Partial functional conservation of IRX10 homologs in physcomitrella patens and Arabidopsis thalianaindicates an evolutionary step contributing to vascular formation in land plants

Abstract

BackgroundPlant cell walls are complex multicomponent structures that have evolved to fulfil an essential function in providing strength and protection to cells. Hemicelluloses constitute a key component of the cell wall and recently a number of the genes thought to encode the enzymes required for its synthesis have been identified in Arabidopsis. The acquisition of hemicellulose synthesis capability is hypothesised to have been an important step in the evolution of higher plants.ResultsAnalysis of the Physcomitrella patens genome has revealed the presence of homologs for all of the Arabidopsis glycosyltransferases including IRX9, IRX10 and IRX14 required for the synthesis of the glucuronoxylan backbone. The Physcomitrella IRX10 homolog is expressed in a variety of moss tissues which were newly formed or undergoing expansion. There is a high degree of sequence conservation between the Physcomitrella IRX10 and Arabidopsis IRX10 and IRX10-L. Despite this sequence similarity, the Physcomitrella IRX10 gene is only able to partially rescue the Arabidopsis irx10 irx10-L double mutant indicating that there has been a neo- or sub-functionalisation during the evolution of higher plants. Analysis of the monosaccharide composition of stems from the partially rescued Arabidopsis plants does not show any significant change in xylose content compared to the irx10 irx10-L double mutant. Likewise, knockout mutants of the Physcomitrella IRX10 gene do not result in any visible phenotype and there is no significant change in monosaccharide composition of the cell walls.ConclusionsThe fact that the Physcomitrella IRX10 (PpGT47A) protein can partially complement an Arabidopsis irx10 irx10-L double mutant suggests that it shares some function with the Arabidopsis proteins, but the lack of a phenotype in knockout lines shows that the function is not required for growth or development under normal conditions in Physcomitrella. In contrast, the Arabidopsis irx10 and irx10 irx10-L mutants have strong phenotypes indicating an important function in growth and development. We conclude that the evolution of vascular plants has been associated with a significant change or adaptation in the function of the IRX10 gene family.

Asparagus IRX9, IRX10, and IRX14A Are Components of an Active Xylan Backbone Synthase Complex that Forms in the Golgi Apparatus

“…Among these, AtIRX9 and AtIRX14 (both GT43 family members) and AtIRX10 (GT47), as well as their functionally redundant homologs, are believed to be involved directly in xylan backbone biosynthesis and to form a complex in the GA (Rennie and Scheller 2014). Orthologs of the AtIRX proteins have been identified in many other species, including wheat (Zeng et al, 2010), Populus trichocarpa (Lee et al, 2012b), Physcomitrella patens (Hörnblad et al, 2013), Plantago ovata (Jensen et al, 2013), rice (Oryza sativa; Chen et al, 2013;Chiniquy et al, 2013), Gossypium hirsutum , Neolamarckia cadamba (Zhao et al, 2014), and garden asparagus (Asparagus officinalis; Song et al, 2015). Enzymes involved in xylan backbone biosynthesis as well as side chain decorations, such as the arabinosyltransferases, glucuronosyltransferases, and acetylation enzymes, were recently reviewed by Rennie and Scheller (2014) and Hao and Mohnen (2014).…”

Arabidopsis thaliana IRX10 and two related proteins from psyllium and Physcomitrella patens are xylan xylosyltransferases

“… Heterologously expressed and purified IRX10 protein has in vitro xylan xylosyltransferase activity, which results in a reaction product that co‐elutes with the β‐1,4 xylan oligosaccharide marker when analyzed by normal phase high‐performance liquid chromatography. Knockout of IRX10 and its redundant homolog IRX10‐L resulted in loss of xylan and loss of endogenous xylan synthase activity (Brown et al ., ; Wu et al ., ). No xylose‐(1→2)‐xylose or xylose‐(1→3)‐xylose linkages have been identified in xylan isolated from A. thaliana , indicating that no enzyme in A. thaliana catalyzes the formation of these glycosidic linkages. Hence, IRX10 likely catalyze xylose transfer to the O ‐4 position rather than to the O ‐2 or O ‐3 positions. IRX10 and PpIRX10 share a high level of amino acid sequence identity (77%), a level at which enzyme specificity is likely conserved. There are no know examples of two homologous glycosyltransferases where one displays a retaining reaction mechanism and the other displays an inverting reaction mechanism (Lombard et al ., ); thus, the linkage formed by IRX10 can be assumed to be a β‐glycosidic linkage, as proven for PpIRX10. Lastly, the presence of xylan in the cell walls of P. patens has been confirmed (Kulkarni et al ., ) and PpIRX10 has been shown to partially complement the A. thaliana irx10 irx10‐l knockout mutant (Hornblad et al ., ), suggesting functional conservation between IRX10 and PpIRX10 . …”

“…This tissue produces large amounts of highly branched xylans and expresses only homologs of IRX10 and not homologs of IRX9 or IRX14 (Jensen et al ., ). Moreover, homologs of IRX10 display a higher level of conservation across large evolutionary distance than homologs of either IRX9 or IRX14 (Hornblad et al ., ; Jensen et al ., ), suggesting that the function of this protein is conserved. These observations suggested to us that IRX10 alone might be capable of synthesizing the xylan backbone.…”

Ceratopteris richardii (C-fern): a model for investigating adaptive modification of vascular plant cell walls

“…As Harholt et al (2012) point out this is in direct contrast with an apparent lack of diversification and specialization within the cellulase synthase (CESA) superfamily. Homologs of IRX10, also involved in vascular formation in land plants, were found in the moss Physcomitrella patens and were recently reported to exhibit functional conservation with those from Arabidopsis (Hörnblad et al, 2013). Taken together these data suggest that at least some components of vascular tissues considered to be a “hallmark” of vascular plants (Weng et al, 2008), are not homologous between the lycophyte and euphyllophyte vascular plant lineages.…”