The Cellulase KORRIGAN Is Part of the Cellulose Synthase Complex

Vain, Thomas; Crowell, Elizabeth; Timpano, Hélène; Biot, Eric; Desprez, Thierry; Mansoori, Nasim; Trindade, Luisa M.; Pagant, Silvère; Robert, Stéphanie; Höfte, Monica; Gonneau, Martine; Vernhettes, Samantha

doi:10.1104/pp.114.241216

Cited by 154 publications

(143 citation statements)

References 46 publications

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“…Plant CesAs have been shown to interact with several components, including cytosolic microtubule-binding proteins, and also with membrane-integrated or -attached factors, such as the β-1,4 glucanase KORRIGAN (33). However, the cellulose biosynthetic activity of recombinantly expressed and reconstituted CesA8 from hybrid aspen demonstrates that no other plant-specific proteinaceous or lipidic factors are required for its catalytic activity.…”

Section: Discussionmentioning

confidence: 97%

A single heterologously expressed plant cellulose synthase isoform is sufficient for cellulose microfibril formation in vitro

Purushotham

Cho

Díaz-Moreno

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Plant cell walls are a composite material of polysaccharides, proteins, and other noncarbohydrate polymers. In the majority of plant tissues, the most abundant polysaccharide is cellulose, a linear polymer of glucose molecules. As the load-bearing component of the cell wall, individual cellulose chains are frequently bundled into micro and macrofibrils and are wrapped around the cell. Cellulose is synthesized by membrane-integrated and processive glycosyltransferases that polymerize UDP-activated glucose and secrete the nascent polymer through a channel formed by their own transmembrane regions. Plants express several different cellulose synthase isoforms during primary and secondary cell wall formation; however, so far, none has been functionally reconstituted in vitro for detailed biochemical analyses. Here we report the heterologous expression, purification, and functional reconstitution of Populus tremula x tremuloides CesA8 (PttCesA8), implicated in secondary cell wall formation. The recombinant enzyme polymerizes UDP-activated glucose to cellulose, as determined by enzyme degradation, permethylation glycosyl linkage analysis, electron microscopy, and mutagenesis studies. Catalytic activity is dependent on the presence of a lipid bilayer environment and divalent manganese cations. Further, electron microscopy analyses reveal that PttCesA8 produces cellulose fibers several micrometers long that occasionally are capped by globular particles, likely representing PttCesA8 complexes. Deletion of the enzyme's N-terminal RING-finger domain almost completely abolishes fiber formation but not cellulose biosynthetic activity. Our results demonstrate that reconstituted PttCesA8 is not only sufficient for cellulose biosynthesis in vitro but also suffices to bundle individual glucan chains into cellulose microfibrils.biopolymer | cellulose | glycosyltransferase | plant cell wall | membrane transport

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Section: Discussionmentioning

confidence: 97%

A single heterologously expressed plant cellulose synthase isoform is sufficient for cellulose microfibril formation in vitro

Purushotham

Cho

Díaz-Moreno

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…This structural information was revealed by freeze-fracture transmission electron microscopy, showing six lobes in a hexagonal arrangement at the point where the transmembrane helices of multiple cellulose synthase proteins (CESAs) cross the plasma membrane. Recently KORRIGAN, a protein with cellulase activity, has also been implicated as an integral component of the CSC (Vain et al, 2014).…”

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confidence: 99%

A Structural Study of CESA1 Catalytic Domain of Arabidopsis Cellulose Synthesis Complex: Evidence for CESA Trimers

et al. 2015

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A cellulose synthesis complex with a "rosette" shape is responsible for synthesis of cellulose chains and their assembly into microfibrils within the cell walls of land plants and their charophyte algal progenitors. The number of cellulose synthase proteins in this large multisubunit transmembrane protein complex and the number of cellulose chains in a microfibril have been debated for many years. This work reports a low resolution structure of the catalytic domain of CESA1 from Arabidopsis (Arabidopsis thaliana; AtCESA1CatD) determined by small-angle scattering techniques and provides the first experimental evidence for the self-assembly of CESA into a stable trimer in solution. The catalytic domain was overexpressed in Escherichia coli, and using a two-step procedure, it was possible to isolate monomeric and trimeric forms of AtCESA1CatD. The conformation of monomeric and trimeric AtCESA1CatD proteins were studied using small-angle neutron scattering and small-angle x-ray scattering. A series of AtCESA1CatD trimer computational models were compared with the small-angle x-ray scattering trimer profile to explore the possible arrangement of the monomers in the trimers. Several candidate trimers were identified with monomers oriented such that the newly synthesized cellulose chains project toward the cell membrane. In these models, the class-specific region is found at the periphery of the complex, and the plant-conserved region forms the base of the trimer. This study strongly supports the "hexamer of trimers" model for the rosette cellulose synthesis complex that synthesizes an 18-chain cellulose microfibril as its fundamental product.

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“…Moreover, yeast split-ubiquitin interaction studies and in vivo bimolecular fluorescence complementation show that CESAs can form homodimers or heterodimers (Desprez et al, 2007;Timmers et al, 2009;Carroll et al, 2012). In addition to the CESAs, other proteins are associated at least with the primary cell wall CSC, such as the membrane-bound cellulase KORRIGAN1 (KOR1; Lei et al, 2014;Vain et al, 2014) and CELLULOSE SYNTHASE INTERACTING PRO-TEIN (CSI), which links CSCs to microtubules . The exact stoichiometry of the CESAs within the CSC is not known.…”

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confidence: 99%

Catalytic Subunit Stoichiometry within the Cellulose Synthase Complex

et al. 2014

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Cellulose synthesis is driven by large plasma membrane-inserted protein complexes, which in plants have 6-fold symmetry. In Arabidopsis (Arabidopsis thaliana), functional cellulose synthesis complexes (CSCs) are composed of at least three different cellulose synthase catalytic subunits (CESAs), but the actual ratio of the CESA isoforms within the CSCs remains unresolved. In this work, the stoichiometry of the CESAs in the primary cell wall CSC was determined, after elimination of CESA redundancy in a mutant background, by coimmunoprecipitation and mass spectrometry using label-free quantitative methods. Based on spectral counting, we show that CESA1, CESA3, and CESA6 are present in a 1:1:1 molecular ratio.

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The Cellulase KORRIGAN Is Part of the Cellulose Synthase Complex

Cited by 154 publications

References 46 publications

A single heterologously expressed plant cellulose synthase isoform is sufficient for cellulose microfibril formation in vitro

A single heterologously expressed plant cellulose synthase isoform is sufficient for cellulose microfibril formation in vitro

A Structural Study of CESA1 Catalytic Domain of Arabidopsis Cellulose Synthesis Complex: Evidence for CESA Trimers

Catalytic Subunit Stoichiometry within the Cellulose Synthase Complex

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