DOE OSTI2018
This DOE project concerns the molecular mechanisms by which plants coordinate carbon (C) and nitrogen (N) metabolism. Specifically, it focuses on a histone methyltransferase protein that targets C/N-metabolism genes for chromatin modifications. How the dynamic changes of chromatin regulate energy metabolism in response to developmental and environmental cues is a new area of study. We have filled this knowledge-gap with our studies of SDG8, a histone methyltransferase, that targets light, C and N metabolism genes. We aimed to understand how plants allocate C resources to assimilate inorganic N onto C-skeletons -- the most energy-intensive pathway in plants – in order to balance energy production and consumption in a changing environment. In order to achieve more C-efficient N-storage under low energy conditions, plants convert assimilated N from “reactive” Gln (C5:N2) into “inert” Asn (C4:N2) through the enzyme ASN1. To identify regulatory factors in this pathway, we used the ASN1 promoter in a genetic selection and discovered SDG8, a histone methyltransferase. We discovered that histone modification by SDG8 coordinates the expression of genes involved in energy generation (light-harvesting and C fixation) with energy consumption (N assimilation and sulfur metabolism). We used this opportunity to explore how histone modifications control energy metabolism genome-wide. Our aims focus on various levels of the regulation of energy related genes at the level of: 1) Chromatin; 2) Transcription; and 3) Metabolites of the C/N regulatory network. Specifically, we asked: 1) How does histone methylation regulate energy metabolism during the N response? 2) How does histone methylation control transcription of the energy metabolism genes? And 3) how does the histone methyltransferase SDG8 and other TF regulators affect C/N metabolism and N use efficiency? Our study indicates that N supply triggers genome-wide changes in chromatin modification via the histone methyltransferase SDG8 which impacts on gene expression, RNA processing, chlorophyll synthesis, N uptake and metabolism. Our study thus revealed a previously overlooked layer of chromatin regulation during nutrient signaling. In broad terms, our study provides a framework to understand how chromatin modifications influence responses to environmental changes and provides unique opportunities for breeding and engineering resilient crops with better performance in the current climate model. Accomplishments AIM 1 CHROMATIN: Role of histone methylation in the ASN1 energy metabolism network. This aim focuses on understanding the role of chromatin modification in response to the environment. We studied the targeting mechanism of histone methylation by SDG8, in particular the role of the CW domain in reading histone methylation status. We also began to study the effect of nitrogen-treatment on genome-wide chromatin modifications in WT and sdg8 mutant plants. AIM 2 TRANSCRIPTION: Transcription control of the ASN1 energy metabolism network. This aim investigated whether SDG8 and the histone modification it mediates affects mRNA processing. We performed genome-wide profiling that uncovered that histone methylation by SDG8 affects the co-transcriptional RNA processing of downstream genes involved in energy and photosynthesis. AIM 3 METABOLISM: A gene regulatory network controlling energy, C/N metabolism and N use efficiency. In Aim 3A, we showed that SDG8 affects the chromatin modification and expression of a transcription factor, CCT, which in turn affects the regulation of carbon responsive genes. In Aim 3B we showed that C and N metabolism is perturbed in the sdg8 mutant through differences in the chlorophyll content, free amino acids, and N uptake. Project activities In Aim 1, we uncovered that the histone methyltransferase SDG8 affects chromatin modification in response to N treatments. The N treatments causes changes in the H3K36me3 distribution off ~200 genes in WT plants, while the N-triggered changes in H3K36me3 of these genes is abrogated in the sdg8-5 mutant. The functional annotation of these 200 genes shows significant enrichment of N-related pathways. Overall, our study suggests that SDG8 mediates chromatin-level adaption of functionally relevant genes in response to changes in N. In Aim 2, we tested if SDG8 plays a role in transcriptional elongation and/or RNA processing. We detected more N-triggered isoform shifting events in the sdg8-5 mutant than in WT, which provides supports for SDG8 being required for proper RNA processing in response to N signaling. In Aim 3, we identified the gene network controlled by a CCT transcription factor under the epigenetic control of SDG8. Isoform abundance of the CCT protein is regulated by SDG8, which leads to an altered response of energy related genes to N level changes. Using physiological, metabolic and genetic approaches, we found that responses to N, including chlorophyll accumulation, N uptake and free amino acids, are altered in sdg8 mutants. Finally, this project provided training to a high school student who performed experiments on the complex interaction of photosynthesis and N availability. The student was named a semi-finalist in both the Regeneron and Siemens National Science competitions for her work.
59 BASIC BIOLOGICAL SCIENCES↗