CELLULOSE SYNTHASE-LIKE A2, a Glucomannan Synthase, Is Involved in Maintaining Adherent Mucilage Structure in Arabidopsis Seed

Abstract

Mannans are hemicellulosic polysaccharides that are considered to have both structural and storage functions in the plant cell wall. However, it is not yet known how mannans function in Arabidopsis (Arabidopsis thaliana) seed mucilage. In this study, CELLULOSE SYNTHASE-LIKE A2 (CSLA2; At5g22740) expression was observed in several seed tissues, including the epidermal cells of developing seed coats. Disruption of CSLA2 resulted in thinner adherent mucilage halos, although the total amount of the adherent mucilage did not change compared with the wild type. This suggested that the adherent mucilage in the mutant was more compact compared with that of the wild type. In accordance with the role of CSLA2 in glucomannan synthesis, csla2-1 mucilage contained 30% less mannosyl and glucosyl content than did the wild type. No appreciable changes in the composition, structure, or macromolecular properties were observed for nonmannan polysaccharides in mutant mucilage. Biochemical analysis revealed that cellulose crystallinity was substantially reduced in csla2-1 mucilage; this was supported by the removal of most mucilage cellulose through treatment of csla2-1 seeds with endo-b-glucanase. Mutation in CSLA2 also resulted in altered spatial distribution of cellulose and an absence of birefringent cellulose microfibrils within the adherent mucilage. As with the observed changes in crystalline cellulose, the spatial distribution of pectin was also modified in csla2-1 mucilage. Taken together, our results demonstrate that glucomannans synthesized by CSLA2 are involved in modulating the structure of adherent mucilage, potentially through altering cellulose organization and crystallization.Mannan polysaccharides are a complex set of hemicellulosic cell wall polymers that are considered to have both structural and storage functions. Based on the particular chemical composition of the backbone and the side chains, mannan polysaccharides are classified into four types: pure mannan, glucomannan, galactomannan, and galactoglucomannan (Moreira and Filho, 2008;Wang et al., 2012;Pauly et al., 2013). Each of these polysaccharides is composed of a b-1,4-linked backbone containing Man or a combination of Glc and Man residues. In addition, the mannan backbone can be substituted with side chains of a-1,6-linked Gal residues. Mannan polysaccharides have been proposed to cross link with cellulose and other hemicelluloses via hydrogen bonds (Fry, 1986;Iiyama et al., 1994;Obel et al., 2007;Scheller and Ulvskov, 2010). Furthermore, it has been reported that heteromannans with different levels of substitution can interact with cellulose in diverse ways (Whitney et al., 1998). Together, these observations indicate the complexity of mannan polysaccharides in the context of cell wall architecture.CELLULOSE SYNTHASE-LIKE A (CSLA) enzymes have been shown to have mannan synthase activity in vitro. These enzymes polymerize the b-1,4-linked backbone of mannans or glucomannans, depending on the substrates (GDP-Man and/or GDP-Glc) provided (Richmond and Some...

MUCILAGE-RELATED10 Produces Galactoglucomannan That Maintains Pectin and Cellulose Architecture in Arabidopsis Seed Mucilage

“…S2). Four muci10 alleles and the csla2-3 mutant, which has dense mucilage (Yu et al, 2014), displayed approximately 30% smaller mucilage capsules than the wild type (Fig. 2Q).…”

“…While most polymers had wild-type levels, the muci10-1 mucilage contained 38% less HM (Student's t test, P , 0.05). Unsubstituted glucomannan is the only known HM component of mucilage (Yu et al, 2014), although some of the available linkage data suggest the presence of GGM . Lower HM content in muci10-1 mucilage resulted from reductions in terminal-Gal (t-Gal), 4-Glc, 4-Man, and 4,6-Man (Student's t test, P , 0.05), with 81% less t-Gal as the most severe defect (Table II; Fig.…”

“…Previously, csla2-1 mucilage capsules were easily digested by an endo-b-1,4-glucanase from Aspergillus niger (Yu et al, 2014). A 90-min treatment with a similar b-glucanase, purified from Trichoderma longibrachiatum, fully detached csla2-3 and muci10-1 adherent mucilage but had minor effects on the wild type (Supplemental Fig.…”

“…Arabidopsis CSLA2, like most other isoforms, can use both GDP-Man and GDP-Glc as substrates in vitro (Liepman et al, 2005 and is responsible for stem glucomannan synthesis in vivo along with CSLA3 and CSLA7 (Goubet et al, 2009). CSLA2 also participates in the synthesis of glucomannan present in mucilage produced by seed coat epidermal (SCE) cells (Yu et al, 2014).…”

Monitoring Polysaccharide Dynamics in the Plant Cell Wall

“…The dynamics of mannan have yet to be explored in great detail, although new insights into their structural roles were gained in recent years (Yu et al, 2014;Voiniciuc et al, 2015a). Some mAb probes are available (e.g.…”