A BAHD acyltransferase is expressed in the tapetum of Arabidopsis anthers and is involved in the synthesis of hydroxycinnamoyl spermidines
A BAHD acyltransferase is expressed in the tapetum of Arabidopsis anthers and is involved in the synthesis of hydroxycinnamoyl spermidines
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DOI：
10.1111/j.1365-313x.2008.03773.x
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发表时间：
2009-04-01
期刊：
影响因子：
7.2
通讯作者：
Legrand, Michel
中科院分区：
文献类型：
--
作者：
Grienenberger, Etienne;Besseau, Sebastien;Legrand, Michel
BAHD酰基转移酶催化许多植物次生代谢物的酰化。我们的特征在于At 2g 19070，拟南芥BAHD基因家族的成员的功能。酰基转移酶基因在花发育早期的花药绒毡层细胞中特异表达。在RNAi植物和敲除（KO）突变株系中研究了基因阻遏的影响。用针对重组蛋白产生的特异性抗血清进行免疫印迹来评估At 2g 19070基因产物在各种拟南芥基因型的花中的积累，所述拟南芥基因型包括KO和RNAi系、雄性不育突变体ms 1和过表达酰基转移酶基因的转化体。从这些遗传背景的花蕾组织的代谢谱表现出正相关的积累酰基转移酶蛋白质和代谢产物的数量，通过串联质谱鉴定为N-1，N-5，N-10-三羟基阿魏酰亚精胺和N-1，N-5-二羟基阿魏酰-N-10-芥子酰亚精胺puestrine。这些产物沉积在花粉衣中，可以很容易地通过花粉洗液提取，并被证明是花粉自发荧光的原因。用各种羟基肉桂酰辅酶A酯和多胺分别作为供体和受体底物测定细菌中产生的重组酶的活性。阿魏酰辅酶A和亚精胺被证明是最好的底物，因此该酶被命名为亚精胺羟基肉桂酰转移酶（SHT）。在花发育过程中与SHT共调控的甲基转移酶基因（At 1g 67990）被证明参与亚精胺缀合物的O-甲基化，通过分析其在RNAi植物中的抑制后果和表征重组酶的甲基化活性。
BAHD acyltransferases catalyze the acylation of many plant secondary metabolites. We characterized the function of At2g19070, a member of the BAHD gene family of Arabidopsis thaliana. The acyltransferase gene was shown to be specifically expressed in anther tapetum cells in the early stages of flower development. The impact of gene repression was studied in RNAi plants and in a knockout (KO) mutant line. Immunoblotting with a specific antiserum raised against the recombinant protein was used to evaluate the accumulation of At2g19070 gene product in flowers of various Arabidopsis genotypes including the KO and RNAi lines, the male sterile mutant ms1 and transformants overexpressing the acyltransferase gene. Metabolic profiling of flower bud tissues from these genetic backgrounds demonstrated a positive correlation between the accumulation of acyltransferase protein and the quantities of metabolites that were putatively identified by tandem mass spectrometry as N-1,N-5,N-10-trihydroxyferuloyl spermidine and N-1,N-5-dihydroxyferuloyl-N-10-sinapoyl spermidine. These products, deposited in pollen coat, can be readily extracted by pollen wash and were shown to be responsible for pollen autofluorescence. The activity of the recombinant enzyme produced in bacteria was assayed with various hydroxycinnamoyl-CoA esters and polyamines as donor and acceptor substrates, respectively. Feruloyl-CoA and spermidine proved the best substrates, and the enzyme has therefore been named spermidine hydroxycinnamoyl transferase (SHT). A methyltransferase gene (At1g67990) which co-regulated with SHT during flower development, was shown to be involved in the O-methylation of spermidine conjugates by analyzing the consequences of its repression in RNAi plants and by characterizing the methylation activity of the recombinant enzyme.