Abstract:Plant cell wall (CW) synthesizing enzymes can be divided into the glycan (i.e. cellulose and callose) synthases, which are multimembrane spanning proteins located at the plasma membrane, and the glycosyltransferases (GTs), which are Golgi localized single membrane spanning proteins, believed to participate in the synthesis of hemicellulose, pectin, mannans, and various glycoproteins. At the Carbohydrate-Active enZYmes (CAZy) database where e.g. glucoside hydrolases and GTs are classified into gene families pri… Show more
“…This domain is highly conserved between KOBITO1 homologs in land plants and green algae but is not present in other organisms. While KOBITO1 is not part of CAZy, a database of carbohydrate active enzymes, a bioinformatics approach aimed at finding additional plant glycosyltransferase genes has identified KOBITO1 as one of 27 non-CAZy-classified putative glycosyltransferases (Egelund et al, 2004). Two genes out of this group of 27 have been proven to encode glycosyltransferases that function in pectin biosynthesis (Egelund et al, 2006).…”
The differentiation of stomata provides a convenient model for studying pattern formation in plant tissues. Stomata formation is induced by a set of basic helix-loop-helix transcription factors and inhibited by a signal transduction pathway initiated by TOO MANY MOUTHS (TMM) and ERECTA family (ERf) receptors. The formation of a proper stomata pattern is also dependent upon the restriction of symplastic movement of basic helix-loop-helix transcription factors into neighboring cells, especially in the backgrounds where the function of the TMM/ERf signaling pathway is compromised. Here, we describe a novel mutant of KOBITO1 in Arabidopsis (Arabidopsis thaliana). The kob1-3 mutation leads to the formation of stomata clusters in the erl1 erl2 background but not in the wild type. Cell-to-cell mobility assays demonstrated an increase in intercellular protein trafficking in kob1-3, including increased diffusion of SPEECHLESS, suggesting that the formation of stomata clusters is due to an escape of cell fate-specifying factors from stomatal lineage cells. While plasmodesmatal permeability is increased in kob1-3, we did not detect drastic changes in callose accumulation at the neck regions of the plasmodesmata. Previously, KOBITO1 has been proposed to function in cellulose biosynthesis. Our data demonstrate that disruption of cellulose biosynthesis in the erl1 erl2 background does not lead to the formation of stomata clusters, indicating that cellulose biosynthesis is not a major determining factor for regulating plasmodesmatal permeability. Analysis of KOBITO1 structure suggests that it is a glycosyltransferase-like protein. KOBITO1 might be involved in a carbohydrate metabolic pathway that is essential for both cellulose biosynthesis and the regulation of plasmodesmatal permeability.
“…This domain is highly conserved between KOBITO1 homologs in land plants and green algae but is not present in other organisms. While KOBITO1 is not part of CAZy, a database of carbohydrate active enzymes, a bioinformatics approach aimed at finding additional plant glycosyltransferase genes has identified KOBITO1 as one of 27 non-CAZy-classified putative glycosyltransferases (Egelund et al, 2004). Two genes out of this group of 27 have been proven to encode glycosyltransferases that function in pectin biosynthesis (Egelund et al, 2006).…”
The differentiation of stomata provides a convenient model for studying pattern formation in plant tissues. Stomata formation is induced by a set of basic helix-loop-helix transcription factors and inhibited by a signal transduction pathway initiated by TOO MANY MOUTHS (TMM) and ERECTA family (ERf) receptors. The formation of a proper stomata pattern is also dependent upon the restriction of symplastic movement of basic helix-loop-helix transcription factors into neighboring cells, especially in the backgrounds where the function of the TMM/ERf signaling pathway is compromised. Here, we describe a novel mutant of KOBITO1 in Arabidopsis (Arabidopsis thaliana). The kob1-3 mutation leads to the formation of stomata clusters in the erl1 erl2 background but not in the wild type. Cell-to-cell mobility assays demonstrated an increase in intercellular protein trafficking in kob1-3, including increased diffusion of SPEECHLESS, suggesting that the formation of stomata clusters is due to an escape of cell fate-specifying factors from stomatal lineage cells. While plasmodesmatal permeability is increased in kob1-3, we did not detect drastic changes in callose accumulation at the neck regions of the plasmodesmata. Previously, KOBITO1 has been proposed to function in cellulose biosynthesis. Our data demonstrate that disruption of cellulose biosynthesis in the erl1 erl2 background does not lead to the formation of stomata clusters, indicating that cellulose biosynthesis is not a major determining factor for regulating plasmodesmatal permeability. Analysis of KOBITO1 structure suggests that it is a glycosyltransferase-like protein. KOBITO1 might be involved in a carbohydrate metabolic pathway that is essential for both cellulose biosynthesis and the regulation of plasmodesmatal permeability.
