Mannitol-1-phosphate dehydrogenase, MpdA, is required for mannitol production in vegetative cells and involved in hyphal branching, heat resistance of conidia and sexual development in Aspergillus nidulans.

Aspergillus nidulans produces cleistothecia as sexual reproductive organs in a process affected by genetic and external factors. To gain a deeper insight into A. nidulans sexual development, we performed comparative proteome analyses based on the wild type developmental periods. We identified sexual development-specific proteins with a more than twofold increase in production during hypoxia or the sexual period compared to the asexual period. Among the sexual development-specific proteins analyzed by gene-deletion experiments and functional assays, MpdA, a putative mannitol-1-phosphate 5-dehydrogenase, plays multiple roles in growth and differentiation of A. nidulans. The most distinct mpdA-deletion phenotype was ascosporogenesis failure. Genetic mpdA deletion resulted in small cleistothecia with no functional ascospores. Transcriptional analyses indicated that MpdA modulates the expression of key development- and meiosis-regulatory genes during sexual development. The mpdA deletion increased hyphal branching and decreased conidial heat resistance. Mannitol production in conidia showed no difference, whereas it was decreased in mycelia and sexual cultures. Addition of mannitol during vegetative growth recovered the defects in conidial heat resistance and ascospore genesis. Taken together, these results indicate that MpdA plays an important role in sexual development, hyphal branching, and conidial heat resistance in Aspergillus nidulans.

The LAMMER kinase is involved in morphogenesis and response to cell wall- and DNA-damaging stresses in Candida albicans.

Dual specificity LAMMER kinase has been reported to be conserved across species ranging from yeasts to animals and has multiple functions. Candida albicans undergoes dimorphic switching between yeast cells and hyphal growth forms as its key virulence factors. Deletion of KNS1, which encodes for LAMMER kinase in C. albicans, led to pseudohyphal growth on YPD media and defects in filamentous growth both on spider and YPD solid media containing 10% serum. These cells exhibited expanded central wrinkled regions and specifically reduced peripheral filaments. Among the several stresses tested, the kns1Δ strains showed sensitivity to cell-wall and DNA-replicative stress. Under fluorescent microscopy, an increase in chitin decomposition was observed near the bud necks and septa in kns1Δ cells. When the expression levels of genes for cell wall integrity (CWI) and the DNA repair mechanism were tested, the kns1 double-deletion cells showed abnormal patterns compared to wild-type cells; The transcript levels of genes for glycosylphosphatidylinositol (GPI)-anchored proteins were increased upon calcofluor white (CFW) treatment. Under DNA replicative stress, the expression of MluI-cell cycle box binding factor (MBF)-targeted genes, which are expressed during the G1/S transition in the cell cycle, was not increased in the kns1 double-deletion cells. This strain showed increased adhesion to the surface of an agar plate and zebrafish embryo. These results demonstrate that Kns1 is involved in dimorphic transition, cell wall integrity, response to DNA replicative stress, and adherence to the host cell surface in C. albicans.

Clostridium kogasensis sp. nov., a novel member of the genus Clostridium, isolated from soil under a corroded gas pipeline.

Two bacterial strains, YHK0403(T) and YHK0508, isolated from soil under a corroded gas pipe line, were revealed as Gram-negative, obligately anaerobic, spore-forming and mesophilic bacteria. The cells were rod-shaped and motile by means of peritrichous flagella. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the isolates were members of the genus Clostridium and were the most closely related to Clostridium scatologenes KCTC 5588(T) (95.8% sequence similarity), followed by Clostridium magnum KCTC 15177(T) (95.8%), Clostridium drakei KCTC 5440(T) (95.7%) and Clostridium tyrobutyricum KCTC 5387(T) (94.9%). The G + C contents of the isolates were 29.6 mol%. Peptidoglycan in the cell wall was of the A1γ type with meso-diaminopimelic acid. The major polar lipid was diphosphatidylglycerol (DPG), and other minor lipids were revealed as phosphatidylglycerol (PG), phosphatidylethanolamine (PE), two unknown glycolipids (GL1 and GL2), an unknown aminoglycolipid (NGL), two unknown aminophospholipids (PN1 and PN2) and four unknown phospholipids (PL1 to PL4). Predominant fatty acids were C16:0 and C16:1cis9 DMA. The major end products from glucose fermentation were identified as butyrate (12.2 mmol) and acetate (9.8 mmol). Collectively, the results from a wide range of phenotypic tests, chemotaxonomic tests, and phylogenetic analysis indicated that the two isolates represent novel species of the genus Clostridium, for which the name Clostridium kogasensis sp. nov. (type strain, YHK0403(T) = KCTC 15258(T) = JCM 18719(T)) is proposed.

