Purification, quantification and gene expression of urate oxidases in rust-infected bean leaves

Bean leaf urate oxidase (host enzyme) activity increases in both compatible and incompatible interactions once the first haustorium has differentiated (1 day post-inoculation) after Uromyces phaseoli infection. Progressive development of the parasite in susceptible leaves leads to further enhancement of urate oxidase activity and both host and fungal enzymes are synergistically involved to give a progressive impetus to purine degradation. The necrogenic process associated with hypersensitivity in the incompatible response blocks fungal growth but does not change the pattern of urate oxidase enhancement. A pronounced 32kDa-subunit accumulation during the late infection stage revealed by SDS–PAGE analysis, greatly exceeds the enzyme activity increase suggesting a relevant drop in urate oxidase specific activity. This de novo enzyme synthesis is connected with greater gene expression, but only in susceptible leaves and concomitantly with the presence of the highest amount of fungal enzyme. These data are consistent with the hypothesis that an up-regulation of plant urate oxidase gene expression occurs at the post-transcriptional level rather than an over expression of the urate oxidase gene. We conclude that a general pathogenic effect and host growth reduction takes place as a consequence of an alteration in the catalytic property of host urate oxidase, and purine pool depauperation. The possibility that excess H2O2, generated by enhanced activity of urate oxidase, may contribute to hypersensitive cell death induction is also discussed.