The PHOA and PHOB cyclin-dependent kinases perform an essential function in Aspergillus nidulans

Unlike Pho85 of Saccharomyces cerevisiae, the highly related PHOA cyclin-dependent kinase (CDK) of Aspergillus nidulans plays no role in regulation of enzymes involved in phosphorous acquisition but instead modulates differentiation in response to environmental conditions, including limited phosphorous. Like PHO85, Aspergillus phoA is a nonessential gene. However, we find that expression of dominant-negative PHOA inhibits growth, suggesting it may have an essential but redundant function. Supporting this we have identified another cyclin-dependent kinase, PHOB, which is 77% identical to PHOA. Deletion of phoB causes no phenotype, even under phosphorous-limited growth conditions. To investigate the function of phoA/phoB, double mutants were selected from a cross of strains containing null alleles and by generating a temperature-sensitive allele of phoA in a deltaphoB background. Double-deleted ascospores were able to germinate but had a limited capacity for nuclear division, suggesting a cell cycle defect. Longer germination revealed morphological defects. The temperature-sensitive phoA allele caused both nuclear division and polarity defects at restrictive temperature, which could be complemented by expression of mammalian CDK5. Therefore, an essential function exists in A. nidulans for the Pho85-like kinase pair PHOA and PHOB, which may involve cell cycle control and morphogenesis.     

acon-3, the Neurospora crassa ortholog of the developmental modifier, medA, complements the conidiation defect of the Aspergillus nidulans mutant

Aspergillus nidulans and Neurospora crassa are ascomycetes that produce asexual spores through morphologically distinct processes. MedA, a protein with unknown function, is required for normal asexual and sexual development in A. nidulans. We determined that the N. crassa ortholog of medA is acon-3, a gene required for early conidiophore development and female fertility. To test hypotheses about the evolutionary origins of asexual development in distinct fungal lineages it is important to understand the degree of conservation of developmental regulators. The amino acid sequences of A. nidulans MedA and N. crassa ACON-3 shared 37% identity and 51% similarity. acon-3 is induced at late time points of conidiation. In contrast, medA is constitutively expressed and MedA protein localizes to nuclei in all tissue types. Nonetheless, expression of acon-3 using its native promoter complemented the conidiation defects of the A. nidulans ΔmedA and medA15 mutants. We conclude that the biochemical activity of the medA orthologs is conserved for conidiation.     

Multiple roles of a heterotrimeric G-protein gamma-subunit in governing growth and development of Aspergillus nidulans

Vegetative growth signaling in the filamentous fungus Aspergillus nidulans is primarily mediated by the heterotrimeric G-protein composed of FadA (G alpha), SfaD (G beta), and a presumed G gamma. Analysis of the A. nidulans genome identified a single gene named gpgA encoding a putative G gamma-subunit. The predicted GpgA protein consists of 90 amino acids showing 72% similarity with yeast Ste18p. Deletion (delta) of gpgA resulted in restricted vegetative growth and lowered asexual sporulation. Moreover, similar to the delta sfaD mutant, the delta gpgA mutant was unable to produce sexual fruiting bodies (cleistothecia) in self-fertilization and was severely impaired with cleistothecial development in outcross, indicating that both SfaD and GpgA are required for fruiting body formation. Developmental and morphological defects caused by deletion of flbA encoding an RGS protein negatively controlling FadA-mediated vegetative growth signaling were suppressed by delta gpgA, indicating that GpgA functions in FadA-SfaD-mediated vegetative growth signaling. However, deletion of gpgA could not bypass the need for the early developmental activator FluG in asexual sporulation, suggesting that GpgA functions in a separate signaling pathway. We propose that GpgA is the only A. nidulans G gamma-subunit and is required for normal vegetative growth as well as proper asexual and sexual developmental progression.     

Sporogenic effect of polyunsaturated fatty acids on development of Aspergillus spp.

Aspergillus spp. are frequently occurring seed-colonizing fungi that complete their disease cycles through the development of asexual spores, which function as inocula, and through the formation of cleistothecia and sclerotia. We found that development of all three of these structures in Aspergillus nidulans, Aspergillus flavus, and Aspergillus parasiticus is affected by linoleic acid and light. The specific morphological effects of linoleic acid include induction of precocious and increased asexual spore development in A. flavus and A. parasiticus strains and altered sclerotium production in some A. flavus strains in which sclerotium production decreases in the light but increases in the dark. In A. nidulans, both asexual spore production and sexual spore production were altered by linoleic acid. Spore development was induced in all three species by hydroperoxylinoleic acids, which are linoleic acid derivatives that are produced during fungal colonization of seeds. The sporogenic effects of these linoleic compounds on A. nidulans are similar to the sporogenic effects of A. nidulans psi factor, an endogenous mixture of hydroxylinoleic acid moieties. Light treatments also significantly increased asexual spore production in all three species. The sporogenic effects of light, linoleic acid, and linoleic acid derivatives on A. nidulans required an intact veA gene. The sporogenic effects of light and linoleic acid on Aspergillus spp., as well as members of other fungal genera, suggest that these factors may be significant environmental signals for fungal development.     