Luteimicrobium xylanilyticum sp. nov., isolated from the gut of a long-horned beetle, Massicus raddei.

A novel strain, designated W-15(T), was isolated from the gut of a long-horned beetle, Massicus raddei, collected in South Korea. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the strains belonged to the suborder Micrococcineae. Strain W-15(T) was most closely related to Luteimicrobium album RI148-Li105(T) (97.9 % similarity). Strain W-15(T) was Gram-stain-positive, rod- and coccus-shaped and non-motile. Growth was observed at 15-37 °C, at pH 4.5-8.5 and in the presence of 0-5.0 % NaCl. The cell-wall peptidoglycan of the strain was A4α (l-Lys-d-Ser-d-Asp). The major menaquinone present in this strain was MK-8 (H2) and the major cellular fatty acids were anteiso-C15 : 0, iso-C16 : 0, iso-C15 : 0 and anteiso-C17 : 0. The major polar lipids were diphosphatidylglycerol, phosphatidylinositol, an unknown lipid, an unknown phospholipid and an unknown phosphoglycolipid. The G+C content of genomic DNA of the strain was 73.8 mol%. On the basis of evidence from our polyphasic taxonomic study, strain W-15(T) is classified as representing a novel species in the suborder Micrococcineae, for which the name Luteimicrobium xylanilyticum sp. nov. is proposed. The type strain of this species is strain W-15(T) ( = KCTC 19882(T) = JCM 18090(T)).

LAMMER Kinase Governs the Expression and Cellular Localization of Gas2, a Key Regulator of Flocculation in Schizosaccharomyces pombe.

It was reported that LAMMER kinase in Schizosaccharomyces pombe plays an important role in cation-dependent and galactose-specific flocculation. Analogous to other flocculating yeasts, when cell wall extracts of the Δlkh1 strain were treated to the wild-type strain, it displayed flocculation. Gas2, a 1,3-ß-glucanosyl transferase, was isolated from the EDTA-extracted cell-surface proteins in the Δlkh1 strain. While disruption of the gas2+ gene was not lethal and reduced the flocculation activity of the ∆lkh1 strain, the expression of a secreted form of Gas2, in which the GPI anchor addition sequences had been removed, conferred the ability to flocculate upon the WT strain. The Gas2-mediated flocculation was strongly inhibited by galactose but not by glucose. Immunostaining analysis showed that the cell surface localization of Gas2 was crucial for the flocculation of fission yeast. In addition, we identified the regulation of mbx2+ expression by Lkh1 using RT-qPCR. Taken together, we found that Lkh1 induces asexual flocculation by regulating not only the localization of Gas2 but also the transcription of gas2+ through Mbx2.

Fission yeast LAMMER kinase Lkh1 regulates the cell cycle by phosphorylating the CDK-inhibitor Rum1.

