ISOLATION AND CHARACTERIZATION OF MUTANTS DEFECTIVE IN PRODUCTION OF LACCASE IN NEUROSPORA-CRASSA

A protein synthesis inhibitor, cycloheximide, induces excretion of lac case in Neurospora crassa. The lah-1 mutation results in excretion of a large amount of laccase even in the absence of cycloheximide. Ten mu tations were induced that suppress derepressed excretion of laccase in the lah-1 mutant. Of these, seven second-site mutations were found to confer a laccase-noninducible phenotype, and were classified into two different complementation groups. Four mutations define a locus desig nated lni-1, found to be closely linked to ylo-1 on linkage group VI. The other three mutations were mapped to second locus, designated lni- 2, that lies between nic-3 and thi-3 on linkage group VII. The lni-2 l ocus was shown to encode laccase by RFLP mapping of the DNA fragment e ncoding laccase and by transformation of the lni-2 mutant with plasmid pBL1 carrying the laccase gene (the locus encoding laccase is hereaft er described as lacc). All lacc mutants examined (whether mutagen-indu ced or inactivated by repeat-induced point mutation) appeared to exhib it no phenotypic deficiency during both asexual and sexual cycles, sug gesting that the laccase gene is dispensable in N. crassa. Northern an alysis of total cellular RNA from the four lni-1 mutants demonstrated that the lni-1 mutations abolish increased transcription of the laccas e gene under inducing conditions. Consequently, the lni-1 locus is inf erred to encode a trans-acting positive regulator required for transcr iptional activation of the laccase gene in response to cycloheximide. Possible functions of the lah-1 gene are also described.