“…This suggests that glycosyltransferase may act across plant species as a defense to very different pathogens. Glycosyltransferases also play a role in cell wall synthesis (Lao et al, 2003;Egelund et al, 2004) and may suggest a role in defense to RKN via this function. Research to better understand the role of this enzyme in RKN resistance is currently in progress.…”
Section: Discussionmentioning
confidence: 99%
“…Experimental downregulation of this gene via virus-induced gene silencing (VIGS) restores susceptibility to M. incognita in 'Motelle,' indicating that this function is necessary for Mi-mediated resistance. Glycosyltransferases have been implicated in carbohydrate biosynthesis and associated in plant stress and defense responses (Dixon, 2001;Qi et al, 2005;Vogt and Jones, 2000) and cell wall synthesis (Egelund et al, 2004;Lao et al, 2003); this is the first report, to our knowledge, of a role for a glycosyltransferase in nematode resistance.…”
Root-knot nematode (RKN; Meloidogyne spp.) is a major crop pathogen worldwide. Effective resistance exists for a few plant species, including that conditioned by Mi in tomato (Solanum lycopersicum). We interrogated the root transcriptome of the resistant (Mi1) and susceptible (Mi-) cultivars 'Motelle' and 'Moneymaker,' respectively, during a time-course infection by the Mi-susceptible RKN species Meloidogyne incognita and the Mi-resistant species Meloidogyne hapla. In the absence of RKN infection, only a single significantly regulated gene, encoding a glycosyltransferase, was detected. However, RKN infection influenced the expression of broad suites of genes; more than half of the probes on the array identified differential gene regulation between infected and uninfected root tissue at some stage of RKN infection. We discovered 217 genes regulated during the time of RKN infection corresponding to establishment of feeding sites, and 58 genes that exhibited differential regulation in resistant roots compared to uninfected roots, including the glycosyltransferase. Using virus-induced gene silencing to silence the expression of this gene restored susceptibility to M. incognita in 'Motelle,' indicating that this gene is necessary for resistance to RKN. Collectively, our data provide a picture of global gene expression changes in roots during compatible and incompatible associations with RKN, and point to candidates for further investigation.
“…Position of the predicted TMD is indicated by the line (dotted line for RGXT1 and solid line for RGXT2; Egelund et al, 2004), and the DxD motif involved in the binding of UDP-sugar is indicated with asterisks above the sequence alignment. 1 online).…”
Section: Rgxt1 and Rgxt2 Expressed In Baculovirus-transfected Insect mentioning
confidence: 99%
“…In addition, the very limited number of donor and especially acceptor substrates has further hampered the identification of novel plant CW GTs. As a significant proportion of Arabidopsis GTs probably remains to be classified, we developed a bioinformatic filtering strategy aimed at identifying unclassified CW GTs in Arabidopsis (Egelund et al, 2004). As a result, 27 putative GTs were identified, and two of these gave rise to a new family in CAZy (GT-family-77; Coutinho et al, 2003a; http://afmb.cnrs-mrs.fr/CAZY/).…”
Section: Gene Identification and Deduced Protein Structurementioning
Two homologous plant-specific Arabidopsis thaliana genes, RGXT1 and RGXT2, belong to a new family of glycosyltransferases (CAZy GT-family-77) and encode cell wall (1,3)-a-D-xylosyltransferases. The deduced amino acid sequences contain single transmembrane domains near the N terminus, indicative of a type II membrane protein structure. Soluble secreted forms of the corresponding proteins expressed in insect cells showed xylosyltransferase activity, transferring D-xylose from UDPa-D-xylose to L-fucose. The disaccharide product was hydrolyzed by a-xylosidase, whereas no reaction was catalyzed by b-xylosidase. Furthermore, the regio-and stereochemistry of the methyl xylosyl-fucoside was determined by nuclear magnetic resonance to be an a-(1,3) linkage, demonstrating the isolated glycosyltransferases to be (1,3)-a-D-xylosyltransferases. This particular linkage is only known in rhamnogalacturonan-II, a complex polysaccharide essential to vascular plants, and is conserved across higher plant families. Rhamnogalacturonan-II isolated from both RGXT1 and RGXT2 T-DNA insertional mutants functioned as specific acceptor molecules in the xylosyltransferase assay. Expression of RGXT1-and RGXT2-enhanced green fluorescent protein constructs in Arabidopsis revealed that both fusion proteins were targeted to a Brefeldin A-sensitive compartment and also colocalized with the Golgi marker dye BODIPY TR ceramide, consistent with targeting to the Golgi apparatus. Taken together, these results suggest that RGXT1 and RGXT2 encode Golgi-localized (1,3)-a-D-xylosyltransferases involved in the biosynthesis of pectic rhamnogalacturonan-II.
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