The Role of Chromatin Accessibility, Chromosome Conformation, and Efficient H2A.Z Incorporation in Acquired Thermotolerance in Arabidopsis thaliana

Accessibility to nuclease cleavage has proven to be a useful genomic marker. Transcribing regions are sensitive to cleavage, and regulatory regions are hypersensitive to cleavage. Using the hypersensitivity assay, we previously found that, unlike in most tissue and conditions, plants undergoing lethal heat stress become accessible in many heat-shock-related gene bodies, as well as, more typically, in noncoding regulatory regions. Here, we examine whether gene body accessibility is merely a symptom of cell death or is functionally associated with acquired thermotolerance, a phenomenon in which plants exposed to mild heat stress temporarily acquire the ability to survive typically-lethal levels of heat stress. We find that those same heat-shock-related gene bodies do become accessible during mild heat stress, and that this accessibility disappears within two hours, reappearing during subsequent heat stress in plants with acquired thermotolerance. Further, we identified 24 additional genes that display heat-induced accessibility only in plants with acquired thermotolerance. We performed a parallel set of experiments on arp6 mutant plants, which are defective in both H2A.Z incorporation and thermomemory. We found slightly lower gene body accessibility in arp6 plants compared to wildtype at the long-term acquired thermotolerance time point. Lastly, to determine if heat stress altered chromosome conformation, we performed Hi-C. We found that most interactions between loci in the nuclear genome did not change significantly during heat stress. Instead, we found that lethal heat stress increased the number of interactions between the chloroplast genome and the nuclear genome, suggesting chloroplast sensitivity during heat shock. Acquired thermotolerance, however, prevented this increase in chloroplast interactions, suggesting a protection to chloroplasts.