An oligogalacturonide‐derived molecular probe demonstrates the dynamics of calcium‐mediated pectin complexation in cell walls of tip‐growing structures

Abstract

Pectic homogalacturonan (HG) is one of the main constituents of plant cell walls. When processed to low degrees of esterification, HG can form complexes with divalent calcium ions. These macromolecular structures (also called egg boxes) play an important role in determining the biomechanics of cell walls and in mediating cell-to-cell adhesion. Current immunological methods enable only steady-state detection of egg box formation in situ. Here we present a tool for efficient real-time visualisation of available sites for HG crosslinking within cell wall microdomains. Our approach is based on calcium-mediated binding of fluorescently tagged long oligogalacturonides (OGs) with endogenous de-esterified HG. We established that more than seven galacturonic acid residues in the HG chain are required to form a stable complex with endogenous HG through calcium complexation in situ, confirming a recently suggested thermodynamic model. Using defined carbohydrate microarrays, we show that the long OG probe binds exclusively to HG that has a very low degree of esterification and in the presence of divalent ions. We used this probe to study real-time dynamics of HG during elongation of Arabidopsis pollen tubes and root hairs. Our results suggest a different spatial organisation of incorporation and processing of HG in the cell walls of these two tip-growing structures.

Monitoring Polysaccharide Dynamics in the Plant Cell Wall

“…Sequential use of COS probes coupled to two distinct fluorophores revealed how cells in Arabidopsis root caps accumulate deesterified HG over time. Recently, another oligosaccharide probe consisting only of GalA units was used to continuously monitor the distribution of calcium-cross-linked HG in elongating pollen tubes (Mravec et al, 2017). Throughout pollen tube growth, tightly linked HG was only detected outside of the tipgrowing region, consistent with the polar localization of PMEI proteins (Röckel et al, 2008).…”

Cell wall traits that influence plant development, immunity, and bioconversion

“…The growth of pollen tubes and root hairs is particularly amenable to study CW dynamics and the complex interplay between synthesis and re‐modeling of the CW polysaccharides (Mravec et al ., 2017a; Cameron and Geitmann, ; Tucker et al ., ). Control of pectin synthesis, de‐methylesterification and assembly is essential to prevent bursting of the growing cells caused by a turgor pressure not properly balanced by the CW tensile strength, with consequent perturbation of the cell wall integrity (CWI).…”

Release, Recycle, Rebuild: Cell-Wall Remodeling, Autodegradation, and Sugar Salvage for New Wall Biosynthesis during Plant Development

“…Recent advances in biochemical and microscopic tools and techniques provide new avenues to explore the physiological roles of cell-wall recycling through more targeted analyses of wall degradation and sugar salvage. Recently, a set of fluorescently labeled chitin oligosaccharide and oligogalacturonide probes was described with the intended purpose of labeling negatively charged, de-methyl-esterified HG (Mravec et al, 2014(Mravec et al, , 2017. Pulse-chase experiments with these probes or fluorescently tagged XGOs, coupled with confocal microscopy, might allow researchers to track the route of endocytosed hemicellulose or pectins in real time.…”