The ability to sense and adapt to a hostile host environment is a crucial element for virulence of pathogenic fungi, including Cryptococcus neoformans . These cellular responses are evoked by diverse signaling cascades, including the stress-activated HOG pathway. Despite previous analysis of central components of the HOG pathway, its downstream signaling network is poorly characterized in C. neoformans . Here we performed comparative transcriptome analysis with HOG signaling mutants to explore stress-regulated genes and their correlation with the HOG pathway in C. neoformans . In this study, we not only provide important insights into remodeling patterns of global gene expression for counteracting external stresses but also elucidate novel characteristics of the HOG pathway in C. neoformans . First, inhibition of the HOG pathway increases expression of ergosterol biosynthesis genes and cellular ergosterol content, conferring a striking synergistic antifungal activity with amphotericin B and providing an excellent opportunity to develop a novel therapeutic method for treatment of cryptococcosis. Second, a number of cadmium-sensitive genes are differentially regulated by the HOG pathway, and their mutation causes resistance to cadmium. Finally, we have discovered novel stress defense and HOG-dependent genes, which encode a sodium/potassium efflux pump, protein kinase, multidrug transporter system, and elements of the ubiquitin-dependent system.

In Saccharomyces cerevisiae, the accepted theory is that due to TCA cycle dysfunction, the Δcit1 mutant lacking the mitochondrial enzyme citrate synthase (Cit1) cannot grow on acetate, regardless of the presence of the peroxisomal isoenzyme (Cit2). In this study, we re‐evaluated the roles of Cit1 and Cit2 in acetate utilization and examined the pathway of acetate metabolism by analysing mutants defective in TCA or glyoxylate cycle enzymes. Although Δcit1 cells showed significantly reduced growth on rich acetate medium (YPA), they exhibited growth similar to Δcit2 and the wild‐type cells on minimal acetate medium (YNBA). Impaired acetate utilization by Δcit1Δcit2 cells on YNBA was restored by ectopic expression of either Cit2 or its cytoplasmically localized variants. Deletion of any of the genes for the enzymes solely involved in the TCA cycle (IDH1, KGD1 and LSC1), except for SDH1, caused little defect in acetate utilization on YNBA but resulted in significant growth impairment on YPA. In contrast, cells lacking any of the genes involved in the glyoxylate cycle (ACO1, FUM1, MLS1, ICL1 and MDH2) did not grow on either YNBA or YPA. Deletion of SFC1 encoding the succinate–fumarate carrier also caused similar growth defects on YNBA. Our results suggest that in S. cerevisiae the glyoxylate cycle functions as a competent metabolic pathway for acetate utilization on YNBA, while both the TCA and glyoxylate cycles are essential for growth on YPA. Copyright © 2010 John Wiley & Sons, Ltd.

Beta-glucans are a heterologous group of fibrous glucose polymers that are a major constituent of cell walls in Ascomycetes and Basidiomycetes fungi. Synthesis of β (1,3)- and (1,6)-glucans is coordinated with fungal cell growth and development, thus, is under tight genetic regulation. Here, we report that β-glucan synthesis in both asexual and sexual spores is turned off by the NF-kB like fungal regulators VosA and VelB in Aspergillus nidulans. Our genetic and genomic analyses have revealed that both VosA and VelB are necessary for proper down-regulation of cell wall biosynthetic genes including those associated with β-glucan synthesis in both types of spores. The deletion of vosA or velB results in elevated accumulation of β-glucan in asexual spores. Double mutant analyses indicate that VosA and VelB play an inter-dependent role in repressing β-glucan synthesis in asexual spores. In vivo chromatin immuno-precipitation analysis shows that both VelB and VosA bind to the promoter region of the β-glucan synthase gene fksA in asexual spores. Similarly, VosA is required for proper repression of β-glucan synthesis in sexual spores. In summary, the VosA-VelB hetero-complex is a key regulatory unit tightly controlling proper levels of β-glucan synthesis in asexual and sexual spores.

Expression of chsE encoding one of the five chitin synthases of Aspergillus nidulans was analyzed. Expression of chsE was moderate in conidiophores, but somewhat weaker in vegetative mycelia. During sexual development, chsE was expressed strongly in young cleistothecia and hülle cells, but little in mature sexual structures. Deletion of chsE caused a significant decrease in the chitin content of the cell wall during early sexual development. Expression of chsE was increased by substituting glucose with lactose or by addition of 0.6M KCl or NaCl, but affected little by substituting glucose with sodium acetate. Consequently, chsE was shown to have a mode of expression distinct from those of the other chitin synthase genes, chsA, chsB and chsC.

Background and objectives: Endothelial and monocytic cells appear to play a key role in the initiation and progression of atherosclerosis and restenosis via the upregulation of inflammatory cytokines and the formation of oxidized low-density lipoprotein (ox-LDL). However, the role of smooth muscle cells (SMCs) has been underestimated and is not well understood. It was investigated for the first time that native LDL stimulates human SMCs to secrete IL-8. The aim of this study was to investigate the signaling pathway involved in the upregulation of IL-8, induced by LDL in human aortic SMCs. Methods and results: LDL-induced IL-8 expression (mRNA and protein) is specific to SMCs and likely to be regulated at the transcription level in dose- and time-dependent manners, as judged by experiments with actinomycin D and ELISA. Although both p38 and ERK 1/2 MAPKs were activated by LDL, only p38 MAPK is responsible for the LDL effects, as evidenced by a complete blockade of IL-8 upregulation by SB203580. Pretreatment with catalase significantly decreased the extent of IL-8 upregulation, indicating that H2O2 is necessary for the LDL response. Activation of activator protein (AP)-1, but not nuclear factor (NF)-κB, by p38 or H2O2 appears to be necessary along with the concomitant upregulation of c- fos and c-JUN, as judged by electrophoretic mobility shift and luciferase reporter assays. Conclusions: These data demonstrated that LDL stimulates SMCs to induce IL-8 production in dose- and time-dependent manners at the transcription level and that the LDL signaling in hAoSMCs is conveyed via the generation of H2O2, the phosphorylation of p38 MAPK, the activation of AP-1, and the participation of NF-κB.