Untersuchungen zur Rolle von stress-induzierbaren Lipoxygenase-Aktivitäten in Weizen

  • Investigations on the Role of Stress-Induced Lipoxygenase Isoforms in Wheat Plants

Seiler, Angelika; Grambow, Hans-Jürgen (Thesis advisor)

Aachen : Publikationsserver der RWTH Aachen University (2007)
Dissertation / PhD Thesis

Aachen, Techn. Hochsch., Diss., 2007


Lipoxygenases (LOX) catalyze the hydroperoxidation of unsaturated fatty acids. Depending upon the site of the hydroperoxy insertion LOX-isoforms are classified into a 9-LOX family and a 13-LOX family, respectively. It has been documented in a large number of earlier investigations that LOX activity increases in plants in response to pathogen attack or elicitation. This study focuses on the LOX-92 (Mr 92,000) and also on the LOX-103 (Mr 103,000) isoforms which were identified in wheat leaves in an earlier study (Bohland et al., Plant Phys. 114, 679-685, 1997). The strongly induced LOX-92, which represents more than 80 % of the LOX activity in stressed wheat leaves and which most probably is a cytosolic LOX species, has been shown to be a member of the 9-LOX family. Attempts to obtain LOX-92 protein in a sufficiently pure state to be able to obtain peptide sequence information failed due to it's extreme lability. Thus, it was not possible to clone LOX-92 by targeted PCR amplification. Northern blot analysis using a general probe for LOX sequences showed that activation of LOX-92 apparently did not occur by de novo synthesis but rather by a posttranslational mechanism. In another attempt to clone LOX-92 using degenerate probes for consensus LOX sequences, a full length wheat LOX cDNA, LOX2 Ta1 was cloned. The results indicate that LOX2 Ta1 is probably identical to LOX-103. LOX2 Ta1 (LOX-103) was further characterized by Northern blot analysis, sequence analysis, and positioning in a phylogenetic tree: (1) LOX-103 is induced by methyljasmonate and wounding at the RNA level. (2) LOX-103 is a 13-LOX species with a putative chloroplast leader sequence. (3) LOX-103 shows a close relationship to two other gramineous LOX species, the LOX2 Hv1 (Vorös et al., Eur. J. Biochem. 251, 36-44, 1998) and the LOX2 Or2 (Zabbai et al., Physiol. Mol. Plant Pathol. 64, 37-43, 2004). These species seem to play a role in induced resistance. Other aspects of this study include the identity and characteristics of LOX in wheat cell suspension cultures and also the possible existence of non-identified cell factors affecting LOX stability. The results are discussed in terms of a hypothesis in relation to other relevant work which accentuates the possible role of both the 9-LOX and 13-LOX isoforms within the intercellular signalling network determining the plant defence response.