1.3.22
DEVELOPMENTALLY CONTROLLED GENE EXPRESSION IN RUST FUNGI AND THE SPECIAL ROLE OF HAUSTORIA IN THE UPTAKE OF AMINO ACIDS

M HAHN, J SOHN and K MENDGEN

Universitšt Konstanz, Fakultšt Biologie, Lehrstuhl Phytopathologie, D-78457 Konstanz, Germany

Background and objectives
Haustoria are specialized hyphae of the obligately biotrophic rust fungi that are assumed to play a crucial role in parasitism. Because rust fungi cannot be cultivated in the absence of their host plants, and because the fungal mycelium cannot be isolated from infected plant tissue in physiologically intact form, studies on the function of rust haustoria are very limited. The rust mycelium consists of intercellular hyphae and haustoria, yet their relative contribution to nutrient uptake is unknown. To study the biotrophic metabolism on a molecular level, we have constructed a haustorium-specific cDNA library of the rust fungus Uromyces fabae and searched this library for genes that are phase-specifically expressed. We found that almost 20% of the genes (PIGs, in planta induced rust genes) in this library are strongly expressed in haustoria and rust-infected leaves, but not or only weakly in germinated spores and infection structures formed in vitro. Thus the transition from the early stages of infection (when the fungus is growing mainly at the expense of nutrients from the spore) to the development of the parasitic mycelium including haustoria is accompanied by a dramatic shift in gene expression [1].

Results and conclusions
From 32 PIGs isolated from the cDNA library, two (PIG2 and PIG27) were identified to encode putative amino-acid permeases. Their genes have a similar organization, both containing 17 introns, 11 of them at homologous positions. The deduced amino acid sequences of pPIG2 and pPIG27 are similar to each other (57% identity) and to amino-acid permeases from yeasts (30-40%). The developmental pattern of expression is different. With immunofluorescence microscopy, the pPIG2 was exclusively localized to haustorial plasma membranes [2]. In contrast, pPIG27 is also expressed in intercellular hyphae.

By heterologous complementation of a yeast histidine uptake-deficient mutant, we were able to functionally confirm (p)PIG27 as an amino-acid permease. 14C-his uptake studies revealed that (p)PIG27 mediates high-affinity transport. Competition of his transport with other amino acids indicated that (p)PIG27 also transports lys, arg, trp and phe. The (p)PIG27-expressing yeast cells accumulated his from the medium more than 1000-fold within 10 min, in a strongly pH-dependent manner. These data strongly indicate an active, proton-driven, low-specificity amino-acid transport by (p)PIG27.

Together with our studies on the plasma membrane H+-ATPase (see abstract by C Struck et al.), these data represent the first molecular evidence for a prominent, but not exclusive, role of rust haustoria in the uptake of amino acids from host cells.

References
1. Hahn M, Mendgen K, 1997. Molecular Plant-Microbe Interactions 10, 427-437.
2. Hahn M, Neef U, Struck C et al., 1997. Molecular Plant-Microbe Interactions 10, 438-445.