Kiyoshi Onai

A new wc-1 mutant of Neurospora crassa shows unique light sensitivity in the circadian conidiation rhythm.

Abstract. A new clock mutant (rhy-2) was isolated by DNA insertion mutagenesis using a plasmid that contains a region located upstream of the cmd gene in the genome of Neurospora crassa. This mutant is arrhythmic with regard to conidiation in continuous darkness but rhythmic under a light-dark cycle. After plasmid rescue from genomic DNA of the rhy-2 strain, the insertion was localized to the gene white collar-1 (wc-1). Plasmid DNA was inserted 3′ to the sequence encoding the polyglutamine region of the WC-1 gene product, and an mRNA encoding a truncated WC-1 protein must be synthesized under the control of the cmd promoter. The new wc-1 mutant, rhy-2, is still sensitive to light, although only weakly. Since the circadian rhythm of conidiation in continuous darkness is eliminated in the mutant, the polyglutamine region in WC-1 may be essential for both clock function and light-induced carotenogenesis in Neurospora.

Mutation of the gene for the second-largest subunit of RNA polymerase I prolongs the period length of the circadian conidiation rhythm in Neurospora crassa

Abstract The period length of the circadian conidiation rhythm was examined in a mutant strain of Neurospora crassa, un-18, that is temperature sensitive for mycelial growth. The un-18 mutant showed a temperature-sensitive phenotype with respect to both mycelial growth and the period length of the conidiation rhythm. Below 22° C, the un-18 mutation did not affect the period length, but at temperatures between 22° C and 32° C, the period length of the un-18 mutant was ∼2 h longer than that of the wild-type strain. The un-18+ gene was cloned and was found to encode the second-largest subunit of RNA polymerase I, which is involved in the synthesis of rRNA. These results indicate that a defect in ribosome synthesis, which must result in a lower rate of protein synthesis, lengthens the period of the circadian conidiation rhythm in Neurospora.

Spermidine Determines the Sensitivity to the Calmodulin Antagonist, Chlorpromazine, for the Circadian Conidiation Rhythm but Not for the Mycelial Growth in Neurospora crassa

The gene that suppresses the phenotype of the cpz-2mutation, which results in changing the sensitivity to chlorpromazine in relation to mycelial growth and circadian rhythms, was cloned in Neurospora crassa.This gene is not the cpz-2 gene itself but rather is identical to the spe-3gene that encodes spermidine synthase in Neurospora. The intracellular content of spermidine was lowered in the cpz-2strain compared to that of the wild-type strain. By integration of the spe-3 gene or by the addition of spermidine into culture medium, the temperature sensitivity of mycelial growth was lost and the conidiation rhythm became sensitive to chlorpromazine in the cpz-2 strain, as was observed in the wild-type strain, but the hypersensitivity of mycelial growth on chlorpromazine in the cpz-2strain was not affected. Therefore, it appears that spermidine determines only the sensitivity of the conidiation rhythm to chlorpromazine.