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  • Title: Targeted mutation of barley (1,3;1,4)-β-glucan synthases reveals complex relationships between the storage and cell wall polysaccharide content.
    Author: Garcia-Gimenez G, Barakate A, Smith P, Stephens J, Khor SF, Doblin MS, Hao P, Bacic A, Fincher GB, Burton RA, Waugh R, Tucker MR, Houston K.
    Journal: Plant J; 2020 Nov; 104(4):1009-1022. PubMed ID: 32890421.
    Abstract:
    Barley (Hordeum vulgare L) grain is comparatively rich in (1,3;1,4)-β-glucan, a source of fermentable dietary fibre that protects against various human health conditions. However, low grain (1,3;1,4)-β-glucan content is preferred for brewing and distilling. We took a reverse genetics approach, using CRISPR/Cas9 to generate mutations in members of the Cellulose synthase-like (Csl) gene superfamily that encode known (HvCslF6 and HvCslH1) and putative (HvCslF3 and HvCslF9) (1,3;1,4)-β-glucan synthases. Resultant mutations ranged from single amino acid (aa) substitutions to frameshift mutations causing premature stop codons, and led to specific differences in grain morphology, composition and (1,3;1,4)-β-glucan content. (1,3;1,4)-β-Glucan was absent in the grain of cslf6 knockout lines, whereas cslf9 knockout lines had similar (1,3;1,4)-β-glucan content to wild-type (WT). However, cslf9 mutants showed changes in the abundance of other cell-wall-related monosaccharides compared with WT. Thousand grain weight (TGW), grain length, width and surface area were altered in cslf6 knockouts, and to a lesser extent TGW in cslf9 knockouts. cslf3 and cslh1 mutants had no effect on grain (1,3;1,4)-β-glucan content. Our data indicate that multiple members of the CslF/H family fulfil important functions during grain development but, with the exception of HvCslF6, do not impact the abundance of (1,3;1,4)-β-glucan in mature grain.
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