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.     

A Large Cluster of Highly Expressed Genes Is Dispensable for Growth and Development in Aspergillus Nidulans

We investigated the functions of the highly expressed, sporulation-specific SpoC1 genes of Aspergillus nidulans by deleting the entire 38-kb SpoC1 gene cluster. The resultant mutant strain did not differ from the wild type in (1) growth rate, (2) morphology of specialized reproductive structures formed during completion of the asexual or sexual life cycles, (3) sporulation efficiency, (4) spore viability or (5) spore resistance to environmental stress. Thus, deletion of the SpoC1 gene cluster, representing 0.15% of the A. nidulans genome, had no readily detectable phenotypic effects. Implications of this result are discussed in the context of major alterations in gene expression that occur during A. nidulans development.     

Sexual development and cryptic sexuality in fungi: insights from Aspergillus species

Major insights into sexual development and cryptic sexuality within filamentous fungi have been gained from investigations using Aspergillus species. Here, an overview is first given into sexual morphogenesis in the aspergilli, describing the different types of sexual structures formed and how their production is influenced by a variety of environmental and nutritional factors. It is argued that the formation of cleistothecia and accessory tissues, such as Hülle cells and sclerotia, should be viewed as two independent but co-ordinated developmental pathways. Next, a comprehensive survey of over 75 genes associated with sexual reproduction in the aspergilli is presented, including genes relating to mating and the development of cleistothecia, sclerotia and ascospores. Most of these genes have been identified from studies involving the homothallic Aspergillus nidulans, but an increasing number of studies have now in addition characterized 'sex-related' genes from the heterothallic species Aspergillus fumigatus and Aspergillus flavus. A schematic developmental genetic network is proposed showing the inter-relatedness between these genes. Finally, the discovery of sexual reproduction in certain Aspergillus species that were formerly considered to be strictly asexual is reviewed, and the importance of these findings for cryptic sexuality in the aspergilli as a whole is discussed.     

A basic helix-loop-helix protein with similarity to the fungal morphological regulators, Phd1p, Efg1p and StuA, controls conidiation but not dimorphic growth in Penicillium marneffei

Members of the APSES protein group are basic helix-loop-helix (bHLH) proteins that regulate processes such as mating, asexual sporulation and dimorphic growth in fungi. Penicillium marneffei is a human pathogen and is the only member of its genus to display a dimorphic growth transition. At 25 degrees C, P. marneffei grows with a filamentous morphology and produces asexual spores from multicellular con-idiophores. At 37 degrees C, the filamentous morphology is replaced by yeast cells that reproduce by fission. We have cloned and characterized an APSES protein-encoding gene from P. marneffei that has a high degree of similarity to Aspergillus nidulans stuA. Deletion of stuA in P. marneffei showed that it is required for metula and phialide formation during conidiation but is not required for dimorphic growth. This suggests that APSES proteins may control processes that require budding (formation of the metulae and phialides, pseudohyphal growth in Saccharomyces cerevisiae and dimorphic growth in Candida albicans) but not those that require fission (dimorphic growth in P. marneffei). The A. nidulans DeltastuA mutant has defects in both conidiation and mating. The P. marneffei stuA gene was capable of complementing the conidiation defect but could only inefficiently complement the sexual defects of the A. nidulans mutant. This suggests that the P. marneffei gene, which comes from an asexual species, has diverged significantly from the A. nidulans gene with respect to sexual but not asexual development.     

FluG-BrlA途径参与构巢曲霉无性发育机制的研究进展

构巢曲霉是丝状真菌的模式生物,已对其无性发育机制进行了比较充分的研究。本文以FluG-BrlA途径参与构巢曲霉无性发育机制的研究为切入点,综述了构巢曲霉无性发育中心调控路径中各主要成员如brlA、abaA、wetA,中心调控路径修饰基因如stuA、medA及中心调控路径激活因子fluG、flbA-E的研究进展,绘制出构巢曲霉无性发育相关基因遗传位置模式图。研究将为其它丝状真菌无性发育机制的研究提供参考。     

brlA requires both zinc fingers to induce development.

Expression of the Aspergillus nidulans brlA gene induces a developmental pathway leading to the production of asexual spores. We have introduced mutations into brlA that are expected to disrupt either or both Cys2-His2 Zn(II) coordination sites postulated to exist in the BrlA polypeptide. The resultant brlA alleles fail to induce either the asexual reproductive pathway or the expression of development-specific genes. These data support the hypothesis that brlA encodes a nucleic acid-binding protein whose activity requires each of two zinc fingers.