Analysis of xyloglucan endotransglucosylase/hydrolase (XTH) gene families in kiwifruit and apple

Abstract

Xyloglucan endotransglucosylase/hydrolase (XTH) enzymes are thought to play a key role in fruit ripening by loosening the cell wall in preparation for further modification by other cell wall-associated enzymes and through disassembly of xyloglucan. Twenty-five XTH genes were isolated and characterised from kiwifruit and apple databases containing >270,000 expressed sequence tags (ESTs). The XTH genes (14 from kiwifruit, 11 from apple) encoded putative proteins with similar molecular weights, but with isoelectric points ranging from acidic to basic. All 25 XTH amino acid sequences contained the two conserved glutamic acid residues at the active site of the XTH enzyme. Phylogenetic analysis produced trees with branches each containing kiwifruit, apple, tomato and Arabidopsis XTH sequences, suggesting the potential for functional orthology. EST expression profiling and quantitative PCR analysis were used to identify kiwifruit and apple XTH transcripts expressed in root tip, flower, young leaf and ripe fruit. In ripe apple fruit, the transcripts for two XTH genes ( Md-XTH2 and Md-XTH10) were the most abundant, whilst in ripe kiwifruit three transcripts predominated ( Ad-XTH4, 5 and 7). Ad-XTH7 was highly expressed in the outer pericarp of firm kiwifruit at commercial harvest and expression of this mRNA decreased during the rapid softening phase. Three kiwifruit XTH genes ( Ad-XTH5, Ad-XTH7 and Ac-XTH14) were expressed in Escherichia coli and the recombinant proteins were shown to have xyloglucan endotransglucosylase activity. This research lays the groundwork for understanding the role of XTHs during fruit ripening and storage in kiwifruit and apple.