The Neurospora circadian clock-controlled gene, ccg-2, is allelic to eas and encodes a fungal hydrophobin required for formation of the conidial rodlet layer.

Abstract

The Neurospora crassa clock-controlled gene (ccg-2) is transcriptionally activated by the circadian clock in a time-of-day-specific manner. Transcript and sequence analyses of ccg-2 reveal that the predicted Ccg-2 polypeptide bears significant similarity to a class of low-molecular-weight, cysteine-rich, hydrophobic proteins (hydrophobins), first identified in Schizophyllum, and including the product of the developmentally regulated Aspergillus gene, rodletless, required for spore surface rodlets. Allelism between ccg-2 and easily wettable (eas) (one of the first developmental genetic loci identified in Neurospora) was predicted on the basis of this similarity, their close genetic linkage, and previous findings demonstrating that eas mutants lack rodlets. In this study allelism is confirmed experimentally by showing that (1) transformation of an eas mutant strain with ccg-2 DNA results in phenotypic complementation, including restoration of surface rodlets, (2) inactivation of the ccg-2 gene, by RIP, results in an eas phenotype including loss of rodlet fascicles, and (3) the original eas strain has dramatically reduced levels of ccg-2 mRNA. Thus, the clock-controlled ccg-2 gene encodes an integral component of fungal asexual spores important for spore dispersal. The dramatic reduction of ccg-2 expression in the eas mutant has no apparent effect on the normal operation of the circadian clock, confirming that there is no feedback of this clock output on the oscillator itself. These data, in conjunction with the previous observation that ccg-2 is light induced, serve to focus attention on the dual interacting role of light and the circadian clock in the regulation of fungal spore development.[Key Words: Clock-controlled gene; ccg-2; easily wettable., hydrophobins; rodlets; circadian clock] Received August 13, 1992; revised version accepted September 15, 1992.Circadian rhythms are biological rhythms that are present in most, if not all, eukaryotes (Dunlap 1990). Daily fluctuations in cellular and organismal activities are controlled by an endogenous oscillator, commonly referred to as the biological clock. The clock exerts its effects on a wide variety of processes ranging from asexual spore formation in the ascomycete Neurospora crassa and the control of cell division in the marine protist Gonyaulax to photosynthesis in plants and cognitive functions in people (for review, see Edmunds 1988). However, at all phylogenetic levels, clocks appear to be assembled at the level of intracellular regulation and not intercellular communication. Thus, simple eukaryotes provide appropriate and powerful systems for molecular genetic analysis of this phenomenon.Of the eukaryotes known to possess a well-defined circadian clock, the filamentous fungus N. crassa provides an ideal model system for clock analysis {for review, see Feldman and Dunlap 1983;Dunlap 1990; Lakin-Thomas tCorresponding authors. et al. 1990). Advantages include an easily measurable circadian rhythm in developmental potential (rhythmic formation of aerial hy...

Regulation by Blue Light of the fluffy Gene Encoding a Major Regulator of Conidiation in Neurospora crassa

“…However, fl is transiently induced 30 min after the induction of conidiation, suggesting an additional role for FL in the formation of aerial hyphae (Correa and BellPedersen 2002). The relevance of FL as a major regulator of conidiation in Neurospora is supported by the observation of conidial development when fl expression is forced in vegetative hyphae (Bailey-Shrode and Ebbole 2004), a condition that leads to the expression of eas (Bailey-Shrode and Ebbole 2004), the gene for the hydrophobin rodlet protein that is located on the surface of matured conidia (Bell-Pedersen et al 1992;Lauter et al 1992). This is consistent with the observed binding of FL to the eas promoter (Rerngsamran et al 2005).…”

Mutants of Neurospora crassa that alter gene expression and conidia development.

“…con-6 and con-10 transcripts and proteins are normally undetectable in vegetative mycelium but appear at high levels during conidiation and in mature conidia; these transcripts and proteins disappear a few hours after conidia germinate (6,13). The expression pattern of these genes closely parallels that of eas (11,12). The con-6 and con -10 proteins are present in a second type of asexual spore, the microconidium (10,13), and the con-10 protein is present in sexual spores (10,13).…”

“…Four of these genes, con-6, con-8, con-10, and con-13, have been sequenced, and their expression has been examined (6)(7)(8)(9)(10). A fifth conidiationspecific gene, eas, also has been characterized (11,12); eas encodes the rodlet protein that coats the surface ofthe mature conidium.…”

Characterization of Adenylate Cyclase Interacting Protein ACI1 in the Rice Blast Fungus, Magnaporthe oryzae

“…Disruption of the well characterized hydrophobin gene MPG1 in M. oryzae results in absence of the rodlet layer and the production of a watersoaked, easily wettable phenotype [27]. The phenotype is commonly associated with several other characterized fungal hydrophobins such as rodA and EAS genes of Aspergillus nidulans and N. crassa, respectively [18][19][20]. ACI1 contains a predicted signal peptide and a predicted extracellular CFEM domain, a conserved motif consisting of eight conserved cysteines similar to that found in hydrophobins although the spacing of cysteines was found to be different.…”

“…An easily wettable phonotype has been reported for several mutants defective in fungal hydrophobins [18][19][20]. Based on the predicted signal peptides and extracellullar domains, we were interested to know whether the CFEM domain was related to hydrophobin function.…”