In eukaryotes, LAMMER kinases are involved in various cellular events, including the cell cycle. However, no attempt has been made to investigate the mechanisms that underlie the involvement of LAMMER kinase. In this study, we performed a functional analysis of LAMMER kinase using the fission yeast, Schizosaccharomyces pombe. FACS analyses revealed that deletion of the gene that encodes the LAMMER kinase Lkh1 made mutant cells pass through the G1/S phase faster than their wild-type counterparts. Co-immunoprecipitation and an in vitro kinase assay also revealed that Lkh1 can interact with and phosphorylate Rum1 to activate this molecule as a cyclin-dependent kinase inhibitor, which blocks cell cycle progression from the G1 phase to the S phase. Peptide mass fingerprinting and kinase assay with Rum1(T110A) confirmed T110 as the Lkh1-dependent phosphorylation residue. In this report we present for the first time a positive acting mechanism that is responsible for the CKI activity of Rum1, in which the LAMMER kinase-mediated phosphorylation of Rum1 is involved.

LAMMER Kinase Modulates Cell Cycle by Phosphorylating the MBF Repressor, Yox1, in Schizosaccharomyces pombe.

Lkh1, a LAMMER kinase homolog in the fission yeast Schizosaccharomyces pombe, acts as a negative regulator of filamentous growth and flocculation. It is also involved in the response to oxidative stress. The lkh1-deletion mutant displays slower cell growth, shorter cell size, and abnormal DNA content compared to the wild type. These phenotypes suggest that Lkh1 controls cell size and cell cycle progression. When we performed microarray analysis using the lkh1-deletion mutant, we found that only four of the up-regulated genes in the lkh1-deletion were associated with the cell cycle. Interestingly, all of these genes are regulated by the Mlu1 cell cycle box binding factor (MBF), which is a transcription complex responsible for regulating the expression of cell cycle genes during the G1/S phase. Transcription analyses of the MBF-dependent cell-cycle genes, including negative feedback regulators, confirmed the up-regulation of these genes by the deletion of lkh1. Pull-down assay confirmed the interaction between Lkh1 and Yox1, which is a negative feedback regulator of MBF. This result supports the involvement of LAMMER kinase in cell cycle regulation by modulating MBF activity. In vitro kinase assay and NetPhosK 2.0 analysis with the Yox1T40,41A mutant allele revealed that T40 and T41 residues are the phosphorylation sites mediated by Lkh1. These sites affect the G1/S cell cycle progression of fission yeast by modulating the activity of the MBF complex.

Survival Factor A (SvfA) Contributes to Aspergillus nidulans Pathogenicity.

Survival factor A (SvfA) in Aspergillus nidulans plays multiple roles in growth and developmental processes. It is a candidate for a novel VeA-dependent protein involved in sexual development. VeA is a key developmental regulator in Aspergillus species that can interact with other velvet-family proteins and enter into the nucleus to function as a transcription factor. In yeast and fungi, SvfA-homologous proteins are required for survival under oxidative and cold-stress conditions. To assess the role of SvfA in virulence in A. nidulans, cell wall components, biofilm formation, and protease activity were evaluated in a svfA-gene-deletion or an AfsvfA-overexpressing strain. The svfA-deletion strain showed decreased production of ß-1,3-glucan in conidia, a cell wall pathogen-associated molecular pattern, with a decrease in gene expression for chitin synthases and ß-1,3-glucan synthase. The ability to form biofilms and produce proteases was reduced in the svfA-deletion strain. We hypothesized that the svfA-deletion strain was less virulent than the wild-type strain; therefore, we performed in vitro phagocytosis assays using alveolar macrophages and analyzed in vivo survival using two vertebrate animal models. While phagocytosis was reduced in mouse alveolar macrophages challenged with conidia from the svfA-deletion strain, the killing rate showed a significant increase with increased extracellular signal-regulated kinase ERK activation. The svfA-deletion conidia infection reduced host mortality in both T-cell-deficient zebrafish and chronic granulomatous disease mouse models. Taken together, these results indicate that SvfA plays a significant role in the pathogenicity of A. nidulans.

Small heat-shock protein Hsp9 has dual functions in stress adaptation and stress-induced G2-M checkpoint regulation via Cdc25 inactivation in Schizosaccharomyces pombe.

