Root Development & Acquired Thermotolerance Regulatory Targets in Arabidopsis thaliana

Abstract

In this thesis, I will describe my work in two research topics using Arabidopsis thaliana: root development and acquired thermotolerance. 1. Arabidopsis roots are an excellent model for studying cell development because the root tissue has a mix of undifferentiated cells, developing cells, and mature cells. The roots are structured such that undifferentiated cells reside at the root tip, and mature cells are closer to the plant shoot. Much of the knowledge gained from researching plant roots was gained from microscopy based approaches and transgenic plant lines. These approaches are quite labor intensive and have not been applied in many plant/ crop species. Here I present research profiling the transcriptome of the root using single cell RNA-seq (scRNA-seq). In just one experiment, the scRNA-seq data was able to capture all the major cell types within the root, to recapitulate known developmental changes in transcription associated with each cell type, and to identify novel genes that change over the course of cell development. Next I describe a collaboration project studying the process of lateral root formation, a process in which the developmental state of a mature cell resets to form a population of cells capable of developing into any other cell type in the root. Using scRNA-seq again, genes known to play major roles in lateral root were identified as well as genes that previously had no association with lateral root development. 2. Acquired thermotolerance is a conserved mechanism that allows an organism to survive lethal heat stress if they were recently exposed to a milder heat stress. Acquired thermotolerance is a form of transcriptional memory that primes an organism to respond more effectively to heat stress. This is achieved through changes in how protein coding genes and other elements of the genome are regulated. The full extent of loci that are regulated in order to achieve acquired thermotolerance is not known. Here I present work assaying chromatin accessibility in Arabidopsis in various stages of acquired thermotolerance to identify loci whose accessibility changes in response to heat stress. Loci that become more accessible during heat stress are able to interact with transcription factors and other DNA binding molecules. Specifically, I focus on a previously identified phenomenon of heat induced gene body accessibility seen in plants without acquired thermotolerance responding to lethal heat stress. Here I show that gene body accessibility does occur during a mild heat stress, but most heat induced accessibility changes are temporary. I also identify genes that are heat accessible only in plants with acquired thermotolerance. Next, the role the histone variant H2A.Z may play in acquired thermotolerance was explored using an H2A.Z incorporation mutant, arp6. It was found that arp6 had a defect in its memory of prior heat stress and could not maintain acquired thermotolerance as long as a wild type plant could. Lastly, the possibility that acquired thermotolerance changed chromosome conformation was explored. Although heat stress did not produce large changes in interactions between regions of the nuclear genome, it was found that the number of interactions between the nuclear genome and the chloroplast genome increased drastically during lethal heat stress. This increase was prevented in plants with acquired thermotolerance.