Phenotyping stomatal closure by thermal imaging for GWAS and TWAS of water use efficiency-related genes

Authors: PIGNON, CHARLES - University Of Illinois; FERNANDES, SAMUEL - University Of Illinois; VALLURU, RAVI - University Of Nebraska; BANDILLO, NONOY - North Dakota State University; LOZANO, ROBERTO - Cornell University; Buckler, Edward - Ed; GORE, MICHAEL - Cornell University; LONG, STEPHEN - University Of Illinois; BROWN, PATRICK - University Of Illinois; LEAKEY, ANDREW - University Of Illinois

Submitted to: Plant Physiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/26/2021
Publication Date: 8/16/2021
Citation: Pignon, C.P., Fernandes, S.B., Valluru, R., Bandillo, N., Lozano, R., Buckler IV, E.S., Gore, M.A., Long, S.P., Brown, P.J., Leakey, A. 2021. Phenotyping stomatal closure by thermal imaging for GWAS and TWAS of water use efficiency-related genes. Plant Physiology. 184(4):2544-2562. https://doi.org/10.1093/plphys/kiab395.
DOI: https://doi.org/10.1093/plphys/kiab395

Interpretive Summary: Plants need to breathe in CO2 in order to grow and the CO2 enters plants through small pores in the leaves called stomata. Yet, there are tradeoffs to opening and closing of stomata in response to changes in light intensity and water availability. How stomata are regulated and demonstrate natural variation is not well understood in C4 grasses like maize and sorghum, but understanding this is essential for optimizing water-use efficiency. This study examined natural genetic variation in how sorghum controls stomata. Overall the trait is quite heritable and controlled by over two hundred genes, which suggests varieties need to be carefully adapted to specific environments. While a third of the genes are known from a model plant studies, the remaining genetic variation is controlled through yet to be understood genes and pathways. Overall, this study shows that there is sufficient variation that can breed on stomatal responses now, but tremendous work remains to understand the mechanisms behind adaptation.

Technical Abstract: Stomata allow CO2 uptake by leaves for photosynthetic assimilation at the cost of water vapor loss to the atmosphere. The opening and closing of stomata in response to fluctuations in light intensity regulate CO2 and water fluxes and are essential to maintenance of water-use efficiency (WUE). However, little is known about the genetic basis for natural variation in stomatal movement, especially in C4 crops. This is partly because the stomatal response to a change in light intensity is difficult to measure at the scale required for association studies. High-throughput thermal imaging was used to bypass the phenotyping bottleneck and assess 10 traits describing stomatal conductance (gs) before, during and after a stepwise decrease in light intensity for a diversity panel of 659 sorghum accessions. Results from thermal imaging significantly correlated with photosynthetic gas-exchange measurements. gs traits varied substantially across the population and were moderately heritable (h2 up to 0.72). An integrated genome-wide and transcriptome-wide association study (GWAS/TWAS) identified candidate genes putatively driving variation in stomatal conductance traits. Of the 239 unique candidate genes identified with greatest confidence, 77 were orthologs of Arabidopsis genes related to functions implicated in WUE, including stomatal opening/closing (24 genes), stomatal/epidermal cell development (35 genes), leaf/vasculature development (12 genes), or chlorophyll metabolism/photosynthesis (8 genes). These findings demonstrate an approach to finding genotype-to-phenotype relationships for a challenging trait as well as candidate genes for further investigation of the genetic basis of WUE in a model C4 grass for bioenergy, food, and forage production.