TY - JOUR

T1 - Cutin:xyloglucan transacylase (CXT) activity covalently links cutin to a plant cell-wall polysaccharide

AU - Xin, Anzhou

AU - Fry, Stephen C.

PY - 2021

Y1 - 2021

N2 - The shoot epidermal cell wall in land-plants is associated with a polyester, cutin, which controls water loss and possibly organ expansion. Covalent bonds between cutin and its neighbouring cell-wall polysaccharides have long been proposed. However, the lack of biochemical evidence makes cutin-polysaccharide linkages largely conjectural. Here we optimised a portfolio of radiochemical assays to look for cutin-polysaccharide ester bonds in the epidermis of pea epicotyls, ice-plant leaves and tomato fruits, based on the hypothesis that a transacylase remodels cutin in a similar fashion to cutin synthase and cutin:cutin transacylase activities. Through in-situ enzyme assays and chemical degradations coupled with chromatographic analysis of the 3H-labelled products, we observed that among several wall-related oligosaccharides tested, only a xyloglucan oligosaccharide ([3H] XXXGol) could acquire ester-bonds from endogenous cutin, suggesting a cutin:xyloglucan transacylase (CXT). CXT activity was heat-labile, time-dependent, and maximal at near-neutral pH values. In-situ CXT activity peaked in nearly fully expanded tomato fruits and ice-plant leaves. CXT activity positively correlated with organ growth rate, suggesting that it contributes to epidermal integrity during rapid expansion. This study uncovers hitherto unappreciated re-structuring processes in the plant epidermis and provides a step towards the identification of CXT and its engineering for biotechnological applications.

AB - The shoot epidermal cell wall in land-plants is associated with a polyester, cutin, which controls water loss and possibly organ expansion. Covalent bonds between cutin and its neighbouring cell-wall polysaccharides have long been proposed. However, the lack of biochemical evidence makes cutin-polysaccharide linkages largely conjectural. Here we optimised a portfolio of radiochemical assays to look for cutin-polysaccharide ester bonds in the epidermis of pea epicotyls, ice-plant leaves and tomato fruits, based on the hypothesis that a transacylase remodels cutin in a similar fashion to cutin synthase and cutin:cutin transacylase activities. Through in-situ enzyme assays and chemical degradations coupled with chromatographic analysis of the 3H-labelled products, we observed that among several wall-related oligosaccharides tested, only a xyloglucan oligosaccharide ([3H] XXXGol) could acquire ester-bonds from endogenous cutin, suggesting a cutin:xyloglucan transacylase (CXT). CXT activity was heat-labile, time-dependent, and maximal at near-neutral pH values. In-situ CXT activity peaked in nearly fully expanded tomato fruits and ice-plant leaves. CXT activity positively correlated with organ growth rate, suggesting that it contributes to epidermal integrity during rapid expansion. This study uncovers hitherto unappreciated re-structuring processes in the plant epidermis and provides a step towards the identification of CXT and its engineering for biotechnological applications.

KW - Epidermis remodelling

KW - Cutin

KW - Xyloglucan

KW - Ester bond

KW - Transacylase

KW - Cell expansion

KW - Polyester-polysaccharide conjugates

KW - LEAF CUTICLE

KW - BIOSYNTHESIS

KW - IDENTIFICATION

KW - DEGRADATION

KW - GLUCANASE

KW - BODYGUARD

KW - ENZYMES

KW - BONDS

KW - ACID

U2 - 10.1016/j.jplph.2021.153446

DO - 10.1016/j.jplph.2021.153446

M3 - Journal article

C2 - 34051591

VL - 262

JO - Journal of Plant Physiology

JF - Journal of Plant Physiology

SN - 0176-1617

M1 - 153446

ER -