Brassinolide-improved development of <Emphasis Type="Italic">Brassica napus</Emphasis> microspore-derived embryos is associated with increased activities of purine and pyrimidine salvage pathways |
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Authors: | Mark Belmonte Mohamed Elhiti Hiroshi Ashihara Claudio Stasolla |
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Institution: | (1) Department of Biological Sciences, University of Manitoba, Winnipeg, MB, R3T 2N2, Canada;(2) Department of Biological Sciences, Graduate School of Humanities and Sciences, Ochanomizu University, Bunkyo-ku, Tokyo 112-8610, Japan;(3) Department of Plant Science, University of Manitoba, Winnipeg, MB, R3T 2N2, Canada; |
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Abstract: | 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 14C-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. |
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