Group III-AXTHGenes of Arabidopsis Encode Predominant Xyloglucan Endohydrolases That Are Dispensable for Normal Growth      

Abstract

The molecular basis of primary wall extension endures as one of the central enigmas in plant cell morphogenesis. Classical cell wall models suggest that xyloglucan endo-transglycosylase activity is the primary catalyst (together with expansins) of controlled cell wall loosening through the transient cleavage and religation of xyloglucan-cellulose cross links. The genome of Arabidopsis (Arabidopsis thaliana) contains 33 phylogenetically diverse XYLOGLUCAN ENDO-TRANSGLYCOSYLASE/ HYDROLASE (XTH) gene products, two of which were predicted to be predominant xyloglucan endohydrolases due to clustering into group III-A. Enzyme kinetic analysis of recombinant AtXTH31 confirmed this prediction and indicated that this enzyme had similar catalytic properties to the nasturtium (Tropaeolum majus) xyloglucanase1 responsible for storage xyloglucan hydrolysis during germination. Global analysis of Genevestigator data indicated that AtXTH31 and the paralogous AtXTH32 were abundantly expressed in expanding tissues. Microscopy analysis, utilizing the resorufin b-glycoside of the xyloglucan oligosaccharide XXXG as an in situ probe, indicated significant xyloglucan endohydrolase activity in specific regions of both roots and hypocotyls, in good correlation with transcriptomic data. Moreover, this hydrolytic activity was essentially completely eliminated in AtXTH31/AtXTH32 double knockout lines. However, single and double knockout lines, as well as individual overexpressing lines, of AtXTH31 and AtXTH32 did not demonstrate significant growth or developmental phenotypes. These results suggest that although xyloglucan polysaccharide hydrolysis occurs in parallel with primary wall expansion, morphological effects are subtle or may be compensated by other mechanisms. We hypothesize that there is likely to be an interplay between these xyloglucan endohydrolases and recently discovered apoplastic exo-glycosidases in the hydrolytic modification of matrix xyloglucans.

Structure-Function Analysis of a Broad Specificity Populus trichocarpa Endo-β-glucanase Reveals an Evolutionary Link between Bacterial Licheninases and Plant XTH Gene Products

“…Deletion of this loop significantly increased the transglycosylation/hydrolysis ratio of the enzyme, making it structurally and functionally more XET-like (11). This loop insertion is likewise found in two homologous A. thaliana XEHs of Group III-A (12).…”

“…S5) previously observed for plant and microbial XEHs (12,18). In addition to the mixture of oligosaccharides expected from cleavage at the unbranched ␤(134)- Table 2, and gymnosperm transcripts in Group III-A are denoted by their accession number at TIGR (24).…”

“…This is due to the remarkable ability of some gene products to catalyze matrix polysaccharide rearrangement with essentially no chain hydrolysis (XET activity, EC 2.4.1.207). On the other hand, a limited number of XTH genes encode predominant hydrolases (XEH, EC 3.2.1.151) that operate in germinating seeds, ripening fruit, or expanding tissues (11,12). Notably, contemporary enzyme structure-function analyses have led to a refined phylogenetic delineation of the disparate transglycosylating and hydrolytic activities individual XTH gene products, which number in the range of ϳ20 -60 in diverse plants (11,13).…”

Xyloglucan Endotransglucosylase-Hydrolase17 Interacts with Xyloglucan Endotransglucosylase-Hydrolase31 to Confer Xyloglucan Endotransglucosylase Action and Affect Aluminum Sensitivity in Arabidopsis

“…For example, XTH27 is involved in the cell wall modification of tracheary elements at a specific stage of rosette leaf development and is essential for tertiary vein development (Matsui et al, 2005), whereas XTH31 is involved in cell wall modification and cell elongation through modulating xyloglucan endotransglucosylase (XET) action under Al stress (Zhu et al, 2012). However, XTH31 is an XTH for which xyloglucan endohydrolase (XEH) activity has been predicted (Baumann et al, 2007), and in our previous report, we demonstrated that XTH31 produced heterologously in Pichia pastoris has high XEH activity but low XET activity in vitro (Zhu et al, 2012), which is in accordance with Kaewthai et al (2013), who reported that XTH31 is a predominant hydrolase using the in vitro activity assays and enzyme product analysis, as well as the use of a fluorogenic substrate in vivo. Unexpectedly, however, the xth31 mutant has very low XET action and activity (Zhu et al, 2012).…”

Soluble and Membrane-Bound β-Glucosidases Are Involved in Trimming the Xyloglucan Backbone

“…These activities could modify xyloglucan chain length as well as its connections to microfibrils and have been shown to alter cell wall extension (Nishitani and Vissenberg, 2007;Miedes et al, 2013;Park and Cosgrove, 2015). The family also includes enzymes that show predominantly endohydrolase activity, using water as the final acceptor, but their physiological role is still unclear (Kaewthai et al, 2013). It is also unclear if xyloglucan can be hydrolyzed by plant endoglucanases from other families.…”