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

The wall is the last frontier of a plant cell involved in modulating growth, development and defense against biotic stresses. Cellulose and additional polysaccharides of plant cell walls are the most abundant biopolymers on earth, having increased in economic value and thereby attracted significant interest in biotechnology. Cellulose biosynthesis constitutes a highly complicated process relying on the formation of cellulose synthase complexes. Cellulose synthase (CesA) and Cellulose synthase-like (Csl) genes encode enzymes that synthesize cellulose and most hemicellulosic polysaccharides. Arabidopsis and rice are invaluable genetic models and reliable representatives of land plants to comprehend cell wall synthesis. During the past two decades, enormous research progress has been made to understand the mechanisms of cellulose synthesis and construction of the plant cell wall. A plethora of cesa and csl mutants have been characterized, providing functional insights into individual protein isoforms. Recent structural studies have uncovered the mode of CesA assembly and the dynamics of cellulose production. Genetics and structural biology have generated new knowledge and have accelerated the pace of discovery in this field, ultimately opening perspectives towards cellulose synthesis manipulation. This review provides an overview of the major breakthroughs gathering previous and recent genetic and structural advancements, focusing on the function of CesA and Csl catalytic domain in plants.

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Endosidin20 targets cellulose synthase catalytic domain to inhibit cellulose biosynthesis

10.1101/2020.02.16.946244 ◽

2020 ◽

Cited By ~ 1

Author(s):

Lei Huang ◽

Xiaohui Li ◽

Weiwei Zhang ◽

Nolan Ung ◽

Nana Liu ◽

...

Keyword(s):

Amino Acids ◽

Plasma Membrane ◽

Catalytic Activity ◽

Catalytic Domain ◽

Cellulose Synthase ◽

Catalytic Site ◽

Cellulose Synthesis ◽

Spatiotemporal Resolution ◽

Chemical Genetic ◽

Biochemical Assays

AbstractCellulose is synthesized by rosette structured cellulose synthase (CESA) complexes (CSCs), each of which is composed of multiple units of CESAs in three different isoforms. CSCs rely on vesicle trafficking for delivery to the plasma membrane where they catalyze cellulose synthesis. Although the rosette structured CSCs were observed decades ago, it remains unclear what amino acids in plant CESA that directly participate in cellulose catalytic synthesis. It is also not clear how the catalytic activity of CSCs influences their efficient transport at the subcellular level. Here we report characterization of the small molecule Endosidin20 (ES20) and present evidence that it represents a new CESA inhibitor. We show data from chemical genetic analyses, biochemical assays, structural modeling, and molecular docking to support our conclusion that ES20 targets the catalytic site of Arabidopsis CESA6. Further, chemical genetic analysis reveals important amino acids that potentially form the catalytic site of plant CESA6. Using high spatiotemporal resolution live-cell imaging, we found that inhibition of CSC catalytic activity by inhibitor treatment, or by creating missense mutation at amino acids in the predicted catalytic site, causes reduced efficiency in CSC transport to the plasma membrane. Our results show that the catalytic activity of plant CSCs is integrated with subcellular trafficking dynamics.One sentence summaryEndosidin20 targets cellulose synthase at the catalytic site to inhibit cellulose synthesis and the inhibition of catalytic activity reduces cellulose synthase complex delivery to the plasma membrane.

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Endosidin20 is a broad-spectrum cellulose synthesis inhibitor with an herbicidal function

10.1101/2020.02.22.961094 ◽

2020 ◽

Author(s):

Lei Huang ◽

Chunhua Zhang

Keyword(s):

