Regulation of hyphal morphogenesis in Candida albicans can occur through quorum sensing (QS). A QS signal, farnesol, is produced during high-density growth and inhibits morphogenesis. However, the signal transduction pathway that regulates QS is unknown. Here, we show that a C. albicans mutant lacking Chk1p but not either the Sln1p or the Nik1p histidine kinase is refractory to the inhibitory effect of farnesol both in cell suspension and during the formation of a biofilm. This study is the first to demonstrate a role for a two-component signal transduction protein in QS by a eukaryotic organism.Cell density is a critical factor in the regulation of Candida albicans hyphal morphogenesis. At a density of Ͼ10 6 cells/ml, yeast cells do not shift (germinate) to hyphae or do so at low frequencies, while at a density of Ͻ10 6 cells/ml, germination occurs (3). The relationship between cell density and new gene transcription (hyphal morphogenesis) resembles quorum sensing (QS) in some bacteria (14). Recent observations indicate that a QS system operates in C. albicans and that the isoprenoid farnesol is the QS autoinducer signal (12). Cells exposed to farnesol do not germinate, even at low cell densities. However, the regulatory and signal transduction events that direct QS are unknown, not only for C. albicans but for other fungi and eukaryotes in general. In some bacteria, two-component signaling regulates QS. Since C. albicans has several two-component signal proteins that are critical to a number of processes, including cell wall biosynthesis, adaptation to stress conditions, and virulence, our rationale was that farnesol sensing could be mediated through two-component proteins. C. albicans has three hybrid-histidine kinases, two of which have orthologues in Saccharomyces cerevisiae (Sln1p) and Neurospora crassa (Nik1p) that are presumed to play a role in an osmotic stress response (1,15,19,20). The third histidine kinase, Chk1p, has some similarity to two Schizosaccharomyces pombe proteins, Mak2p and Mak3p, which are known to function as sensors for oxidative stress (2, 5). In addition to the histidine kinases, C. albicans has two response regulator proteins, Ssk1p and Skn7p, whose S. cerevisiae homologs act downstream of the Sln1p histidine kinase (11). In C. albicans, Ssk1p and Skn7p function in the adaptation of cells to oxidant stress, while Ssk1p, in addition, regulates the expression of structural cell wall proteins and negatively regulates Chk1p expression (7, 18).The C. albicans strains used for this study have been described previously (4, 6, 9, 20). Unless noted, cells were routinely cultured in YPD (1% yeast extract, 2% dextrose, 2% peptone) or YNB (0.67% yeast nitrogen base [pH 7.0], 50 mM glucose) at 30°C. To assess whether the two-component signal transduction proteins of C. albicans play a role in QS, all strains (see Table 1) were first cultured overnight at 30°C in YPD. Subsequently, the cells were washed twice and then inoculated into 10 ml of prewarmed medium 199 (pH 7.5) with or without 250 M t...

Fatty acid synthase (FAS) has been identified as a potential antifungal target. FAS prepared from Saccharomyces cerevisiae was employed for bioactivity-guided fractionation of Chlorophora tinctoria,Paspalum conjugatum, Symphonia globulifera, Buchenavia parviflora, and Miconia pilgeriana. Thirteen compounds (1-13), including three new natural products (1, 4, 12), were isolated and their structures identified by spectroscopic interpretation. They represented five chemotypes, namely, isoflavones, flavones, biflavonoids, hydrolyzable tannin-related derivatives, and triterpenoids. 3'-Formylgenistein (1) and ellagic acid 4-O-alpha-l-rhamnopyranoside (9) were the most potent compounds against FAS, with IC(50) values of 2.3 and 7.5 microg/mL, respectively. Furthermore, 43 (14-56) analogues of the five chemotypes from our natural product repository and commercial sources were tested for their FAS inhibitory activity. Structure-activity relationships for some chemotypes were investigated. All these compounds were further evaluated for antifungal activity against Candida albicans and Cryptococcus neoformans. Although there were several antifungal compounds in the set, correlation between the FAS inhibitory activity and antifungal activity could not be defined.

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Two spontaneous cerulenin-resistant mutants of Candida albicans, 4918-2 and 4918-10, were unable to adhere in vitro in fibrin-platelet clots. Because in vitro adherence correlates well with colonization of nonbacterial thrombotic endocarditis on traumatized valvular endocardium, 50% infectious dose studies were performed with a rabbit model of endocarditis. Wild-type C. albicans required 10(3.6) +/- 0.12 cfu in comparison with 10(5.73) +/- 0.31 and 10(7.3) +/- 0.21 cfu for mutants 4918-2 and 4918-10, respectively. The relative avirulence of mutant strains in producing endocarditis was not attributed to accelerated clearance of these strains from the bloodstream. In fact, clearance of wild-type and mutant strains was almost identical. In the same animals renal candidiasis was observed with all strains of C. albicans, although the number of cfu per gram of kidney was higher after infection with wild-type C. albicans. Thus, strains of C. albicans with reduced ability to adhere in vitro to a fibrin-platelet matrix are relatively avirulent in the rabbit endocarditis model.