Abstract
P-HYDROXYPHENYLPYRUVATE DIOXYGENASE (HPPD) is the first committed enzyme involved in the biosynthesis of vitamin E, and is characterized by catalyzing the conversion of p-hydroxyphenyl pyruvate (HPP) to homogentisic acid (HGA). Here, an HPPD gene was cloned from Medicago sativa L. and designated MsHPPD, which was expressed at high levels in alfalfa leaves. PEG 6000 (polyethylene glycol), NaCl, abscisic acid and salicylic acid were shown to significantly induce MsHPPD expression, especially in the cotyledons and root tissues. Overexpression of MsHPPD was found to significantly increase the level of β-tocotrienol and the total vitamin E content in Arabidopsis seeds. Furthermore, these transgenic Arabidopsis seeds exhibited an accelerated germination time, compared with wild-type seeds under normal conditions, as well as under NaCl and ABA treatments. Meanwhile, the expression level of several genes associated with ABA biosynthesis (NCED3, NCED5 and NCED9) and the ABA signaling pathway (RAB18, ABI3 and ABI5) were significantly down-regulated in MsHPPD-overexpressing transgenic lines, as well as the total free ABA content. Taken together, these results demonstrate that MsHPPD functions not only in the vitamin E biosynthetic pathway, but also plays a critical role in seed germination via affecting ABA biosynthesis and signaling.
Highlights
Vitamin E is an essential nutrient for animals and humans, the physiological significance of this substance has been studied widely
The results showed that the expression of the abscisic acid (ABA) biosynthetic genes ABA1, NCED3, NCED5 and NCED9 was not altered in dry seeds of the MsHPPD-overexpressing lines; in imbibed seeds, NCED3, NCED5 and NCED9 expression was significantly reduced, compared to the expression in wild-type seeds
Full-length HYDROXYPHENYLPYRUVATE DIOXYGENASE (HPPD) cDNA from alfalfa was isolated, and subsequent amino acid sequence alignment showed that MsHPPD from alfalfa shared high similarity with LsHPPD from Lactuca sativa, MtHPPD from Medicago truncatula, and AtHPPD from Arabidopsis thaliana
Summary
Vitamin E is an essential nutrient for animals and humans, the physiological significance of this substance has been studied widely. Vitamin E is not a single compound, but rather the collective name for a group of eight lipid-soluble antioxidants consisting of a polar chromanol head group and a hydrophobic prenyl tail[7], which are derived from the methylerythritol-4-phosphate (MEP) and shikimate pathways Four of these compounds are termed tocopherols, and the other four are termed tocotrienols, and depending on the saturation level of the hydrophobic tail and the number and position of the methyl groups on the chromanol ring, members of the vitamin E group are classified into α-, β-, γ-and δ-forms[8]. Overexpression of AtHPPD increased the total vitamin E level by seven-fold in Synechocystis[12], and significantly increased vitamin E accumulation in transgenic potato tubers[13] These observations indicate that modifying HPPD gene expression is a valid strategy to utilize during attempts to modulate the total vitamin E content of plant tissues. The growth inhibitor abscisic acid (ABA) is widely recognized as an important phytohormone involved in plant stress response and seed germination[14]. As such, discovering and characterizing related genes will enrich our knowledge on the biosynthetic mechanisms of this essential nutrient in forage crops, including alfalfa
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