Essential amino acids in the Plant-Conserved and Class-Specific Regions of cellulose synthases
Essential amino acids in the Plant-Conserved and Class-Specific Regions of cellulose synthases
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DOI：
10.1093/plphys/kiac479
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发表时间：
2022-10-17
期刊：
影响因子：
7.4
通讯作者：
Carpita，Nicholas C.
中科院分区：
文献类型：
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作者：
Olek，Anna T.;Rushton，Phillip S.;Carpita，Nicholas C.
植物保守区（Plant-Conserved Region，P-CR）和类特异性区（Class-Specific Region，CSR）是纤维素酶催化核心中的两个植物特有序列，其功能尚未确定。在这里，我们使用定点突变来取代这些序列中的氨基酸和基序预测是必不可少的CESA的组装和功能。我们开发了一种体内方法，以确定突变的CesA 1转基因的能力，以补充拟南芥（拟南芥）的温度敏感根膨胀1（rsw 1）突变体。CSR中的Cys残基的替换，在体外阻断了二聚化，使得AtCesA 1转基因不能补充其sw 1突变。检查的CSR序列从33个不同的被子植物物种显示领域的高序列的保守性在一个类特异性的方式，但与变化的程度的障碍，表明非冗余的作用的CSR结构在不同的CESA亚型类。二聚化所必需的Cys残基并不总是位于内在紊乱的结构域。AtCesA 1转基因构建体的表达，其中Pro417和Arg 453取代了P-CR卷曲螺旋中的Ala或Lys，也不能补充其sw 1突变。尽管Arg 457在CESA蛋白的三聚化中具有预期的作用，但具有Arg 457 Ala突变的AtCesA 1转基因能够完全恢复野生型表型inrsw 1。我们的数据支持拟南芥CESA 1的CSR中的Cys 662和P-CR中的Pro 417和Arg 453是功能性合成酶复合物形成的必需残基，但我们的数据不支持Arg 457在天然CESA复合物的三聚化中的特定作用。
The Plant-Conserved Region (P-CR) and the Class-Specific Region (CSR) are two plant-unique sequences in the catalytic core of cellulose synthases (CESAs) for which specific functions have not been established. Here, we used site-directed mutagenesis to replace amino acids and motifs within these sequences predicted to be essential for assembly and function of CESAs. We developed an in vivo method to determine the ability of mutatedCesA1transgenes to complement an Arabidopsis (Arabidopsis thaliana) temperature-sensitiveroot-swelling1(rsw1) mutant. Replacement of a Cys residue in the CSR, which blocks dimerization in vitro, rendered theAtCesA1transgene unable to complement thersw1mutation. Examination of the CSR sequences from 33 diverse angiosperm species showed domains of high-sequence conservation in a class-specific manner but with variation in the degrees of disorder, indicating a nonredundant role of the CSR structures in different CESA isoform classes. The Cys residue essential for dimerization was not always located in domains of intrinsic disorder. Expression ofAtCesA1transgene constructs, in which Pro417and Arg453were substituted for Ala or Lys in the coiled-coil of the P-CR, were also unable to complement thersw1mutation. Despite an expected role for Arg457in trimerization of CESA proteins,AtCesA1transgenes with Arg457Ala mutations were able to fully restore the wild-type phenotype inrsw1. Our data support that Cys662within the CSR and Pro417and Arg453within the P-CR of Arabidopsis CESA1 are essential residues for functional synthase complex formation, but our data do not support a specific role for Arg457in trimerization in native CESA complexes.