The small heat-shock protein Hsp9 from Schizosaccharomyces pombe was previously reported to be a homologue of Saccharomyces cerevisiae HSP12. Although Hsp9 is expressed in response to heat shock and nutritional limitation, its function is still not completely understood. Here, we explored the biological function of Hsp9 in S. pombe. The hsp9 gene might play a role in stress adaptation; hsp9 deletion caused heat sensitivity and overexpression induced heat tolerance. In addition, Hsp9 also contribute to cell cycle regulation in the nucleus. Δhsp9 cells grew more quickly and were shorter in length than wild-type cells. Moreover, Δhsp9 cells did not achieve checkpoint arrest under stress conditions, leading to cell death, and exhibited a short doubling time and short G2 phase. Overexpression of hsp9 induced cell cycle delay, increased the population of G2 phase cells, and rescued the phenotypes of cdc2-33, cdc25-22, Δrad24, and Δrad25 mutants, suggesting that Hsp9 probably regulates Cdc2 phosphorylation by modulating the Cdc25 activity. Indeed, immunoprecipitation experiments revealed that Hsp9 is associated with 14-3-3 and Cdc25. In Δhsp9 cells, the association of 14-3-3 with Cdc25 was weakened and Cdc2 phosphorylaton was reduced. Together, our data suggest that Hsp9 has dual functions in stress adaptation and regulating a G2-M checkpoint by the Cdc25 inactivation; this differs from S. cerevisiae HSP12, which maintains cell membrane stability under stress conditions.

Gryllotalpicola gen. nov., with descriptions of Gryllotalpicola koreensis sp. nov., Gryllotalpicola daejeonensis sp. nov. and Gryllotalpicola kribbensis sp. nov. from the gut of the African mole cricket, Gryllotalpa africana, and reclassification of Curtobacterium ginsengisoli as Gryllotalpicola ginsengisoli comb. nov.

Strains RU-16(T), RU-28, RU-04(T) and PU-02(T) were isolated from the gut of the African mole cricket, Gryllotalpa africana. Phylogenetic analyses based on 16S rRNA gene sequences revealed that the strains belonged to the family Microbacteriaceae. All four strains were most closely related to Curtobacterium ginsengisoli DCY26(T) (below 97% 16S rRNA gene sequence similarity). These isolates were Gram-stain-positive, motile (by gliding), rod-shaped and exhibited ivory-coloured colonies. Their chemotaxonomic properties included MK-11 as the major respiratory quinone, ornithine as the cell-wall diamino acid, acetyl as the acyl type of the peptidoglycan, cyclohexyl-C(17:0) as the major fatty acid and phosphatidylglycerol and diphosphatidylglycerol as the major polar lipids. On the basis of phenotypic, chemotaxonomic and phylogenetic analyses, we propose a new genus in the family Microbacteriaceae, Gryllotalpicola gen. nov., with three novel species, Gryllotalpicola daejeonensis sp. nov. (type strain RU-04(T)  =KCTC 13809(T)  =JCM 17590(T)), Gryllotalpicola koreensis sp. nov. (type strain RU-16(T)  = KCTC 13810(T)  =JCM 17591(T)) and Gryllotalpicola kribbensis sp. nov. (type strain PU-02(T)  =KCTC 13808(T)  =JCM 17593(T)). Gryllotalpicola koreensis is the type species of the genus. Additionally, we propose that Curtobacterium ginsengisoli should be reclassified in the genus as Gryllotalpicola ginsengisoli comb. nov. (type strain DCY26(T)  =KCTC 13163(T)  = JCM 14773(T)).

The Gß-like Protein AfCpcB Affects Sexual Development, Response to Oxidative Stress and Phagocytosis by Alveolar Macrophages in Aspergillus fumigatus.

