Brassinolide-improved development of Brassica napus microspore-derived embryos is associated with increased activities of purine and pyrimidine salvage pathways
Brassinolide-improved development of Brassica napus microspore-derived embryos is associated with increased activities of purine and pyrimidine salvage pathways
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DOI：
10.1007/s00425-010-1287-6
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发表时间：
2011-01-01
期刊：
影响因子：
4.3
通讯作者：
Stasolla, Claudio
中科院分区：
文献类型：
--
作者：
Belmonte, Mark;Elhiti, Mohamed;Stasolla, Claudio
细胞油菜素内酯 (BL) 水平调节甘蓝型油菜小孢子衍生胚 (MDE) 的发育。核苷酸的合成和降解是在用 BL 或 BL 生物合成抑制剂芸苔唑 (BrZ) 处理的开发 MDE 上进行测量的。通过跟踪 C-14 标记的腺嘌呤和腺苷（挽救途径的底物）以及肌苷（挽救途径和降解途径的中间体）的代谢命运来研究嘌呤代谢。对于嘧啶、乳清酸、尿苷和尿嘧啶用作从头（乳清酸）、补救（尿苷和尿嘧啶）和降解（尿嘧啶）途径的标记物。我们的结果表明，在 BL 处理的胚胎中，第 15 天时，腺嘌呤、腺苷和尿苷对核苷酸和核酸的利用显着增加，并且在整个培养期间保持较高水平。这些代谢变化归因于相应挽救酶的活性：腺嘌呤磷酸核糖转移酶 (EC 2.4.2.7)、腺苷激酶 (EC 2.7.1.20) 和尿苷激酶 (EC 2.7.1.48)，这些酶是由 BL 应用诱导的。 BL 对补救合成的促进伴随着降解途径活性的减少，表明存在利用标记前体的竞争性合成代谢和分解代谢机制。在 BrZ 处理的胚胎中，BL 水平耗尽，嘌呤和嘧啶核苷酸的挽救活性均降低，这与结构异常和不良胚胎性能有关。在这些胚胎中，主要挽救酶的活性始终低于对照（未处理）对应物中测量的活性。
Cellular brassinolide (BL) levels regulate the development of Brassica napus microspore-derived embryos (MDEs). Synthesis and degradation of nucleotides were measured on developing MDEs treated with BL or brassinazole (BrZ), a biosynthetic inhibitor of BL. Purine metabolism was investigated by following the metabolic fate of C-14-labelled adenine and adenosine, substrates of the salvage pathway, and inosine, an intermediate of both salvage and degradation pathways. For pyrimidine, orotic acid, uridine and uracil were employed as markers for the de novo (orotic acid), salvage (uridine and uracil), and degradation (uracil) pathways. Our results indicate that utilization of adenine, adenosine, and uridine for nucleotides and nucleic acids increased significantly in BL-treated embryos at day 15 and remained high throughout the culture period. These metabolic changes were ascribed to the activities of the respective salvage enzymes: adenine phosphoribosyltransferase (EC 2.4.2.7), adenosine kinase (EC 2.7.1.20), and uridine kinase (EC 2.7.1.48), which were induced by BL applications. The BL promotion of salvage synthesis was accompanied by a reduction in the activities of the degradation pathways, suggesting the presence of competitive anabolic and catabolic mechanisms utilizing the labelled precursors. In BrZ-treated embryos, with depleted BL levels, the salvage activity of both purine and pyrimidine nucleotides was reduced and this was associated to structural abnormalities and poor embryonic performance. In these embryos, the activities of major salvage enzymes were consistently lower to those measured in their control (untreated) counterparts.