The Aspergillus nidulans rcoA gene is required for veA-dependent sexual development

The Aspergillus nidulans rcoADelta mutant exhibits growth and developmental defects. We show that the rcoADelta mutant lacks cleistothecia and is self-sterile. In crosses with wild-type strains, rcoADelta nuclei do not contribute to the cleistothecial walls. Furthermore, sexual development resulting from veA overexpression is rcoA dependent, indicating that rcoA lies downstream of veA in the sexual development pathway.     

Neurospora crassa ve-1 affects asexual conidiation

The velvet factor of the homothallic fungus Aspergillus nidulans promotes sexual fruiting body formation. The encoding veA gene is conserved among fungi, including the ascomycete Neurospora crassa. There, the orthologous ve-1 gene encodes a deduced protein with high similarity to A. nidulans VeA. Cross-complementation experiments suggest that both the promoter and the coding sequence of N. crassa ve-1 are functional to complement the phenotype of an A. nidulans deletion mutant. Moreover, ve-1 expression in the heterologous host A. nidulans results in development of reproductive structures in a light-dependent manner, promoting sexual development in the darkness while stimulating asexual sporulation under illumination. Deletion of the N. crassa ve-1 locus by homologous gene replacement causes formation of shortened aerial hyphae accompanied by a significant increase in asexual conidiation, which is not light-dependent. Our data suggest that the conserved velvet proteins of A. nidulans and N. crassa exhibit both similar and different functions to influence development of these two ascomycetes.     

veA is required for toxin and sclerotial production in Aspergillus parasiticus

It was long been noted that secondary metabolism is associated with fungal development. In Aspergillus nidulans, conidiation and mycotoxin production are linked by a G protein signaling pathway. Also in A. nidulans, cleistothecial development and mycotoxin production are controlled by a gene called veA. Here we report the characterization of a veA ortholog in the aflatoxin-producing fungus A. parasiticus. Cleistothecia are not produced by Aspergillus parasiticus; instead, this fungus produces spherical structures called sclerotia that allow for survival under adverse conditions. Deletion of veA from A. parasiticus resulted in the blockage of sclerotial formation as well as a blockage in the production of aflatoxin intermediates. Our results indicate that A. parasiticus veA is required for the expression of aflR and aflJ, which regulate the activation of the aflatoxin gene cluster. In addition to these findings, we observed that deletion of veA reduced conidiation both on the culture medium and on peanut seed. The fact that veA is necessary for conidiation, production of resistant structures, and aflatoxin biosynthesis makes veA a good candidate gene to control aflatoxin biosynthesis or fungal development and in this way to greatly decrease its devastating impact on health and the economy.     

The VeA regulatory system and its role in morphological and chemical development in fungi

In fungi, the velvet gene, or veA, is involved in the regulation of diverse cellular processes, including control of asexual and sexual development as well as secondary metabolism. This global regulator is conserved in numerous fungal species. Interestingly, in Aspergilli, where most of the studies on veA have been carried out, this gene has been described to mediate development in response to light. In recent years the knowledge of this important regulatory system has expanded through the use of Aspergillus nidulans as a model organism, and through the study of veA orthologs across fungal genera. This review includes information on the current understanding of veA function and its mechanism of action. The fact that veA has only been found in fungi, together with advances in the elucidation of the veA mechanism, might be useful in designing future control strategies to decrease the detrimental effects of fungi while enhancing those qualities that are beneficial.     

Synthesis of specific proteins for sexual development in Dictyostelium discoideum

The development of Dictyostelium discoideum may proceed by two pathways, macrocyst or fruiting-body formation, the former being the sexual and the latter the asexual cycle. The pathway of development depends on the presence or absence of zygote giant cells which are produced through fusion of opposite mating-type cells in a population, in heterothallic strains. During the early stages of macrocyst development the patterns of developmentally regulated proteins were noted to differ considerably from those during fruiting-body development. Furthermore, the haploid cells around zygote giant cells synthesized a large number of specific proteins for macrocyst development through the influence of giant cells.     

