Researchers Discover the Core Components of Xylan Synthase Complex in Rice

Shaping plant architecture involves the precise assembly of structural polysaccharides, which are cross-linked into complex networks that are essential for mechanical support and morphogenesis as well as biomass recalcitrance. Xylan is a major interlocking polysaccharide in the cell walls of seed plants, possessing a high structural diversity and cellular specificity in the degree of chain polymerization, side chains and modifications. Xylans also incorporate most of acetyl esters in the cell wall, which patterns determine xylan folding, thereby governing the interlocking with cellulose, lignin, and other wall components. Plants have thus evolved complex mechanisms to precisely control the xylan synthesis at multiple levels, including the formation of protein complexes. However, the biochemical mechanism of xylan synthase complexes (XSCs) and the functions of core components within XSC remain unclear.

 

In a study published in The Plant Cell (DOI: https://doi.org/10.1093/plcell/koae322), ZHANG Baocai's group and ZHOU Yihua's group from the Institute of Genetics and Developmental Biology of the Chinese Academy of Sciences and the collaborators reported that XYLAN O-ACETYLTRANSFERASE 6 promotes xylan polymerization and folding by forming a complex with IRX10 and governs wall strength and recalcitrance in rice.

 

Identifying a complex of transmembrane proteins is always a challenging task. After several years of effort and numerous attempts, the authors successfully identified the components of XSC by performing co-fractionation mass spectrometry assay using the membrane proteins extracted from rice internodes. Xylan O-acetyltransferase 6 (XOAT6) was found co-fractionated with IRRERULAR XYLEM10 (IRX10), a verified xylan synthase in rice. Further experiments, including luciferase complementation and co-immunoprecipitation assays, confirmed that IRX10 and XOAT6 interact directly. Genetic studies revealed that the mutants deficient in IRX10 and XOAT6 exhibited similar phenotypes, such as a reduction in acetyl ester and xylose content, leading to brittleness. The double mutant exhibited additive effects on the above phenotypes, providing genetic proofs for that XOAT6 and IRX10 are critical components of the XSC.

 

A series of in vitro biochemical analyses demonstrated that XOAT6 is an authentic xylan acetyltransferase. More interestingly, the author unexpectedly found that the recombinant XOAT6 protein can enhance xylan backbone elongation mediated by IRX10, and this promotion is at least not fully dependent on its acetyltransferase activity. To obtain evidence at a single-molecule level, fluorescence correlation spectroscopy approach was further developed to visualize the xylooligomer polymerization process. Moreover, solid-state nuclear magnetic resonance spectroscopy, field emission scanning electron microscopy, and nanoindentation analyses revealed that XOAT6 and IRX10 play important roles in xylan folding and cellulose nanofibril organization, thereby facilitating cell wall mechanical strength. Additionally, mutations in XOAT6 and/or IRX10 significantly improved the saccharification efficiency in the assays without acid pretreatment, indicating potentials for diverse uses.

 

Therefore, this study reveals the synergistic mechanism of xylan biosynthesis, outlining the coordination of backbones polymerization and acetylation modifications, and offers a tool for crop trait design, biomass utilization, and artificial synthesis of polysaccharides.

 

 

Contact:

Prof. ZHANG Baocai

Institute of Genetics and Developmental Biology, Chinese Academy of Sciences

E-mail: bczhang@genetics.ac.cn