G-protein signaling is important for signal transduction, allowing various stimuli that are external to a cell to affect its internal molecules. In Aspergillus fumigatus, the roles of Gß-like protein CpcB on growth, asexual development, drug sensitivity, and virulence in a mouse model have been previously reported. To gain a deeper insight into Aspergillus fumigatus sexual development, the ΔAfcpcB strain was generated using the supermater AFB62 strain and crossed with AFIR928. This cross yields a decreased number of cleistothecia, including few ascospores. The sexual reproductive organ-specific transcriptional analysis using RNAs from the cleistothecia (sexual fruiting bodies) indicated that the CpcB is essential for the completion of sexual development by regulating the transcription of sexual genes, such as veA, steA, and vosA. The ΔAfcpcB strain revealed increased resistance to oxidative stress by regulating genes for catalase, peroxiredoxin, and ergosterol biosynthesis. The ΔAfcpcB strain showed decreased uptake by alveolar macrophages in vitro, decreased sensitivity to Congo red, decreased expression of cell wall genes, and increased expression of the hydrophobin genes. Taken together, these findings indicate that AfCpcB plays important roles in sexual development, phagocytosis by alveolar macrophages, biosynthesis of the cell wall, and oxidative stress response.

Different inhaled corticosteroid doses in triple therapy for chronic obstructive pulmonary disease: systematic review and Bayesian network meta-analysis.

A systematic review and Bayesian network meta-analysis is necessary to evaluate the efficacy and safety of triple therapy with different doses of inhaled corticosteroids (ICS) in stable chronic obstructive pulmonary disease (COPD). We selected 26 parallel randomized controlled trials (41,366 patients) comparing triple therapy with ICS/long-acting beta-agonist (LABA), LABA/long-acting muscarinic antagonist (LAMA), and LAMA in patients with stable COPD for ≥ 12 weeks from PubMed, EMBASE, the Cochrane Library, and clinical trial registries (search from inception to June 30, 2022). Triple therapy with high dose (HD)-ICS exhibited a lower risk of total exacerbation in pre-specified subgroups treated for ≥ 48 weeks than that with low dose (LD)-ICS (odds ratio [OR] = 0.66, 95% credible interval [CrI] = 0.52-0.94, low certainty of evidence) or medium dose (MD)-ICS (OR = 0.66, 95% CrI = 0.51-0.94, low certainty of evidence). Triple therapy with HD-ICS exhibited a lower risk of moderate-to-severe exacerbation in pre-specified subgroups with forced expiratory volume in 1 s < 65% (OR = 0.6, 95% CrI = 0.37-0.98, low certainty of evidence) or previous exacerbation history (OR = 0.6, 95% CrI = 0.36-0.999, very low certainty of evidence) than triple therapy with MD-ICS. Triple therapy with HD-ICS may reduce acute exacerbation in patients with COPD treated with other drug classes including triple therapy with LD- or MD-ICS or dual therapies.

The LAMMER kinase homolog, Lkh1, regulates Tup transcriptional repressors through phosphorylation in Schizosaccharomyces pombe.

Disruption of the fission yeast LAMMER kinase, Lkh1, gene resulted in diverse phenotypes, including adhesive filamentous growth and oxidative stress sensitivity, but an exact cellular function had not been assigned to Lkh1. Through an in vitro pull-down approach, a transcriptional repressor, Tup12, was identified as an Lkh1 binding partner. Interactions between Lkh1 and Tup11 or Tup12 were confirmed by in vitro and in vivo binding assays. Tup proteins were phosphorylated by Lkh1 in a LAMMER motif-dependent manner. The LAMMER motif was also necessary for substrate recognition in vitro and cellular function in vivo. Transcriptional activity assays using promoters negatively regulated by Tup11 and Tup12 showed 6 or 2 times higher activity in the Delta lkh1 mutant than the wild type, respectively. Northern analysis revealed derepressed expression of the fbp1(+) mRNA in Delta lkh1 and in Delta tup11 Delta tup12 mutant cells under repressed conditions. Delta lkh1 and Delta tup11 Delta tup12 mutant cells showed flocculation, which was reversed by co-expression of Tup11 and -12 with Ssn6. Here, we presented a new aspect of the LAMMER kinase by demonstrating that the activities of global transcriptional repressors, Tup11 and Tup12, were positively regulated by Lkh1-mediated phosphorylation.

