The barley (Hordeum vulgare ssp. vulgare) Respiratory Burst Oxidase Homolog (HvRBOH) gene family and their plausible role on malting quality

Authors: Mahalingam, Ramamurthy; Graham, Danielle; Walling, Jason

Submitted to: Frontiers in Plant Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/27/2021
Publication Date: 2/19/2021
Citation: Mahalingam, R., Graham, D.L., Walling, J.G. 2021. The barley (Hordeum vulgare ssp. vulgare) Respiratory Burst Oxidase Homolog (HvRBOH) gene family and their plausible role on malting quality. Frontiers in Plant Science. 12. Article 608541. https://doi.org/10.3389/fpls.2021.608541.
DOI: https://doi.org/10.3389/fpls.2021.608541

Interpretive Summary: Plant Respiratory Burst Oxidase Homologs (RBOHs) are key sources of reactive oxygen species (ROS) in plants that regulate growth, development and responses to external environment. We identified seven new barley RBOH genes using in silico analysis, in addition to the six identified earlier. Phylogenetic analysis of barley RBOHs and other plants identified moncot and dicot RBOHs form distinct clades. We propose to name the barley RBOHs based on rice genes. Barley RBOH genes exhibited unique transcriptional profile in various tissues. During the barley malting process, ROS levels increased during steeping and one day after germination. ROS was localized to embryo and aleurone layer. This increase in ROS led to increase in the protein oxidation and lipid peroxidation. Nine RBOHs were differentially expressed during malting and most notable was the HvRBOHA/C. Seeds of the RNAi knockdown line of HvRBOHA/C genes exhibited poor malting quality profile such as low alpha-amylase activity, high levels of beta-glucan and poor malt extractability. Our study demonstrates that RBOH-induced ROS is a key player in determining malt quality and a potential novel biomarker that can be harnessed by the malting and brewing industry.

Technical Abstract: Controlled generation of reactive oxygen species (ROS) is pivotal for normal plant development and adaptation to changes in the external milieu. One of the major enzymatic sources of ROS in plants are the plasma-membrane localized NADPH oxidases, also called as Respiratory Burst Oxidase Homologs (RBOH). Seven novel members of RBOH gene family were identified in barley using in silico analysis, in addition to the six previously reported. Conservation of genomic structure and key residues important for catalytic activity and co-factor binding was observed in barley RBOH genes and other plant RBOHs. Based on the tendency of the monocot RBOHs to form distinct phylogenetic clades, we propose to use the rice nomenclature for barley RBOH genes. Temporal changes in ROS profiles were observed during barley malting and was accompanied by changes in protein carbonylation and lipid peroxidation. Nine of the 13 HvRBOH genes were differentially expressed during various malting stages. Among them, the HvRBOHA/C genes showed significant sustained increases in expression during various malting stages. Seeds from the RNAi knockdown lines with reduced expression of HvRBOHC/A gene showed poor malting quality profile such as low alpha-amylase activity and high levels of beta-glucan. We surmise that the ROS produced by HvRBOHs play a critical role in modulating the redox milieu during the early stages of malting via changes in enzyme activities that impact carbohydrate metabolism.