纤维素是最具经济价值的细胞壁成分，其高级结构与纤维素的理化性质和经济价值密切相关。然而纤维素的组装过程极其复杂，除了纤维素链自组装假说外，一直没有证据证明纤维素的组装需要一些关键蛋白的参与。另一方面，早在2001年已有报道表明COBRA蛋白参与了纤维素的生物合成过程，但其作用机理一直未知，使其成为本领域的“未解之谜”。
BC1是一个类COBRA蛋白，李家洋院士课题组在2003年首次报道了所编码基因的克隆及其影响次生壁纤维素合成的效应。近期，中科院遗传与发育生物研究所周奕华课题组与李家洋课题组及中国水稻所钱前课题组合作，从BC1的生化特性入手，利用生物化学、细胞生物学、遗传学、生物物理学、植物化学等多种手段，以充分的证据证明BC1蛋白是一个磷脂酰肌醇（GPI）锚定的膜蛋白，切除GPI锚定的BC1定位在细胞壁、尤其是次生壁上。BC1蛋白的N端有一个碳水化合物结合结构域（CBM）。体外生化分析和体内遗传学实验证实，该结构域可特异结合晶体化纤维素，是一个重要的功能域。此外还发现，BC1蛋白的细胞壁定位需要GPI和CBM结构域共同发挥作用。虽然BC1与CESAs基因高度共表达，但遗传学证据表明BC1的作用位置与CESA不同，可能在CESA蛋白合成纤维素葡聚糖链后发挥作用。利用广角X衍射(WAXS)方法分析了突变体和相关材料中纤维素的高级结构，证实该基因突变可导致纤维素微晶尺寸显著变小，而过量表达BC1可以提高纤维素的相对结晶度。表达CBM中特定芳香族氨基酸点突变的BC1变异形式可使转基因材料的纤维素微晶大小改变，并与其体外结合纤维素能力相符。
这一发现首次证明了纤维素的组装需要蛋白的参与。因此，BC1可作为改良植物纤维素性质的关键元件，其应用前景令人期待。
该项工作已于8月22日在PLoS Genetics杂志上在线发表(DOI：10.1371/journal.pgen.1003704)。周奕华课题组刘立锋和上官科科为该论文的共同第一作者。
该研究得到了“973”、国家自然科学基金和转基因专项的资助。（生物谷Bioon.com）
doi:10.1371/journal.pgen.1003704
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Brittle Culm1, a COBRA-Like Protein, Functions in Cellulose Assembly through Binding Cellulose Microfibrils
Lifeng Liu equal contributor, Keke Shang-Guan equal contributor, Baocai Zhang, Xiangling Liu, Meixian Yan, Lanjun Zhang, Yanyun Shi, Mu Zhang, Qian Qian, Jiayang Li, Yihua Zhou mail
Cellulose represents the most abundant biopolymer in nature and has great economic importance. Cellulose chains pack laterally into crystalline forms, stacking into a complicated crystallographic structure. However, the mechanism of cellulose crystallization is poorly understood. Here, via functional characterization, we report that Brittle Culm1 (BC1), a COBRA-like protein in rice, modifies cellulose crystallinity. BC1 was demonstrated to be a glycosylphosphatidylinositol (GPI) anchored protein and can be released into cell walls by removal of the GPI anchor. BC1 possesses a carbohydrate-binding module (CBM) at its N-terminus. In vitro binding assays showed that this CBM interacts specifically with crystalline cellulose, and several aromatic residues in this domain are essential for binding. It was further demonstrated that cell wall-localized BC1 via the CBM and GPI anchor is one functional form of BC1. X-ray diffraction (XRD) assays revealed that mutations in BC1 and knockdown of BC1 expression decrease the crystallite width of cellulose; overexpression of BC1 and the CBM-mutated BC1s caused varied crystallinity with results that were consistent with the in vitro binding assay. Moreover, interaction between the CBM and cellulose microfibrils was largely repressed when the cell wall residues were pre-stained with two cellulose dyes. Treating wild-type and bc1 seedlings with the dyes resulted in insensitive root growth responses in bc1 plants. Combined with the evidence that BC1 and three secondary wall cellulose synthases (CESAs) function in different steps of cellulose production as revealed by genetic analysis, we conclude that BC1 modulates cellulose assembly by interacting with cellulose and affecting microfibril crystallinity.
