1.9.7
CONSTITUTIVE DEFENCE MECHANISMS OF WHITE LUPIN (LUPINUS ALBUS L.)

P WOJTASZEK, M PISLEWSKA, M STOBIECKI and P BEDNAREK

Polish Academy of Sciences, Poznan, Poland

Background and objectives
The genus Lupinus is one of the oldest known legumes, and various aspects of its biology are considerably different from those commonly observed in Leguminosae. Lupin plants form characteristic symbiotic nodules, they are not colonized by AM fungi, and usually they contain considerable amounts of quinolizidine alkaloids found only in the tribe Genisteae. To date, however, very little is known on lupin responses to pathogen infection.

Two model systems: white lupin (Lupinus albus) plants and suspension-cultured cells, treated with either abiotic (CuCl2) or biotic elicitors (preparations from purified cell walls of yeast or Colletotrichum lindemuthianum), were used to examine the mechanisms of defence responses. Two classes of secondary metabolites, phenolic compounds and quinolizidine alkaloids, were analysed, paralleling the observations of changes in protein profiles of the exocellular matrix (glyco)proteins.

Results and conclusions
Quinolizidine alkaloids: 14 alkaloids and seven alkaloid esters were identified in a high-alkaloid cultivar of white lupin [1]. Both qualitative and quantitative changes of alkaloid profiles were observed in elicitor-treated lupin plants. The extent of changes depended on the mode of elicitation and plant tissue analysed.
Isoflavonoids: these are present in all white lupin tissues, and most of them are conjugated with sugars. However, the levels of free aglycones differ depending on the tissue analysed, with the highest amounts in roots and the lowest in cotyledons and stems. Following elicitation, the aglycones of genistein and 2-hydroxygenistein are released from their respective glycosides and used for further prenylations [2]. Prenylated isoflavones are fungicidal and form the putative lupin's equivalent of pterocarpan phytoalexins in other legumes.
Isoflavone glucoside glucosidase (IGG): the enzyme is constitutively expressed in white lupin and associated with cell walls. The distribution of IGG parallels that of free isoflavone aglycones with the highest activity found in roots. Its activity increases following elicitor treatment. IGG specifically cleaves beta-glucosides of both simple phenolic compounds (salicylic alcohol, phenol) and isoflavones (genistein, 2'-hydroxygenistein). No activity of the enzyme towards cell wall oligosaccharides could be detected.
Exocellular (glyco)proteins: defence-related proteins were found to be secreted constitutively by suspension-cultured cells in the absence of any elicitor. They were also present in the walls of elicitor-treated cells [3]. Three of them were identified as: exocellular chitinase, polygalacturonase-inhibiting protein, and germin-like oxalate oxidase. The latter was further characterized and its properties were found to be very similar to germins found in Gramineae. This homopentamer of about 140-160 kDa (monomer of 32 kDa) is SDS- and proteinase-resistant. It is also a thermostable protein. For the first time the activity of germin-like protein from dicotyledoneous species is also demonstrated.

White lupin plants seem to exploit mainly the constitutive line of defence, with very little activation of inducible defence mechanisms in response to elicitation. Most of the elements of defence reactions studied were also present in normally growing plants or suspension-cultured cells.

This work was funded by State Committee for Scientific Research grants 6 P04C 107 08 and 6 P04C 022 13 to P.W.

References
1. Stobiecki M, Wojtaszek P, Bednarek P, Gulewicz K, 1996. Acta Physiologiae Plantarum 18, 313-319.
2. Wojtaszek P, Stobiecki M, 1997. Plant Physiology and Biochemistry 35, 129-135.
3. Wojtaszek P, Stobiecki M, Bolwell GP, 1997. Journal of Experimental Botany 48, 2015-2021.