AtBXL1 Encodes a Bifunctional β-d-Xylosidase/α-l-Arabinofuranosidase Required for Pectic Arabinan Modification in Arabidopsis Mucilage Secretory Cells      

Abstract

Following pollination, the epidermal cells of the Arabidopsis (Arabidopsis thaliana) ovule undergo a complex differentiation process that includes the synthesis and polar secretion of pectinaceous mucilage followed by the production of a secondary cell wall. Wetting of mature seeds leads to the rapid bursting of these mucilage secretory cells to release a hydrophilic gel that surrounds the seed and is believed to aid in seed hydration and germination. A novel mutant is identified where mucilage release is both patchy and slow and whose seeds display delayed germination. While developmental analysis of mutant seeds reveals no change in mucilage secretory cell morphology, changes in monosaccharide quantities are detected, suggesting the mucilage release defect results from altered mucilage composition. Plasmid rescue and cloning of the mutant locus revealed a T-DNA insertion in AtBXL1, which encodes a putative bifunctional b-D-xylosidase/a-L-arabinofuranosidase that has been implicated as a b-D-xylosidase acting during vascular development. Chemical and immunological analyses of mucilage extracted from bxl1 mutant seeds and antibody staining of developing seed coats reveal an increase in (1/5)-linked arabinans, suggesting that BXL1 is acting as an a-L-arabinofuranosidase in the seed coat. This implication is supported by the ability to rescue mucilage release through treatment of bxl1 seeds with exogenous a-L-arabinofuranosidases. Together, these results suggest that trimming of rhamnogalacturonan I arabinan side chains is required for correct mucilage release and reveal a new role for BXL1 as an a-L-arabinofuranosidase acting in seed coat development.

GALACTURONOSYLTRANSFERASE-LIKE5 Is Involved in the Production of Arabidopsis Seed Coat Mucilage

“…4M). The increased width of the columella is consistent with the phenotype seen in other mutants with severely (mum4 and ttg1) and moderately (men4) reduced mucilage production (Western et al, 2004;Arsovski et al, 2009). Besides the mucilage defect and changes in mucilage secretory cell structure, the atgatl5-1 mutant also shows less adhesion between the hexagonal seed coat epidermal cell walls and wild-type seeds, as evidenced by the appearance of two distinct walls side by side in some areas of the seed coat, rather than a single thick wall as seen for wild-type cells, in either the scanning electron microscopy images or in the calcofluor white-stained seeds (Fig.…”

“…CELLULOSE SYNTHASE5 (CESA5) was found to be involved in cellulose synthesis (Sullivan et al, 2011), while MUCILAGE-MODIFIED4/RHAMNOSE SYNTHESIS2 (MUM4/RHM2) encodes a UDP-Rha synthase (Usadel et al, 2004;Western et al, 2004;Oka et al, 2007). The mum2, atsbt1.7, atbxl1, pmei6, fly1, and per36 mutants were found to be defective in pectin modification (Dean et al, 2007;Macquet et al, 2007b;Rautengarten et al, 2008;Arsovski et al, 2009;Saez-Aguayo et al, 2013;Voiniciuc et al, 2013) or in the degradation of the outer cell wall of the outer integument (Kunieda et al, 2013), all of which show a mucilage-release defect.…”

SALT-OVERLY SENSITIVE5 Mediates Arabidopsis Seed Coat Mucilage Adherence and Organization through Pectins

“…JIM5 is specific for partially methylesterified HG (up to 40%; Vandenbosch et al, 1989;Knox et al, 1990;Willats et al, 2000), whereas CCRC-M36 is specific for the unsubstituted RG I backbone that is found in Arabidopsis seed mucilage (Young et al, 2008;Arsovski et al, 2009;Pattathil et al, 2010). CCRC-M36 strongly labeled the periphery of the mucilage halo in wild-type seeds ( Fig.…”

“…Galactans and arabinans are also present in mucilage, and their regulation by glycosidases is required for correct mucilage hydration (Dean et al, 2007;Macquet et al, 2007b;Arsovski et al, 2009). For example, b-XYLOSIDASE1 encodes a bifunctional b-D-xylosidase/a-L-arabinofuranosidase required for arabinan modification in mucilage, and b-xylosidase1 mutant seeds have a delayed mucilage release phenotype (Arsovski et al, 2009). MUCILAGE MODIFIED2 (MUM2) encodes a b-D-galactosidase, and mum2 seeds fail to release mucilage when hydrated in water (Dean et al, 2007;Macquet et al, 2007b).…”

“…Mucilage separates into two fractions: a loose nonadherent fraction and an inner adherent fraction that can only be released by vigorous shaking, strong bases, or glycosidases (for review, see North et al, 2014). Galactans and arabinans are also present in mucilage, and their regulation by glycosidases is required for correct mucilage hydration (Dean et al, 2007;Macquet et al, 2007b;Arsovski et al, 2009). For example, b-XYLOSIDASE1 encodes a bifunctional b-D-xylosidase/a-L-arabinofuranosidase required for arabinan modification in mucilage, and b-xylosidase1 mutant seeds have a delayed mucilage release phenotype (Arsovski et al, 2009).…”