The rpl16a gene for ribosomal protein L16A identified from expressed sequence tags is differentially expressed during sexual development of Aspergillus nidulans

We obtained 305 expressed sequence tags (ESTs), which are from the poly(A) site to the most proximal MboI site, from mycelia at the early sexual developmental (ESD) stage of Aspergillus nidulans. By comparison of these ESTs with those obtained previously from the vegetative stage and from the late sexual developmental stage followed by Northern blot analyses, genes of 17 ESTs were identified as being expressed more abundantly at the ESD stage than at the vegetative stage. Five of 17 genes were expressed more abundantly in the presence of the veA gene or the nsdD gene, suggesting that these 5 genes may be involved in sexual development. In a gene of one EST, appearing three times among 305 ESTs and identified by GenBank, polyadenylation seemed to occur at two sites. Nucleotide sequences of the gene having the EST and its cDNA revealed that the gene can code for a 202-amino-acid polypeptide with an estimated molecular mass of 23 kDa. The deduced amino acid showed 73% identity to Saccharomyces cerevisiae ribosomal protein L16A (RPL16A), and therefore the gene was named rpl16a. A. nidulans RPL16A had a putative leucine zipper motif and a basic leucine zipper motif like those of other organisms. The expression level of the rpl16a gene, present as a single copy in this organism, reached a maximum after 2 h, decreased thereafter, and increased again 30 to 50 h after the end of induction of sexual development. These results clearly indicated that the rpl16a gene is expressed differentially during sexual development.     

Genetic connection between fatty acid metabolism and sporulation in Aspergillus nidulans

In the Ascomycete fungus Aspergillus nidulans, the ratio of conidia (asexual spores) to ascospores (sexual spores) is affected by linoleic acid moieties including endogenous sporogenic factors called psi factors. Deletion of odeA (Delta odeA), encoding a Delta-12 desaturase that converts oleic acid to linoleic acid, resulted in a strain depleted of polyunsaturated fatty acids (18:2 and 18:3) but increased in oleic acid (18:1) and total percent fatty acid content. Linoleic acid-derived psi factors were absent in this strain but oleic acid-derived psi factors were increased relative to wild type. The Delta odeA strain was reduced in conidial production and mycelial growth; these effects were most noticeable when cultures were grown at 26 degrees C in the dark. Under these environmental conditions, the Delta odeA strain was delayed in ascospore production but produced more ascospores than wild type over time. This suggests a role for oleic acid-derived psi factors in affecting the asexual to sexual spore ratio in A. nidulans. Fatty acid composition and spore development were also affected by veA, a gene previously shown to control light driven conidial and ascospore development. Taken together our results indicate an interaction between veA and odeA alleles for fatty acid metabolism and spore development in A. nidulans.     

A putative G protein-coupled receptor negatively controls sexual development in Aspergillus nidulans

G protein-coupled receptors (GPCRs) are key components of heterotrimeric G protein-mediated signalling pathways that detect environmental signals and confer rapid cellular responses. To broaden our understanding of signalling mechanisms in the filamentous fungus Aspergillus nidulans, intensive analyses of the Aspergillus nidulans genome have been carried out and nine genes (gprA approximately gprI) that are predicted to encode seven transmembrane spanning GPCRs have been identified. Six of nine putative GPCRs have been disrupted and the gprD gene was found to play a central role in coordinating hyphal growth and sexual development. Deletion of gprD (Delta gprD) causes extremely restricted hyphal growth, delayed conidial germination and uncontrolled activation of sexual development resulting in a small colony covered by sexual fruiting bodies. Genetic studies indicate that GprD may not signal through the FadA (G alpha)-protein kinase A (PKA) pathway. Elimination of sexual development rescues both growth and developmental abnormalities caused by Delta gprD, suggesting that the primary role of GprD is to negatively regulate sexual development. This is supported by the fact that environmental conditions inhibiting sexual development suppress growth defects of the Delta gprD mutant. We propose that the GprD-mediated signalling cascade negatively regulates sexual development, which is required for proper proliferation of A. nidulans.     