The activation of p38 MAPK primarily contributes to UV-induced RhoB expression by recruiting the c-Jun and p300 to the distal CCAAT box of the RhoB promoter.

The Ras-related small GTP-binding protein RhoB is rapidly induced in response to genotoxic stresses caused by ionizing radiation. It is known that UV-induced RhoB expression results from the binding of activating transcription factor 2 (ATF2) via NF-Y to the inverted CCAAT box (-23) of the RhoB promoter. Here, we show that the association of c-Jun with the distal CCAAT box (-72) is primarily involved in UV-induced RhoB expression and p38 MAPK regulated RhoB induction through the distal CCAAT box. UV-induced RhoB expression and apoptosis were markedly attenuated by pretreatment with the p38 MAPK inhibitor. siRNA knockdown of RhoB, ATF2 and c-Jun resulted in decreased RhoB expression and eventually restored the growth of UV-irradiated Jurkat cells. In the reporter assay using luciferase under the RhoB promoter, inhibition of RhoB promoter activity by the p38 inhibitor and knockdown of c-Jun using siRNA occurred through the distal CCAAT box. Immunoprecipitation and DNA affinity protein binding assays revealed the association of c-Jun and p300 via NF-YA and the dissociation of histone deacetylase 1 (HDAC1) via c-Jun recruitment to the CCAAT boxes of the RhoB promoter. These results suggest that the activation of p38 MAPK primarily contributes to UV-induced RhoB expression by recruiting the c-Jun and p300 proteins to the distal CCAAT box of the RhoB promoter in Jurkat cells.

N,N-Diethyl-4-[9-meth-oxy-6-(4-methoxy-phen-yl)-5-methyl-2-phenyl-2H-benzo[h]chromen-2-yl]aniline.

In the title compound, C(38)H(37)NO(3), the pyran ring has an envelope conformation with the quaternary C(q) atom as the flap atom. The dihedral angle formed between the meth-oxy-phenyl group and the naphthalene ring system is 67.32 (6)°. The ethyl-amino groups lie to the same side of the plane through the phenyl ring and form dihedral angles of 84.6 (3) and 75.8 (2)° with it.

Methyl 9-diethyl-amino-2,2-bis-(4-meth-oxy-phen-yl)-2H-benzo[h]chromene-5-carboxyl-ate.

In the title compound, C(31)H(29)NO(5), the methyl carboxyl-ate and dimethyl-amino groups on the naphtho-pyran group are almost coplanar with the naphtho-pyran ring system [r.m.s. deviations = 0.08 (2) and 0.161 (2) Å, respectively]. The dihedral angle between the methyl carboxyl-ate and dimethyl-amino groups is 4.9 (1)°. The pyran ring has an envelope conformation with the quaternary C atom out of plane by 0.4739 (13) Å. The meth-oxy-phenyl substituent forms a dihedral angle of 16.6 (1)° with the plane of the benzene ring, while the other meth-oxy-phenyl group is almost coplanar, making a dihedral angle of 1.4 (1)°.

Corrigendum: The LAMMER Kinase, LkhA, Affects Aspergillus fumigatus Pathogenicity by Modulating Reproduction and Biosynthesis of Cell Wall PAMPs.

[This corrects the article DOI: 10.3389/fcimb.2021.756206.].

The LAMMER Kinase, LkhA, Affects Aspergillus fumigatus Pathogenicity by Modulating Reproduction and Biosynthesis of Cell Wall PAMPs.

