Nitrate assimilation in Fungi, a tool for studying transcription factors and cellular defence mechanisms

START project Y 114 Nitrate modelsystem: transcripionsfactors & cellular defence Joseph STRAUSS 18.06.1999 Aspergillus nidulans has become a major model organism for the study of gene regulation in general and for the regulation of nitrate assimilation in particular. In A. nidulans, two positively acting regulatory genes - nirA, a GAL4-type Zn++-cluster protein mediating induction, and areA, a GATA factor mediating nitrogen metabolite repression, - are required for the expression of the structural genes niaD (encoding nitrate reductase), niaA (encoding nitrite reductase) and crnA (encoding a nitrate permease). In the current proposal , we focus on experiments which should unravel the mechanisms by which modification-of- function mutations in nirA alleviate the need for a functional GATA factor, which is normally involved in chromatin remodelling in the bidirectional promoter of the divergently transcribed niaA and niaD genes. In the second part we plan to clone a gene which is involved in transmitting the nitrate induction signal. Finally, in the third part we plan to exploit a particular feature of mutants in the nitrate utilisation pathway to clone genes involved in the formation of nitric oxide (NO). This short-living intermediate of L-arginine metabolism seems to play an essential role in triggering cellular defence reactions in various experimental systems. Besides the well established role of nitric oxide in the mammalian immune system, NO has recently been shown to function also as a signal molecule in plant disease resistance by potentiating the effect of the oxidative burst necessary for the hypersensitive disease-resistance response. Influence of the proposed work on the development of the field In the context of the current knowledge on how transcription factors work it is certainly a question of general interest how a DNA binding protein of the Zn ++ -cluster family of transcription factors can alter its specificity such that it mediates chromatin remodelling. The fact, that it thereby replaces the function of a GATA factor is important in the view of how GATA factors determine their specificity They have to fulfil activation roles in metabolism, development and cell differentiation. Our experimental system with a number of defined modification-of-function mutants will help to define their role better. The results obtained from the cloning of a factor putatively involved in nitrate signalling might provide interesting new insights into the regulation of the nitrate assimilation pathway and could give new hints for the nitrate research in plant systems. The availability of mutants and genes from the nitric oxide synthesis pathway will help to better understand the role of the NO intermediate in cell defence reactions and will contribute to our knowledge in the field of molecular events in plant-microbe interactions.