Differential expression pattern of an acidic 9/13-lipoxygenase in flower opening and senescence and in leaf response to phloem feeders in the tea plant

Shouan Liu,Baoyu Han

doi:10.1186/1471-2229-10-228

Abstract

BackgroundLipoxygenase (LOXs) is a large family of plant enzymes that catalyse the hydroperoxidation of free polyunsaturated fatty acids into diverse biologically active compounds, collectively named phyto-oxylipins. Although multiple isoforms of LOXs have been identified in a wide range of annual herbaceous plants, the genes encoding these enzymes in perennial woody plants have not received as much attention. In Camellia sinensis (L.) O. Kuntze, no LOX gene of any type has been isolated, and its possible role in tea plant development, senescence, and defence reaction remains unknown. The present study describes the isolation, characterization, and expression of the first tea plant LOX isoform, namely CsLOX1, and seeks to clarify the pattern of its expression in the plant's defence response as well as in flower opening and senescence.ResultsBased on amino acid sequence similarity to plant LOXs, a LOX was identified in tea plant and named CsLOX1, which encodes a polypeptide comprising 861 amino acids and has a molecular mass of 97.8 kDa. Heterologous expression in yeast analysis showed that CsLOX1 protein conferred a dual positional specificity since it released both C-9 and C-13 oxidized products in equal proportion and hence was named 9/13-CsLOX1. The purified recombinant CsLOX1 protein exhibited optimum catalytic activity at pH 3.6 and 25°C. Real-time quantitative PCR analysis showed that CsLOX1 transcripts were detected predominantly in flowers, up-regulated during petal senescence, and down-regulated during flower bud opening. In leaves, the gene was up-regulated following injury or when treated with methyl jasmonate (MeJA), but salicylic acid (SA) did not induce such response. The gene was also rapidly and highly induced following feeding by the tea green leafhopper Empoasca vitis, whereas feeding by the tea aphid Toxoptera aurantii resulted in a pattern of alternating induction and suppression.ConclusionsAnalysis of the isolation and expression of the LOX gene in tea plant indicates that the acidic CsLOX1 together with its primary and end products plays an important role in regulating cell death related to flower senescence and the JA-related defensive reaction of the plant to phloem-feeders.

Highlights

Lipoxygenase (LOXs) is a large family of plant enzymes that catalyse the hydroperoxidation of free polyunsaturated fatty acids into diverse biologically active compounds, collectively named phyto-oxylipins
Isolation and sequence analysis of CsLOX1 To clone the C. sinensis lipoxygenase gene, three PCR fragments were first amplified by RT-PCR reaction using degenerate primers that correspond to the conserved amino acid sequences from other plant LOXs
Based on the BLAST P network service, the predicted amino acid sequence of CsLOX1 showed a high degree of identity with other plant LOXs such as common olive (77.9%, Genbank: EU678670), almond (77.2%, Genbank: AJ404331), potato (77%, Genbank: U60202), and hazelnut (76.9%, Genbank: AJ417975), which belong to type 1, but the gene shared its identity to a small extent with LOX isoforms from the type 2 group

Summary

Introduction

Lipoxygenase (LOXs) is a large family of plant enzymes that catalyse the hydroperoxidation of free polyunsaturated fatty acids into diverse biologically active compounds, collectively named phyto-oxylipins. The type 1 subfamily lacks a plasmid transit peptide and consists of both 9- and 13-LOXs, and the type 2 subfamily possesses 13-LOXs, which have a chloroplast transit peptide [1] Both 9Sand 13S-hydroperoxides are rapidly converted into a series of biologically active molecules by the action of the LOX pathway enzymes, and are referred to as phyto-oxylipins [1,2]. In Petunia inflata, a highly up-regulated allene oxide synthase (AOS) gene involved in petal senescence was identified and assumed to have a role in programmed cell death [19] whereas in Alstroemeria peruviana and two orchid species, no increase in LOXspecific activity was observed over time [20,21]. It is likely that the function of LOXs during petal senescence varies from species to species [22] and needs further study

Methods

Results

Discussion

Conclusion