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Plant and Cell Physiology Advance Access published online on August 12, 2005
Plant and Cell Physiology, doi:10.1093/pcp/pci187
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Plant and Cell Physiology 2005 © The Japanese Society of Plant Physiologists (JSPP); all rights reserved.
Received December 28, 2004
Accepted August 3, 2005

Regular Paper

Expression of Allene Oxide Cyclase and Accumulation of Jasmonates During Organogenic Nodule Formation from Hop (Humulus lupulus var. Nugget) Internodes

Ana M. Fortes 1*, Otto Miersch 2, Peter R. Lange 2, Rui Malhó 1, Pilar S. Testillano 3, Maria del Carmen Risueño 3, Claus Wasternack 2, and Maria S. Pais 1

1 Lab of Plant Molecular Biology and Biotechnology, Ed. ICAT, FCUL, Campo Grande, 1749-016 Lisboa, Portugal
2 Leibniz Institute of Plant Biochemistry, Dept. Natural Product Biotechnology, Weinberg 3, D-06120 Halle/ S., Germany
3 Development and Nuclear Organization Group, Centro de Investigaciones Biologicas, CSIC, Madrid, Spain

* To whom correspondence should be addressed.
Ana M. Fortes, E-mail: amfortes{at}fc.ul.pt


  Abstract

A crucial step in the biosynthesis of jasmonic acid (JA) is the formation of its stereoisomeric precursor, cis-(+)-12-oxophytodienoic acid (OPDA), which is catalyzed by allene oxide cyclase (AOC, EC 5.3.99.6). A cDNA of AOC was isolated from Humulus lupulus var. Nugget. The ORF of 765 bp encodes a 255 amino acid protein, which carries a putative chloroplast targeting sequence. The recombinant protein without its putative chloroplast target sequence showed significant AOC activity. Previously we demonstrated that wounding induces organogenic nodule formation in hop (Fortes and Pais 2000). Here we show that the AOC transcript level increased in response to wounding of internodes, peaking between 2 and 4 h upon wounding. In addition, Western blot analysis showed elevated levels of AOC peaking 24 h after internode inoculation. The AOC increase was accompanied by increased JA levels 24 h after wounding, whereas OPDA reached its highest level already after 12 h. AOC is mostly present in the vascular bundles of inoculated internodes. During prenodule and nodule formation, AOC levels were still high. JA and OPDA levels decreased down to 10 and 118 pmol/ g FW, respectively, during nodule formation, but increased during plantlet regeneration. Double immuno-localisation analysis of AOC and Rubisco in connection with lugol staining showed that AOC is present in amyloplasts of prenodular cells and in the chloroplasts of vacuolated nodular cells, whereas meristematic cells accumulated little AOC. These data suggest a role of AOC and jasmonates in organogenic nodule formation and plantlet regeneration from these nodules.

Keywords: allene oxide cyclase; Humulus lupulus; jasmonates; organogenic nodule.
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