Xyloglucan-pectin linkages are formed intra-protoplasmically, contribute to wall-assembly, and remain stable in the cell wall

Zoe A. Popper, Stephen C. Fry

Research output: Contribution to journalArticlepeer-review

Abstract

We tested two hypotheses for the mechanism by which xyloglucan-pectin covalent bonds are formed in Arabidopsis cell cultures. Hypothesis 1 proposed hetero-transglycosylation, with xyloglucan as donor substrate and a rhamnogalacturonan-I (RG-I) side-chain as acceptor. We looked for enzyme activities that catalyse this reaction using alpha-(1 -> 5)-L-[H-3]arabino- or beta-(1 -> 4)-L-[H-3]galacto-oligosaccharides as model acceptor substrates. The H-3-oligosaccharides were supplied (with or without added xyloglucans) to living Arabidopsis cell-cultures, permeabilised cells, cell-free extracts, or four authentic XTHs. No hetero-transglycosylation occurred. Therefore, we cannot support hypothesis 1. Hypothesis 2 proposed that some xyloglucan is manufactured de novo as a side-chain on RG-I. To test this, we pulse-labelled Arabidopsis cell-cultures with [H-3]arabinose and monitored the radiolabelling of anionic (pectin-bonded) xyloglucan, which was resolved from free xyloglucan by ion-exchange chromatography. [H-3]Xyloglucan-pectin complexes were detectable < 4 min after [H-3]arabinose feeding, which is shorter than the transit-time for polysaccharide secretion, indicating that xyloglucan-pectin bonds were formed intra-protoplasmically. Thereafter, the proportion of the wall-bound [H-3]xyloglucan that was anionic remained almost constant at similar to 50% for >= 6 days, showing that the xyloglucan-pectin bond was stable in vivo. Some [H-3]xyloglucan was rapidly sloughed into the medium instead of becoming wall-bound. Only similar to 30% of the sloughed [H-3]xyloglucan was anionic, indicating that bonding to pectin promoted the integration of xyloglucan into the wall. We conclude that similar to 50% of xyloglucan in cultured Arabidopsis cells is synthesised on a pectic primer, then secreted into the apoplast, where the xyloglucan-pectin bonds are stable and the pectic moiety aids wall-assembly.

Original languageEnglish
Pages (from-to)781-794
Number of pages14
JournalPlanta
Volume227
Issue number4
DOIs
Publication statusPublished - Mar 2008

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