Hetero-trans-β-Glucanase Produces Cellulose–Xyloglucan Covalent Bonds in the Cell Walls of Structural Plant Tissues and Is Stimulated by Expansin

Klaus Herburger,Lenka Franková,Martina Pičmanová,Jia Wooi Loh,Marcos Valenzuela-Ortega,Frank Meulewaeter,Andrew D Hudson,Christopher E French,Stephen C Fry

doi:10.1016/j.molp.2020.04.011

Klaus Herburger, Lenka Franková + Show 7 more

Open Access

https://doi.org/10.1016/j.molp.2020.04.011

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Journal: Molecular plant	Publication Date: May 4, 2020
Citations: 33	License type: cc-by

Affiliation: University of Edinburgh

Abstract

Current cell-wall models assume no covalent bonding between cellulose and hemicelluloses such as xyloglucan or mixed-linkage β-d-glucan (MLG). However, Equisetum hetero-trans-β-glucanase (HTG) grafts cellulose onto xyloglucan oligosaccharides (XGOs) – and, we now show, xyloglucan polysaccharide – in vitro, thus exhibiting CXE (cellulose:xyloglucan endotransglucosylase) activity. In addition, HTG also catalyzes MLG-to-XGO bonding (MXE activity). In this study, we explored the CXE action of HTG in native plant cell walls and tested whether expansin exposes cellulose to HTG by disrupting hydrogen bonds. To quantify and visualize CXE and MXE action, we assayed the sequential release of HTG products from cell walls pre-labeled with substrate mimics. We demonstrated covalent cellulose–xyloglucan bonding in plant cell walls and showed that CXE and MXE action was up to 15% and 60% of total transglucanase action, respectively, and peaked in aging, strengthening tissues: CXE in xylem and cells bordering intercellular canals and MXE in sclerenchyma. Recombinant bacterial expansin (EXLX1) strongly augmented CXE activity in vitro. CXE and MXE action in living Equisetum structural tissues potentially strengthens stems, while expansin might augment the HTG-catalyzed CXE reaction, thereby allowing efficient CXE action in muro. Our methods will enable surveys for comparable reactions throughout the plant kingdom. Furthermore, engineering similar hetero-polymer formation into angiosperm crop plants may improve certain agronomic traits such as lodging tolerance.

Highlights

Plant cells are surrounded by cell walls, which are composites of complex polysaccharides and crucial for plant function and survival (Popper et al, 2011)
Monitoring extractable transglucanase activities from various Equisetum tissues by in-vitro assays using soluble xyloglucan or mixed-linkage b-D-glucan (MLG) or insoluble cellulose as donor substrates and [3H]XXXGol as acceptor showed that both the MXE:xyloglucan endotransglucosylase (XET) and the CXE:XET ratio increased with shoot age independently of the season
We show that endogenous cellulose can be covalently attached to xyloglucan oligosaccharides (XGOs) in living Equisetum tissues, and provide evidence that native cellulose can undergo enzymic covalent modifications in muro other than hydrolysis

Summary

Introduction

Plant cells are surrounded by cell walls, which are composites of complex polysaccharides and crucial for plant function and survival (Popper et al, 2011). Recent in-vitro studies have revealed that certain hetero-transglucanase activities exist that can catalyze the cleavage of a cellulose molecule (donor substrate) followed by its covalent attachment to a xyloglucan-oligosaccharide (XGO) (acceptor substrate; Simmons et al, 2015; Shinohara et al, 2017). This hetero-transglucosylation differs from homotransglucosylation of hemicelluloses (Fry et al, 1992), where the donor and acceptor substrate, usually xyloglucan, are chemically identical (Frankovaand Fry, 2013)

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