Abstract
The Reduced Wall Acetylation (RWA) proteins are involved in cell wall acetylation in plants. Previously, we described a single mutant, rwa2, which has about 20% lower level of O-acetylation in leaf cell walls and no obvious growth or developmental phenotype. In this study, we generated double, triple, and quadruple loss-of-function mutants of all four members of the RWA family in Arabidopsis (Arabidopsis thaliana). In contrast to rwa2, the triple and quadruple rwa mutants display severe growth phenotypes revealing the importance of wall acetylation for plant growth and development. The quadruple rwa mutant can be completely complemented with the RWA2 protein expressed under 35S promoter, indicating the functional redundancy of the RWA proteins. Nevertheless, the degree of acetylation of xylan, (gluco)mannan, and xyloglucan as well as overall cell wall acetylation is affected differently in different combinations of triple mutants, suggesting their diversity in substrate preference. The overall degree of wall acetylation in the rwa quadruple mutant was reduced by 63% compared with the wild type, and histochemical analysis of the rwa quadruple mutant stem indicates defects in cell differentiation of cell types with secondary cell walls.
Highlights
The Reduced Wall Acetylation (RWA) proteins are involved in cell wall acetylation in plants
REDUCED WALL ACETYLATION2 (RWA2) is a member of a small family consisting of four proteins in Arabidopsis, and its loss-of-function mutants display 20% reduction of acetylation in a range of polysaccharides that include XyG and pectins
We generated all combinations of double, triple, and quadruple rwa mutants to further dissect the role of RWA proteins in planta
Summary
The Reduced Wall Acetylation (RWA) proteins are involved in cell wall acetylation in plants. Plant cell walls are multifunctional viscoelastic networks mainly composed of polysaccharides Many of these polysaccharides, including xylans, (gluco)mannans, xyloglucans (XyGs), and pectins, have various degrees and patterns of acetyl esterification (Gille and Pauly, 2012; Pawar et al, 2013). RWA2 is a member of a small family consisting of four proteins in Arabidopsis, and its loss-of-function mutants display 20% reduction of acetylation in a range of polysaccharides that include XyG and pectins. We have generated all the combinations of double, triple, and quadruple mutants of all four members of RWA family to further investigate the functional diversity and redundancy and to explore the function of cell wall acetylation and the role of RWAs in the network of acetylation-related enzymes. RWAs have partially redundant functions in the process of cell wall acetylation and show distinct impacts upon different cell wall polysaccharides
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