coordinate gene regulatory responses to changes in environmental stimuli. Under growth limiting temperatures, time of day modulates the accumulation of polyadenylated mRNAs. In response to heat stress, plants will conserve energy and selectively translate mRNAs. How the clock and/or time of day regulates polyadenylated mRNAs bound by ribosomes in response to heat stress is unknown. In-depth analysis of Arabidopsis thaliana translating mRNAs found that time of day gates the response of approximately one-third of the circadian regulated heat responsive translatome. Specifically, time of day and heat stress interact to prioritize the pool of mRNAs in cue to be translated. For a subset of mRNAs, we observed a stronger gated response during the day, and preferentially before the peak of expression. We propose previously overlooked transcription factors (TFs) as regulatory nodes and show that the clock plays a role in the temperature response for select TFs. When the stress was removed, the redefined priorities for translation recovered within one-hour, though slower recovery was observed for abiotic stress regulators. Through hierarchical network connections between clock genes and prioritized TFs, our work provides a framework to target key nodes underlying heat stress tolerance throughout the day.

ORGANISM(S): Arabidopsis thaliana

PROVIDER: GSE158444 | GEO | 2021/04/21

REPOSITORIES: GEO

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Project description:Purpose: To understand time of day variation in Arabidopsis heat shock responses. We have observed significant differences in terms of plant survivability in response to a heat shock across various times of day. Because of this, we believe that transcriptional responses may be a significant underlying factor which contributes to the observed physiology and surivability after heat stress. Methods: Whole plant tissue mRNA profiles in Col-0 Arabidopsis were generated by deep sequencing using Illumina HiSeq 2500. Tissue samples of four replicates were collected at two times of day (Dawn and Dusk, ZT0/ZT12) and were either control or heat stressed (30oC) for 1 hour. An additional 4 samples were collected in constant light conditions (single replicate) to quantify circadian effects. Sequence reads from each sample were mapped to the TAIR10 reference genome with Tophat2 and counts were obtained using Htseq-count. Differential expression analysis was done using DeSeq2 to calculate differential expression levels of each gene in the dataset. Results: We identified 5512 differentially expressed transcripts with our DESeq2 based data analysis pipeline that involves characterizing gene expression profiles based on basal expression level, time of differential expression, and direction of differential expression. An additional interaction term analysis revealed 572 genes which showe time of day dependent responses to heat stress, meaning the response to heat stress differs depending on the time of day that the heat stress is perceived by the plant. Downstream analysis using gene ontology analysis tools (Pantherdb.org) and Cytoscape revealed strong differences in terms of response happening in the AM and PM gene expression networks. The significance of this work is that it is the first study to show that there is a significant difference in terms of transcriptional responses to heat stress in Arabidopsis where the differences were quantified by levels of mRNA expression.

Project description:The Eastern North American monarch butterfly, Danaus plexippus, is notorious for its spectacular seasonal long-distance migration. In recent years, it has also emerged as a novel system to study how animal circadian clocks keep track of time and regulate ecologically relevant daily rhythmic activities and seasonal behavioral outputs. However, unlike Drosophila and the mouse, little work has been undertaken in the monarch to identify clock-controlled output genes and elucidate the regulation of their rhythmic expression. Here, we used RNA-sequencing and Assay for Transposase-Accessible Chromatin (ATAC)-sequencing to profile the diurnal transcriptome, open chromatin regions, and transcription factor (TF) footprints in the brain of wild-type monarchs and Cryptochrome 2 (Cry2), Clock (Clk), and Bmal1 (named DCyc-like) butterfly mutants with impaired clock function. We identified 366 rhythmic transcripts under circadian clock control belonging to biological processes key to brain function, such as neurotransmission, neuropeptide signaling, and glucose metabolism. Surprisingly, we found no significant time of day and genotype-dependent changes in chromatin accessibility (i.e., cis-regulatory elements) in the brain. Instead, we found the existence of a temporal regulation of TFs occupancy within open chromatin regions in the vicinity of rhythmic genes in the brains of wild-type monarchs, which is abolished in clock deficient mutants. Our data suggest that TFs binding specifically in the middle of the day display pioneer-like activity by increasing the accessibility of the surrounding chromatin, while TFs binding specifically in the middle of the night would bind DNA with a longer residency time without affecting accessibility of the surrounding chromatin. Together, this work identifies for the first time the clock-controlled genes and modes of regulation by which diurnal transcription rhythms are regulated in the monarch brain. It also illustrates the power of ATAC-seq to profile genome-wide regulatory elements and TF binding in unconventional organisms.

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Time of day prioritizes the pool of translating mRNAs in response to heat stress

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