Genome-Scale Investigation of Integrated Nuclear and Cytoplasmic Gene Regulatory Control in Arabidopsis
- Author(s): Lee, Travis
- Advisor(s): Bailey-Serres, Julia
- et al.
Plants are resilient to transient limitations in the availability of oxygen for efficient energy production. The highly reversible response to oxygen deprivation (hypoxia) is characterized by dynamics in accumulation and differential translation of a subset of genes in the model plant Arabidopsis thaliana. Transcriptional upregulation of genes associated with survival in response to hypoxia are mediated by group VII ETHYLENE RESPONSIVE FACTOR (ERFVII) transcription factors via binding to a conserved Hypoxia Responsive Promoter Element (HRPE) cis-element. Nonetheless, there is little knowledge of the effects of hypoxia on nuclear processes, including histone modification, chromatin accessibility, RNA polymerase II (RNAPII) elongation or RNA export. Additionally, in vivo ERFVII binding sites and dynamics are largely unexplored at the global scale.
In this dissertation, the modulation of chromatin features, RNAPII elongation and nascent transcripts were contrasted with the total polyadenylated and ribosome-associated sub-populations of mRNA of seedlings exposed to non-stress, hypoxic, or re-oxygenation conditions. The technologies utilized in this study to generate datasets included Chromatin Immunoprecipitation (ChIP-seq), Isolation of Nuclei Tagged in Specific Cell Types (INTACT), Assay for Transposase Accessible Chromatin (ATAC-seq), RNA sequencing (RNA-seq), and Translating Ribosome Affinity Purification (TRAP-seq). In vivo binding sites were mapped for the Arabidopsis ERFVII HYPOXIA RESPONSIVE ERF2 (HRE2), and the rice (Oryza sativa) ERFVIIs SUBMERGENCE 1A and C (SUB1A/C).
Integrated bioinformatic analyses were performed to investigate coordination in regulation from chromatin to translation. This led to the identification of multiple nuclear-regulated processes that contribute to dynamics in gene activity. The ~50 hypoxia-responsive genes that display coordinate transcript accumulation and translation were characterized by increased chromatin accessibility, pronounced elevation of Histone 3-lysine 9 acetylation and depletion of Histone 2A.Z under hypoxic stress. HRE2 bound just 5’ of the transcription start site for many of these genes. Universal stress response genes including those associated with heat stress displayed an early increase in RNAPII engagement along the transcription unit with elevation of mRNA only after the stress was prolonged. This study revealed genes and nascent transcripts poised for expression in anticipation of prolonged stress or reoxygenation that were previously unrecognized in widely utilized whole-cellular RNA-seq analyses.