VEA1 is required for cleistothecial formation and virulence in Histoplasma capsulatum

Histoplasma capsulatum is a pathogenic fungus dependent on dimorphism for virulence. Among the four described Velvet family genes, two of them, Ryp2 and Ryp3, have been shown to be required for dimorphism. It is known that Velvet A (VeA) is necessary for sexual development and toxin production in Aspergillus nidulans. However, the role of the VeA ortholog in H. capsulatum has not yet been explored. Vea1, H. capsulatum homolog of VeA, was studied to determine its role in cleistothecial formation, dimorphism, and virulence. H. capsulatum Vea1 restores cleistothecial formation and partially restores sterigmatocystin production in an A. nidulans veA deletion strain. Furthermore, silencing VEA1 in an H. capsulatum strain capable of forming cleistothecia abolishes cleistothecial formation. Silenced strains also switch to mycelial phase faster, and show impaired switching to the yeast phase once in mycelial phase. Virulence in mice and macrophages is attenuated in VEA1 silenced strains and silenced strains demonstrate increased sensitivity during growth under acidic conditions. These results indicate that H. capsulatum Vea1 shares a similar role in development as VeA. H. capsulatum is also more susceptible to growth in acidic conditions when VEA1 is silenced, which may contribute to the silenced strains' attenuated virulence in mice and macrophages.     

Aspergillus nidulans VeA subcellular localization is dependent on the importin alpha carrier and on light

The veA gene is a light-dependent regulator governing development and secondary metabolism in Aspergillus nidulans. We have identified a putative bipartite nuclear localization signal (NLS) motif in the A. nidulans VeA amino acid sequence and demonstrated its functionality when expressed in yeast. Furthermore, migration of VeA to the nucleus was dependent on the importin alpha. This bipartite NLS is also functional when VeA is expressed in A. nidulans. Interestingly, we found that VeA migration to the nucleus is light-dependent. While in the dark VeA is located mainly in the nuclei, under light VeA is found abundantly in the cytoplasm. The VeA1 mutant protein (lacking the first 36 amino acids at the N-terminus) was found predominantly in the cytoplasm independent of illumination. This indicates that the truncated bipartite NLS in VeA1 is not functional and fails to respond to light. These results might explain the lack of the morphological light-dependent response in strains carrying the veA1 allele. We also evaluated the effect of light on production of the mycotoxin sterigmatocystin in a veA wild-type and the veA1 mutant strains and found that the highest amount of toxin was produced by the veA+ strain growing in the dark, condition favouring accumulation of VeA in the nucleus.     

The Aspergillus nidulans phytochrome FphA represses sexual development in red light

Phytochrome photoreceptors sense red and far-red light through photointerconversion between two stable conformations, a process mediated by a linear tetrapyrrole chromophore. Originally, phytochromes were thought to be confined to photosynthetic organisms including cyanobacteria, but they have been recently discovered in heterotrophic bacteria and fungi, where little is known about their functions. It was shown previously in the ascomycetous fungus Aspergillus nidulans that asexual sporulation is stimulated and sexual development repressed by red light. The effect was reminiscent of a phytochrome response, and indeed phytochrome-like proteins were detected in several fungal genomes. All fungal homologs are more similar to bacterial than plant phytochromes and have multifunctional domains where the phytochrome region and histidine kinase domain are combined in a single protein with a C-terminal response-regulator domain. Here, we show that the A. nidulans phytochrome FphA binds a biliverdin chromophore, acts as a red-light sensor, and represses sexual development under red-light conditions. FphA-GFP is cytoplasmic and excluded from the nuclei, suggesting that red-light photoperception occurs in the cytoplasm. This is the first phytochrome experimentally characterized outside the plant and bacterial kingdoms and the second type of fungal protein identified that functions in photoperception.