Pattern of Deposition of Cell Wall Polysaccharides and Transcript Abundance of Related Cell Wall Synthesis Genes during Differentiation in Barley Endosperm

Immunolabeling, combined with chemical analyses and transcript profiling, have provided a comprehensive temporal and spatial picture of the deposition and modification of cell wall polysaccharides during barley (Hordeum vulgare) grain development, from endosperm cellularization at 3 d after pollination (DAP) through differentiation to the mature grain at 38 DAP. (1 -> 3)-beta-D-Glucan appears transiently during cellularization but reappears in patches in the subaleurone cell walls around 20 DAP. (1 -> 3, 1 -> 4)-beta-Glucan, the most abundant polysaccharide of the mature barley grain, accumulates throughout development. Arabino-(1-4)-beta-D-xylan is deposited significantly earlier than we previously reported. This was attributable to the initial deposition of the polysaccharide in a highly substituted form that was not recognized by antibodies commonly used to detect arabino-(1-4)-beta-D-xylans in sections of plant material. The epitopes needed for antibody recognition were exposed by pretreatment of sections with alpha-L-arabinofuranosidase; this procedure showed that arabino-(1-4)-beta-D-xylans were deposited as early as 5 DAP and highlighted their changing structures during endosperm development. By 28 DAP labeling of hetero-(1 -> 4)-beta-D-mannan is observed in the walls of the starchy endosperm but not in the aleurone walls. Although absent in mature endosperm cell walls we now show that xyloglucan is present transiently from 3 until about 6 DAP and disappears by 8 DAP. Quantitative reverse transcription-polymerase chain reaction of transcripts for GLUCAN SYNTHASE-LIKE, Cellulose Synthase, and CELLULOSE SYNTHASE-LIKE genes were consistent with the patterns of polysaccharide deposition. Transcript profiling of some members from the Carbohydrate-Active Enzymes database glycosyl transferase families GT61, GT47, and GT43, previously implicated in arabino-(1-4)-beta-D-xylan biosynthesis, confirms their presence during grain development.