Cell Growth ◽

Plant Growth ◽

Broad Spectrum ◽

Plant Cell Wall ◽

Catalytic Domain ◽

Cellulose Synthesis ◽

Missense Mutations ◽

Cellulose Biosynthesis ◽

Synthesis Inhibitor ◽

Biosynthesis Inhibitor

AbstractCellulose is an important component of plant cell wall that controls anisotropic cell growth. Disruption of cellulose biosynthesis often leads to inhibited cell growth. Endosidin20 (ES20) was recently identified as a cellulose biosynthesis inhibitor (CBI) that targets the catalytic domain of Arabidopsis cellulose synthase 6 (CESA6) to inhibit plant growth. Here, we characterized the effects of ES20 on the growth of some other plant species and found that ES20 is a broad-spectrum plant growth inhibitor. We compared the inhibitory effects of ES20 and other CBIs on the growth of cesa6 plants that have reduced sensitivity to ES20. We found that most of the cesa6 with reduced sensitivity to ES20 show normal inhibited growth by other CBIs. ES20 also shows synergistic inhibitory effect on plant growth when applied together with other CBIs. We show ES20 has a different mode of action than tested CBIs isoxaben, indaziflam and C17. ES20 not only inhibits Arabidopsis growth under tissue culture condition, it inhibits plant growth under soil condition after direct spraying. We demonstrate that plants carrying two missense mutations can tolerate dual inhibition by ES20 and isoxaben.One sentence summaryCellulose biosynthesis inhibitor Endosidin20 has synergistic effect with other cellulose synthesis inhibitors and has the potential to be used as a spray herbicide.

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Structure of Arabidopsis CESA3 catalytic domain with its substrate UDP-glucose provides insight into the mechanism of cellulose synthesis

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2024015118 ◽

2021 ◽

Vol 118 (11) ◽

pp. e2024015118

Author(s):

Zhu Qiao ◽

Edwin R. Lampugnani ◽

Xin-Fu Yan ◽

Ghazanfar Abbas Khan ◽

Wuan Geok Saw ◽

...

Keyword(s):

Amino Acids ◽

Arabidopsis Thaliana ◽

Catalytic Domain ◽

Specific Region ◽

Cellulose Synthesis ◽

Uridine Diphosphate ◽

In Planta ◽

Core Domain ◽

Cellulose Synthases ◽

Insight Into

Cellulose is synthesized by cellulose synthases (CESAs) from the glycosyltransferase GT-2 family. In plants, the CESAs form a six-lobed rosette-shaped CESA complex (CSC). Here we report crystal structures of the catalytic domain of Arabidopsis thaliana CESA3 (AtCESA3CatD) in both apo and uridine diphosphate (UDP)-glucose (UDP-Glc)–bound forms. AtCESA3CatD has an overall GT-A fold core domain sandwiched between a plant-conserved region (P-CR) and a class-specific region (C-SR). By superimposing the structure of AtCESA3CatD onto the bacterial cellulose synthase BcsA, we found that the coordination of the UDP-Glc differs, indicating different substrate coordination during cellulose synthesis in plants and bacteria. Moreover, structural analyses revealed that AtCESA3CatD can form a homodimer mainly via interactions between specific beta strands. We confirmed the importance of specific amino acids on these strands for homodimerization through yeast and in planta assays using point-mutated full-length AtCESA3. Our work provides molecular insights into how the substrate UDP-Glc is coordinated in the CESAs and how the CESAs might dimerize to eventually assemble into CSCs in plants.

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Structural study of the catalytic domain of PKCzeta using infrared spectroscopy and two-dimensional infrared correlation spectroscopy

FEBS Journal ◽

10.1111/j.1742-4658.2006.05338.x ◽

2006 ◽

Vol 273 (14) ◽

pp. 3273-3286 ◽

Cited By ~ 8

Author(s):

Sonia Sanchez-Bautista ◽

Andris Kazaks ◽

Melanie Beaulande ◽

Alejandro Torrecillas ◽

Senena Corbalan-Garcia ◽

...

Keyword(s):

Infrared Spectroscopy ◽

Structural Study ◽

Catalytic Domain ◽

Correlation Spectroscopy ◽

Two Dimensional

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An Ultra-Structural Study of Human Melanoma in Vivo and Vitro

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100091275 ◽

1976 ◽

Vol 34 ◽

pp. 258-259

Author(s):

James R. Gaylor ◽

Fredda Schafer ◽

Robert E. Nordquist

Keyword(s):

Endoplasmic Reticulum ◽

Golgi Apparatus ◽

Protein Aggregation ◽

Structural Study ◽

Human Melanoma ◽

Protein Matrix ◽

Early Form ◽

Endoplasmic Reticulum Protein ◽

Mitochondrial Origin

Several theories on the origin of the melanosome exist. These include the Golgi origin theory, in which a tyrosinase-rich protein is "packaged" by the Golgi apparatus, thus forming the early form of the melanosome. A second theory postulates a mitochondrial origin of melanosomes. Its author contends that the melanosome is a modified mitochondria which acquires melanin during its development. A third theory states that a pre-melanosome is formed in the smooth or rough endoplasmic reticulum. Protein aggregation is suggested by one author as a possible source of the melanosome. This fourth theory postulates that the melanosome originates when the protein products of several genetic loci aggregate in the cytoplasm of the melanocyte. It is this protein matrix on which the melanin is deposited. It was with these theories in mind that this project was undertaken.

