Network-Guided Discovery of Extensive Epistasis between Transcription Factors Involved in Aliphatic Glucosinolate Biosynthesis

Abstract

Plants use diverse mechanisms influenced by vast regulatory networks of indefinite scale to adapt to their environment. These regulatory networks have an unknown potential for epistasis between genes within and across networks. To test for epistasis within an adaptive trait genetic network, we generated and tested 47 double mutant combinations for 20 transcription factors, which all influence the accumulation of aliphatic glucosinolates, the defense metabolites that control fitness. The epistatic combinations were used to test if there is more or less epistasis depending on gene membership within the same or different phenotypic subnetworks. Extensive epistasis was observed between the transcription factors, regardless of subnetwork membership. Metabolite accumulation displayed antagonistic epistasis, suggesting the presence of a buffering mechanism. Epistasis affecting enzymatic estimated activity was highly conditional on the tissue and environment and shifted between both antagonistic and synergistic forms. Transcriptional analysis showed that epistasis shifts depend on how the trait is measured. Because the 47 combinations described here represent a small sampling of the potential epistatic combinations in this genetic network, there is potential for significantly more epistasis. Additionally, the main effect of the individual gene was not predictive of the epistatic effects, suggesting that there is a need for further studies.

Epistatic Transcription Factor Networks Differentially Modulate Arabidopsis Growth and Defense

“…The 20 selected TFs for this study were originally identified as binding aliphatic GLS related promoters and their knockout mutants influence aliphatic GLS accumulation (29, 30). To measure growth variation in this mutant collection, we utilized digital image analysis to measure plant size in two chambers, CEF clean chamber and LSA herbivore chamber (30). Growth was measured every other day from 9 days post-germination to 27 days at which time the leaves were overlapping.…”

“…In agreement with the coordination model, a large scale yeast one hybrid study identified numerous TFs influencing both GLS accumulation and development (29). Critically, these TFs showed a large number of epistatic interactions in influencing the accumulation of GLS (30). However, previous studies did not inform on whether these TFs linked to aliphatic GLS have individual or epistatic effects on growth or flowering.…”

“…To test if growth and defense are linked by this epistatic network of TFs, we used a high throughput method to measure growth and flowering time of 20 single TF mutants and 48 double mutants previously shown to influence the Arabidopsis aliphatic GLS pathway in two environments (30, 31). We show that 17 of the 20 TFs significantly influence plant growth and/or plant flowering time.…”

Symphony of the Regulators: How Do Plants Control Complex Responses to Environmental Signals?

“…The dynamic and quantitative nature of GLS biosynthesis makes it an attractive system for understanding complex transcriptional regulation, especially given that the genes and enzymes involved are almost entirely characterized (Halkier & Gershenzon 2006). Li et al (2018) have taken on this challenge, focusing on epistatic (non-additive) interactions of transcriptional regulators in aliphatic GLS biosynthesis. They generated 47 double mutants of T-DNA insertion lines for 20 TFs that regulate this process.…”

“…Across these categories, in two tissues and two environments, Li et al (2018) find widespread and largely antagonistic epistasis in the regulatory network of aliphatic GLS biosynthesis on GLS accumulation phenotypes (e.g. Figure, which shows epistasis in short chain GLS accumulation).…”