A Complementary Bioinformatics Approach to Identify Potential Plant Cell Wall Glycosyltransferase-Encoding Genes  

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 primarily based on amino acid sequence similarities, 415 Arabidopsis GTs have been classified. Although much is known with regard to composition and fine structures of the plant CW, only a handful of CW biosynthetic GT genes-all classified in the CAZy system-have been characterized. In an effort to identify CW GTs that have not yet been classified in the CAZy database, a simple bioinformatics approach was adopted. First, the entire Arabidopsis proteome was run through the Transmembrane Hidden Markov Model 2.0 server and proteins containing one or, more rarely, two transmembrane domains within the N-terminal 150 amino acids were collected. Second, these sequences were submitted to the SUPERFAMILY prediction server, and sequences that were predicted to belong to the superfamilies NDP-sugartransferase, UDP-glycosyltransferase/glucogen-phosphorylase, carbohydrate-binding domain, Gal-binding domain, or Rossman fold were collected, yielding a total of 191 sequences. Fifty-two accessions already classified in CAZy were discarded. The resulting 139 sequences were then analyzed using the Three-Dimensional-Position-Specific Scoring Matrix and mGenTHREADER servers, and 27 sequences with similarity to either the GT-A or the GT-B fold were obtained. Proof of concept of the present approach has to some extent been provided by our recent demonstration that two members of this pool of 27 non-CAZy-classified putative GTs are xylosyltransferases involved in synthesis of pectin rhamnogalacturonan II

Regulation of Plasmodesmatal Permeability and Stomatal Patterning by the Glycosyltransferase-Like Protein KOBITO1

“…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).…”

Comprehensive Transcriptome Profiling in Tomato Reveals a Role for Glycosyltransferase in Mi-Mediated Nematode Resistance

“…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.…”

“…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.…”

Arabidopsis thaliana RGXT1 and RGXT2 Encode Golgi-Localized (1,3)-α-d-Xylosyltransferases Involved in the Synthesis of Pectic Rhamnogalacturonan-II

“…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).…”

“…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/).…”