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Single molecule optical diffraction: A tool for understanding the structure of triple stranded left-hand helical cellulose

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100157103 ◽

1989 ◽

Vol 47 ◽

pp. 1024-1025

Author(s):

George C. Ruben ◽

William Krakow

Keyword(s):

Single Molecule ◽

Helical Structure ◽

Optical Diffraction ◽

Maximum Diameter ◽

Primary Cell Wall ◽

Cellulose Synthesis ◽

Left Hand ◽

Left Handed ◽

Freeze Dried ◽

Diffraction Patterns

Tobacco primary cell wall and normal bacterial Acetobacter xylinum cellulose formation produced a 36.8±3Å triple-stranded left-hand helical microfibril in freeze-dried Pt-C replicas and in negatively stained preparations for TEM. As three submicrofibril strands exit the wall of Axylinum , they twist together to form a left-hand helical microfibril. This process is driven by the left-hand helical structure of the submicrofibril and by cellulose synthesis. That is, as the submicrofibril is elongating at the wall, it is also being left-hand twisted and twisted together with two other submicrofibrils. The submicrofibril appears to have the dimensions of a nine (l-4)-ß-D-glucan parallel chain crystalline unit whose long, 23Å, and short, 19Å, diagonals form major and minor left-handed axial surface ridges every 36Å.The computer generated optical diffraction of this model and its corresponding image have been compared. The submicrofibril model was used to construct a microfibril model. This model and corresponding microfibril images have also been optically diffracted and comparedIn this paper we compare two less complex microfibril models. The first model (Fig. 1a) is constructed with cylindrical submicrofibrils. The second model (Fig. 2a) is also constructed with three submicrofibrils but with a single 23 Å diagonal, projecting from a rounded cross section and left-hand helically twisted, with a 36Å repeat, similar to the original model (45°±10° crossover angle). The submicrofibrils cross the microfibril axis at roughly a 45°±10° angle, the same crossover angle observed in microflbril TEM images. These models were constructed so that the maximum diameter of the submicrofibrils was 23Å and the overall microfibril diameters were similar to Pt-C coated image diameters of ∼50Å and not the actual diameter of 36.5Å. The methods for computing optical diffraction patterns have been published before.

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Structural aspects of tracheary element differentiation in suspension cultures of Zinnia elegans

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100155815 ◽

1989 ◽

Vol 47 ◽

pp. 768-769

Author(s):

C. H. Haigler ◽

A. W. Roberts

Keyword(s):

Tracheary Element ◽

Vesicle Fusion ◽

Zinnia Elegans ◽

Cellulose Synthesis ◽

Tracheary Elements ◽

Animal Cells ◽

Mesophyll Cells ◽

Fixation Techniques ◽

Localized Patterns ◽

Structural Aspects

Tracheary elements, the water-conducting cells in plants, are characterized by their reinforced walls that became thickened in localized patterns during differentiation (Fig. 1). The synthesis of this localized wall involves abundant secretion of Golgi vesicles that export preformed matrix polysaccharides and putative proteins involved in cellulose synthesis. Since the cells are not growing, some kind of endocytotic process must also occur. Many researchers have commented on where exocytosis occurs in relation to the thickenings (for example, see), but they based their interpretations on chemical fixation techniques that are not likely to provide reliable information about rapid processes such as vesicle fusion. We have used rapid freezing to more accurately assess patterns of vesicle fusion in tracheary elements. We have also determined the localization of calcium, which is known to regulate vesicle fusion in plant and animal cells.Mesophyll cells were obtained from immature first leaves of Zinnia elegans var. Envy (Park Seed Co., Greenwood, S.C.) and cultured as described previously with the following exceptions: (a) concentration of benzylaminopurine in the culture medium was reduced to 0.2 mg/l and myoinositol was eliminated; and (b) 1.75ml cultures were incubated in 22 x 90mm shell vials with 112rpm rotary shaking. Cells that were actively involved in differentiation were harvested and frozen in solidifying Freon as described previously. Fractures occurred preferentially at the cell/planchet interface, which allowed us to find some excellently-preserved cells in the replicas. Other differentiating cells were incubated for 20-30 min in 10(μM CTC (Sigma), an antibiotic that fluoresces in the presence of membrane-sequestered calcium. They were observed in an Olympus BH-2 microscope equipped for epi-fluorescence (violet filter package and additional Zeiss KP560 barrier filter to block chlorophyll autofluorescence).

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