The LAMMER kinase in eukaryotes is a well-conserved dual-specificity kinase. Aspergillus species cause a wide spectrum of diseases called aspergillosis in humans, depending on the underlying immune status of the host, such as allergy, aspergilloma, and invasive aspergillosis. Aspergillus fumigatus is the most common opportunistic fungal pathogen that causes invasive aspergillosis. Although LAMMER kinase has various functions in morphology, development, and cell cycle regulation in yeast and filamentous fungi, its function in A. fumigatus is not known. We performed molecular studies on the function of the A. fumigatus LAMMER kinase, AfLkhA, and reported its involvement in multiple cellular processes, including development and virulence. Deletion of AflkhA resulted in defects in colonial growth, production of conidia, and sexual development. Transcription and genetic analyses indicated that AfLkhA modulates the expression of key developmental regulatory genes. The AflkhA-deletion strain showed increased production of gliotoxins and protease activity. When conidia were challenged with alveolar macrophages, enodocytosis of conidia by macrophages was increased in the AflkhA-deletion strain, resulting from changes in expression of the cell wall genes and thus content of cell wall pathogen-associated molecular patterns, including ß-1,3-glucan and GM. While T cell-deficient zebrafish larvae were significantly susceptible to wild-type A. fumigatus infection, AflkhA-deletion conidia infection reduced host mortality. A. fumigatus AfLkhA is required for the establishment of virulence factors, including conidial production, mycotoxin synthesis, protease activity, and interaction with macrophages, which ultimately affect pathogenicity at the organismal level.

The putative C2H2 transcription factor RocA is a novel regulator of development and secondary metabolism in Aspergillus nidulans.

Multiple transcriptional regulators play important roles in the coordination of developmental processes, including asexual and sexual development, and secondary metabolism in the filamentous fungus Aspergillus nidulans. In the present study, we characterized a novel putative C2H2-type transcription factor (TF), RocA, in relation to development and secondary metabolism. Deletion of rocA increased conidiation and caused defective sexual development. In contrast, the overexpression of rocA exerted opposite effects on both phenotypes. Additionally, nullifying rocA resulted in enhanced brlA expression and reduced nsdC expression, whereas its overexpression exerted the opposite effects. These results suggest that RocA functions as a negative regulator of asexual development by repressing the expression of brlA encoding a key asexual development activator, but as a positive regulator of sexual development by enhancing the expression of nsdC encoding a pivotal sexual development activator. Deletion of rocA increased the production of sterigmatocystin (ST), as well as the expression of its biosynthetic genes, aflR and stcU. Additionally, the expression of the biosynthetic genes for penicillin (PN), ipnA and acvA, and for terrequinone (TQ), tdiB and tdiE, was increased by rocA deletion. Thus, it appears that RocA functions as a negative transcriptional modulator of the secondary metabolic genes involved in ST, PN, and TQ biosynthesis. Taken together, we propose that RocA is a novel transcriptional regulator that may act either positively or negatively at multiple target genes necessary for asexual and sexual development and secondary metabolism.

Survival factor SvfA plays multiple roles in differentiation and is essential for completion of sexual development in Aspergillus nidulans.

The first member of the velvet family of proteins, VeA, regulates sexual development and secondary metabolism in the filamentous fungus Aspergillus nidulans. In our study, through comparative proteome analysis using wild type and veA-deletion strains, new putative regulators of sexual development were identified and functionally analyzed. Among these, SvfA, containing a yeast survival factor 1 domain, plays multiple roles in the growth and differentiation of A. nidulans. Deletion of the svfA gene resulted in increased sensitivity to oxidative and cold stress as in yeast. The svfA-deletion strain showed an increase in bi-polar germination and a decrease in radial growth rate. The deletion strain formed structurally abnormal conidiophores and thus produced lower amounts of conidiospores during asexual development. The svfA-deletion strain produced few Hülle cells and small cleistothecia with no ascospores, indicating the requirement of svfA for the completion of sexual development. Transcription and genetic analyses indicated that SvfA modulates the expression of key development regulatory genes. Western blot analysis revealed two forms of SvfA. The larger form showed sexual-specific and VeA-dependent production. Also, the deletion of svfA caused decreased ST (sterigmatocystin) production. We propose that SvfA is a novel central regulator of growth, differentiation and secondary metabolism in A. nidulans.