A new elicitor of the hypersensitive response in tobacco: a fungal glycoprotein elicits cell death, expression of defence genes, production of salicylic acid, and induction of systemic acquired resistance.

A 32 kDa glycoprotein whose effects in tobacco and other Nicotianae mimic a typical hypersensitive response, was isolated from Phytophthora megasperma. Infiltration of a few nanograms of the protein into leaves caused the formation of lesions that closely resemble hypersensitive response lesions. Transcripts of genes encoding enzymes of the phenylpropanoid and sesquiterpenoid pathways accumulated rapidly after elicitor application followed by salicylic acid production. Cellular damage, restricted to the infiltrated zone, occurred only several hours later, at a time when expression of PR protein genes was activated. After several days systemic acquired resistance was also induced. Thus, tobacco plant cells that perceived the glycoprotein generated a cascade of signals acting at local, short, and long distances, and causing the coordinate expression of specific defence responses in a way similar to hypersensitivity to tobacco mosaic virus. The glycoprotein represents a powerful tool to investigate further the signals and their transduction pathways involved in induced disease resistance. It may also be useful to engineer broad disease protection in a Nicotianae and possibly into crop plant species.

Pathogenesis-related PR-1 proteins are antifungal. Isolation and characterization of three 14-kilodalton proteins of tomato and of a basic PR-1 of tobacco with inhibitory activity against Phytophthora infestans.

Three distinct basic 14-kD proteins, P14a, P14b, and P14c, were isolated from tomato (Lycopersicon esculentum Mill. cv Baby) leaves infected with Phytophthora infestans. They exhibited antifungal activity against P. infestans both in vitro (inhibition of zoospore germination) and in vivo with a tomato leaf disc assay (decrease in infected leaf surface). Serological cross-reactions and amino acid sequence comparisons showed that the three proteins are members of the PR-1 group of pathogenesis-related (PR) proteins. P14a and P14b showed high similarity to a previously characterized P14, whereas P14c was found to be very similar to a putative basic-type PR-1 from tobacco predicted from isolated DNA clones. This protein, named PR-1 g, was purified from virus-infected tobacco (Nicotiana tabacum Samsun NN) leaves and characterized by amino acid microsequencing, along with the well-known acidic tobacco PR-1a, PR-1b, and PR-1c. The various tomato and tobacco PR-1 proteins were compared for their biological activity and found to display differential fungicidal activity against P. infestans in both the in vitro and in vivo assays, the most efficient being the newly characterized tomato P14c and tobacco PR-1g.

ASSIMILATION OF 15 NH4 + BY BEECH (FAGUS SYLVATICA L.) ECTOMYCORRHIZAS.

Ammonia assimilation has been followed in ectomycorrhizal roots of Fagus sylvatica. The absorption of ammonium ions was associated with a rapid synthesis of free amino acids in mycorrhizal tissues, glutamine being the most prominent. In the presence of [15 N]ammonium, glutamate, glutamine and alanine became the most strongly labelled metabolites of ectomycor-rhizas. Nitrogen-15 nuclear magnetic resonance spectroscopy demonstrated that the glutamine amide-N was the most highly enriched component of the extracts. Methionine sulphoximine and albizine, inhibitors of glutamine synthetase and glutamate synthase, almost completely blocked the incorporation of 15 N label into amino acids and induced an accumulation of NH4 + . These observations suggest that in the ammonia-fed beech ectomycorrhizas, ammonia assimilation occurs mainly via the glutamine synthetase/glutamate synthase pathway, and that glutamate dehydrogenase plays little, if any, part in this process. Alternative models for the nitrogen assimilation pathways in fungal and host tissues are presented.

Nitrogen Assimilation in Mycorrhizas : Ammonium Assimilation in the N-Starved Ectomycorrhizal Fungus Cenococcum graniforme.

Ammonium assimilation was followed in N-starved mycelia from the ectomycorrhizal Ascomycete Cenococcum graniforme. The evaluation of free amino acid pool levels after the addition of 5 millimolar NH(4) (+) indicated that the absorbed ammonium was assimilated rapidly. Post-feeding nitrogen content of amino acids was very different from the initial values. After 8 hours of NH(4) (+) feeding, glutamine accounted for the largest percentage of free amino acid nitrogen (43%). The addition of 5 millimolar methionine sulfoximine (MSX) to NH(4) (+)-fed mycelia caused an inhibition of glutamine accumulation with a corresponding increase in glutamate and alanine levels.Using (15)N as a tracer, it was found that the greatest initial labeling was into glutamine and glutamate followed by aspartate, alanine, and ornithine. On inhibiting glutamine synthetase using MSX, (15)N enrichment of glutamate, alanine, aspartate, and ornithine continued although labeling of glutamine was quite low. Moreover, the incorporation of (15)N label in insoluble nitrogenous compounds was lower in the presence of MSX. From the composition of free amino acid pools, the (15)N labeling pattern and effects of MSX, NH(4) (+) assimilation in C. graniforme mycelia appears to proceed via glutamate dehydrogenase pathway. This study also demonstrates that glutamine synthesis is an important reaction of ammonia utilization.