A mixed-linkage (1,3;1,4)-β-D-glucan specific hydrolase mediates dark-triggered degradation of this plant cell wall polysaccharide

The presence of mixed-linkage (1,3;1,4)-beta-D-glucan (MLG) in plant cell walls is a key feature of grass species such as cereals, the main source of calorie intake for humans and cattle. Accumulation of this polysaccharide involves the coordinated regulation of biosynthetic and metabolic machineries. While several components of the MLG biosynthesis machinery have been identified in diverse plant species, degradation of MLG is poorly understood. In this study, we performed a large-scale forward genetic screen for maize (Zea mays) mutants with altered cell wall polysaccharide structural properties. As a result, we identified a maize mutant with increased MLG content in several tissues, including adult leaves and senesced organs, where only trace amounts of MLG are usually detected. The causative mutation was found in the GRMZM2G137535 gene, encoding a GH17 licheninase as demonstrated by an in vitro activity assay of the heterologously expressed protein. In addition, maize plants overexpressing GRMZM2G137535 exhibit a 90% reduction in MLG content, indicating that the protein is not only required, but its expression is sufficient to degrade MLG. Accordingly, the mutant was named MLG hydrolase 1 (mlgh1). mlgh1 plants show increased saccharification yields upon enzymatic digestion. Stacking mlgh1 with lignin-deficient mutations results in synergistic increases in saccharification. Time profiling experiments indicate that wall MLG content is modulated during day/night cycles, inversely associated with MLGH1 transcript accumulation. This cycling is absent in the mlgh1 mutant, suggesting that the mechanism involved requires MLG degradation, which may in turn regulate MLGH1 gene expression.

Automated summary

In this study, a maize mutant named mlgh1 was identified with increased MLG content due to a mutation in the GRMZM2G137535 gene, which encodes a GH17 licheninase capable of degrading MLG. Overexpression of this gene resulted in a significant reduction in MLG content and increased saccharification yields. Moreover, cycling of MLG content during day/night cycles was disrupted in the mlgh1 mutant, suggesting a regulatory role of MLG degradation in gene expression.