IDENTIFICATION OF DEVELOPMENTAL REGULATORY GENES IN ASPERGILLUS-NIDULANS BY OVEREXPRESSION

Overexpression of several Aspergillus nidulans developmental regulator y genes has been shown to cause growth inhibition and development at i nappropriate times. We set out to identify previously unknown developm ental regulators by constructing a nutritionally inducible A. nidulans expression library containing small, random genomic DNA fragments ins erted next to the alcA promoter [alcA(p)] in an A. nidulans transforma tion vector. Among 20,000 transformants containing random alcA(p) geno mic DNA fusion constructs, we identified 66 distinct mutant strains in which alcA(p) induction resulted in growth inhibition as well as caus ing other detectable phenotypic changes. These growth inhibited mutant s were divided into 52 FIG ( Forced expression Inhibition of Growth) a nd 14 FAB (Forced expression Activation of brlA) mutants based on whet her or not alcA(p) induction resulted in accumulation of mRNA for the developmental regulatory gene brlA. In four FAB mutants, alcA(p) induc tion not only activated brlA expression but also caused hyphae to diff erentiate into reduced conidiophores that produced viable spores from the tips as is observed after alcA(p)::brlA induction. Sequence analys es of the DNA fragments under alcA(p) control in three of these four s porulating strains showed that in two cases developmental activation r esulted from overexpression of previously uncharacterized genes, where as in the third strain, the alcA(p) was fused to brlA. The potential u ses for this strategy in identifying genes whose overexpression result s in specific phenotypic changes like developmental induction are disc ussed.