A Novel Auxin Conjugate Hydrolase from Wheat with Substrate Specificity for Longer Side-Chain Auxin Amide Conjugates

Abstract

This study investigates how the ILR1-like indole acetic acid (IAA) amidohydrolase family of genes has functionally evolved in the monocotyledonous species wheat (Triticum aestivum). An ortholog for the Arabidopsis IAR3 auxin amidohydrolase gene has been isolated from wheat (TaIAR3). The TaIAR3 protein hydrolyzes negligible levels of IAA-Ala and no other IAA amino acid conjugates tested, unlike its ortholog IAR3. Instead, TaIAR3 has low specificity for the ester conjugates IAA-Glc and IAAmyoinositol and high specificity for the conjugates of indole-3-butyric acid (IBA-Ala and IBA-Gly) and indole-3-propionic-acid (IPA-Ala) so far tested. TaIAR3 did not convert the methyl esters of the IBA conjugates with Ala and Gly. IBA and IBA conjugates were detected in wheat seedlings by gas chromatography-mass spectrometry, where the conjugate of IBA with Ala may serve as a natural substrate for this enzyme. Endogenous IPA and IPA conjugates were not detected in the seedlings. Additionally, crude protein extracts of wheat seedlings possess auxin amidohydrolase activity. Temporal expression studies of TaIAR3 indicate that the transcript is initially expressed at day 1 after germination. Expression decreases through days 2, 5, 10, 15, and 20. Spatial expression studies found similar levels of expression throughout all wheat tissues examined.In vascular plants, auxins, primarily indole-3-acetic acid (IAA), regulate gene expression, cell division, and cell elongation and differentiation in plant tissue. Auxins also affect vascularization, phototropism, geotropism, fruit development, flower development, and apical dominance (Davies, 1995). While IAA in low concentrations stimulates growth and development, higher concentrations can be toxic to the plant (Bandurski et al., 1995). Therefore, tight control of IAA concentration is necessary for proper plant development.IAA is stored in conjugated forms that are mostly considered to be inactive. Two main types of conjugated molecules have been studied: the amide-linked IAA forms bound to one or more amino acids and the ester-linked forms primarily bound to a sugar(s). These two types of conjugates appear to be found at varying concentrations in the diverse tissues of angiosperms (Domagalski et al., 1987). On average, 95% of all IAA in a plant is conjugated into these storage forms (Cohen and Bandurski, 1982;Bandurski et al., 1995;Campanella et al., 1996;Walz et al., 2002).There have been a variety of amide conjugates found in the plants studied to date. IAA-Asp has been identified as a natural conjugate in Scots pine (Pinus sylvestris; Andersson and Sandberg, 1982) and, together with IAA-Glu, in cucumber (Sonner and Purves, 1985) and soybean (Cohen, 1982). IAA-Ala has been detected in Picea abies Karst (Ö stin et al., 1992). Additionally, IAA-Ala, IAA-Asp, IAA-Leu, and IAA-Glu have been detected in Arabidopsis L. Heynh (Tam et al., 2000;Kowalczyk and Sandberg, 2001), although recent data suggest that IAA peptides may account for the majority of amide conjugates in this and other plant ...

Comparison of Dynamic Changes of Endogenous Plant Hormones and Hormone-Related Gene Expression between Near Isogenic Lines with Normal Spike and Branched Spike Wheat

“…Temporal expression studies of TaIAR3 indicate that the transcript is initially expressed at day 1 after germination and then expression decreases through days 2, 5, 10, 15, and 20. Spatial expression studies found similar levels of expression throughout all wheat tissues examined (Campanella et al, 2004). Our study results showed that the transcription level of TaIAR3 in branched spike line increased much higher than those in normal spike lines in young shoot and young leaves after the two ridges stage.…”

Arabidopsis Seed Development and Germination Is Associated with Temporally Distinct Metabolic Switches

“…These changes were associated with high mRNA levels of genes encoding enzymes of amino acid biosynthesis, implying that amino acid biosynthesis apparently continues during the desiccation period, and that at least some amino acids significantly accumulate in their free forms rather then being incorporated into storage proteins. These lines of evidences taken in the context of current models of seed metabolism suggest that the seed desiccation process preferentially favors (1) the decrease of the C/N ratio, (2) the accumulation of free amino acids rather than storage proteins, and (3) the production of shikimate-derived secondary metabolites associated with defense (Hull et al, 2000;Glawischnig et al, 2004;Piotrowski et al, 2004) and indole-3-acetic acid biosynthesis (Campanella et al, 2004;Sharaf and Farrag, 2004) that are necessary triggers for seed germination (Normanly, 1997;Slavov et al, 2004). In addition to the accumulation of free amino acids and various carbon metabolites, the period of seed desiccation was also associated with small increases in the levels of various fatty acids, fatty acid-related compounds such as glycerol, as well as intermediates of fatty acid degradation (Fig.…”

Perturbation of Indole-3-Butyric Acid Homeostasis by the UDP-GlucosyltransferaseUGT74E2ModulatesArabidopsisArchitecture and Water Stress Tolerance

“…Besides IBA synthetase activities in maize and Arabidopsis, IBA glucosylation activity in Arabidopsis Ludwig-Mü ller, 2007) and IBA-Ala hydrolase in wheat (Triticum aestivum) (Campanella et al, 2004), the identity of the plant genes involved in IBA biosynthesis, conjugation, and/or hydrolysis remained unknown . Here, we demonstrate that UGT74E2 is an IBA glucosyltransferase with a high and preferred activity toward IBA in in vitro assays and that its overexpression resulted in increased IBA-Glc levels in planta (Figure 2, Table 1).…”