Comprehensive genomic analysis of the TYROSINE AMINOTRANSFERASE (TAT) genes in apple (Malus domestica) allows the identification of MdTAT2 conferring tolerance to drought and osmotic stresses in plants

Highlights

• Four MdTATs were identified in apple genome.

• MdTATs might play important roles in responses to environments.

• MdTATs showed distinct expression patterns in different apple tissues or under drought stress.

• MdTAT2 overexpression increased resistance to drought and osmotic stress.

Abstract

Tyrosine aminotransferase (TAT, EC 2.6.1.5) is the first key enzyme that catalyzes the reversible interconversion of tyrosine and 4-hydroxyphenylpyruvate in the tyrosine-derived pathway for syntheses of important secondary metabolites and compounds. Although plant TAT genes have been proposed to be important in response to abiotic stress, there is little information about TAT genes in woody perennial tree species, especially in economic fruit trees. Based on TAT domain searching, sequence homology screening and phylogenetic analysis, we identified four TATs in apple genome. Then, we carried out a detailed phylogenetic analysis of TAT genes from multi-species, focusing on apple (Malus domestica). The result showed that the TAT family comprises three major classes corresponding to genes from angiosperms, mammals, and bacteria. Angiosperm TAT genes could be further divided into six subclasses. Analysis of intron-exon structure revealed that the typical TAT gene contains six introns and seven exons, with exons of similar size at each exon location. Promoter analysis showed that the 5′-flanking region of apple MdTATs contain multiple cis-acting elements including those implicated in light, biotic stress, abiotic stress, and hormone response. MdTATs were expressed to various levels in all apple structures and organs evaluated, and showed distinct expression patterns under water deficit stress. Ectopic expression of MdTAT2 in Arabidopsis or over-expression of MdTAT2 in apple callus tissue conferred enhanced tolerance to drought and osmotic stress. Collectively, these results suggest a role for TAT genes in drought and osmotic stresses and provide valuable information for further research of TAT genes and their function in plants.

Introduction

Tyrosine aminotransferase (TAT, EC 2.6.1.5) is a key enzyme in the tyrosine-derived pathway in rosmarinic acid biosynthesis (Petersen and Simmonds, 2003). It catalyzes the reversible transamination of tyrosine and 2-oxoglutarate, yielding glutamate and 4-hydroxyphenylpyruvate (Riewe et al., 2012). In plants, 4-hydroxyphenylpyruvate is a precursor for biosynthesis of various secondary metabolites and compounds including plastoquinone, tocopherols, rosmarinic acid and benzylisoquinoline alkaloids (Huang et al., 2008; Lee and Facchini, 2011; Riewe et al., 2012; Wang et al., 2015).

Previous studies showed that TAT belongs to a multigene family (Riewe et al., 2012; Wang et al., 2015). In the Arabidopsis genome, there are eight putative TAT genes (Prabhu and Hudson, 2010; Riewe et al., 2012), four of which (AtTAT1, AtTAT2, CORI3, and SUR1) have been demonstrated to have TAT enzyme activity. TAT was shown to be the target of 5-benzyloxymethyl-1,2-isoxazolines derivatives and the herbicide cinmethylin (Grossmann et al., 2012; Riewe et al., 2012).

TAT in mammals and fungi is understood in considerable detail because it has been subjected to comprehensive biochemical and structural analysis (Blankenfeldt et al., 1999; Lee and Facchini, 2011; Mehere et al., 2010; Schneider et al., 2008; Sobrado et al., 2003). However, TAT in plants has not been well studied. In Arabidopsis, and in Chenopodium quinoa and Amaranthus caudatus cell cultures, TAT has been implicated as the first enzyme in tocopherol biosynthesis, and is induced by methyl jasmonate (MeJA), wounding, and coronatine (Antognoni et al., 2009; Holländer-Czytko et al., 2005; Lee and Facchini, 2011; Lopukhina et al., 2001). Additionally, TAT activity was found in MeJA treated Salvia miltiorrhiza hairy root cultures and in rosmarinic acid-producing Coleus blumei and Anchusa officinalis cell suspension cultures (De-Eknamkul and Ellis, 1987; Lee and Facchini, 2011; Xiao et al., 2009). Moreover, overexpression of a TAT gene from Perilla frutescens, designated PfTAT, resulted to increased rosmarinic acid yield (Lu et al., 2013). In plants, rosmarinic acid and tocopherols are capable of scavenging free radicals, and enhance protection against a variety of abiotic and biotic environmental stressors (Lee and Facchini, 2011). These findings suggested that TAT genes might participate in regulating tolerance of plants to biotic and abiotic stress.

Water scarcity, including drought conditions potentially exacerbated by climate change, is an increasing challenge for production of agricultural crops (Breshears et al., 2005). Apple (Malus domestica) is one of most economically important horticultural crops worldwide. In China, which is the primary producer of apples in the world by volume, apple production is centered on the Loess Plateau. Although the climate of this area is usually ideal for apple production, production is often negatively affected by severe drought (Yan et al., 2015). Thus, in China and elsewhere, a primary breeding goal for apple cultivar improvement is drought tolerance (Wang et al., 2018). Based on this, and the previous findings about the responses of plant TAT genes to various abiotic stresses (Antognoni et al., 2009; Holländer-Czytko et al., 2005; Lee and Facchini, 2011; Lopukhina et al., 2001), here, we carried out the first thorough analysis of the TAT genes in apple (MdTATs) and their potential function in tolerance to abiotic stresses. Our results lay the foundation for future multifunctional analysis of these genes and improvement of apple tolerance to abiotic stresses.