Planta (2006) 224: 792–800 DOI 10.1007/s00425-006-0262-8 ORIGINAL ARTICLE André Frenzel · Nadine Tiller · Bettina Hause Franziska Krajinski The conserved arbuscular mycorrhiza-specific transcription of the secretory lectin MtLec 5 is mediated by a short upstream sequence containing specific protein binding sites Received: 10 January 2006 / Accepted: 6 March 2006 / Published online: 5 April 2006  Springer-Verlag 2006 Abstract In Medicago truncatula a family of mycor- rhiza-speciWc expressed lectins has been identiWed recently, but the function and regulation of these lectins during the arbuscular mycorrhiza symbiosis are still unknown. In order to characterize a Wrst member of this protein family, MtLec5 was analyzed concerning its local- ization and regulation. Confocal laser scanning micros- copy showed that MtLec5 is a secretory protein indicating a role as a vegetative storage protein, which is speciWcally expressed in mycorrhizal root systems. To study the molecular mechanisms leading to the mycor- rhiza-speciWc transcription, deletion studies of pMtLec5 were done using reporter gene fusions. Potential cis-act- ing elements could be narrowed down to a 150 bp frag- ment that was located approximately at ¡300/¡150 according to the transcription start, suggesting the bind- ing of positive regulators to this area. Similar expression pattern of the reporter gene was found after transforming roots of the non-legume Nicotiana tabacum with the het- erologous promoter–reporter fusions. This indicated that the observed mycorrhiza-speciWc transcriptional induc- tion is not legume-speciWc. Electrophoretic mobility shift assays showed that several factors which were exclusively present in mycorrhizal roots bind within the 150 bp pro- moter area. This strengthens the hypothesis of positive regulators mediating the AM-speciWc gene expression. Keywords Arbuscular mycorrhiza · Confocal laser scanning microscopy · Electrophoretic mobility shift assay · Lectin · Medicago · Promoter Abbreviations AM: Arbuscular mycorrhiza · CLSM: Confocal laser scanning microscopy · EMSA: Electro- phoretic mobility shift assay · GFP: Green Xuorescent protein · RACE: Rapid ampliWcation of cDNA ends Introduction In natural ecosystems more than 80% of all terrestrial plants are capable of developing a mutualistic interac- tion with the fungi of the phylum Glomeromycota (Smith and Read 1997; Schüssler et al. 2001). The arbuscular mycorrhiza (AM) symbiosis is several 100 million years old and fossil records show that AM fungi have been associated with the root systems of Wrst land plants (Remy et al. 1994). Symbiosis is initiated by the attach- ment of the fungus to the root surface forming appresso- ria. After colonization of the root cortex, the fungus forms highly dichotomous branched intracellular struc- tures referred to as arbuscules. The symbiosis is charac- terized by a bidirectional exchange of nutrients: carbon components are bestowed from the plant to the fungus whereas the latter mainly transfers phosphorous, and also increased uptake of other minerals such as zinc and copper to its host is reported (Ortas et al. 2002). Research has mostly been concentrated on the exchange of phosphorous, which resulted in the identiWcation of the corresponding mycorrhiza-speciWc transporters (Rausch et al. 2001; Harrison et al. 2002; Nagy et al. 2005). Furthermore, AM fungi can mediate higher levels of resistance against abiotic stress (Augé 2001) and pro- tection against plant pathogens (Newsham et al. 1995; Cordier et al. 1996; Slezack et al. 2000). As the AM interaction is quite ancient, it can be antici- pated that important steps are highly conserved in diVer- ent AM forming species: a conserved regulatory mechanism has been identiWed in two legumes (Endre et al. 2002; Stracke et al. 2002). Analyses of mutants, which are defective in Rhizobium-legume, and AM sym- bioses gave evidence that both interactions share com- mon signal transduction pathways (Duc et al. 1989; A. Frenzel · N. Tiller · F. Krajinski (&) Lehrgebiet Molekulargenetik, Universität Hannover, Herrenhäuser Str. 2, 30419 Hannover, Germany E-mail: krajinski@lgm.uni-hannover.de Tel.: +49-511-7625548 Fax: +49-511-7624088 B. Hause Department of Secondary Metabolism, Leibniz Institute of Plant Biochemistry, P.O. Box 110432, 06018 Halle, Germany