Role of cysteine in regulating morphogenesis and mitochondrial activity in the dimorphic fungus Histoplasma capsulatum.

Abstract

Three stages can be distinguished in the temperature-induced mycelial-to-yeast phase transition of Hwtoplasma capsulatum. Stage one is characterized by a progressive decrease in the respiration rate and in the intracellular concentrations of cysteine and other amino acids. By stage two, respiration has ceased completely and free cysteine has fallen to low levels. Exogenous cysteine is required during the second stage for activation of mitochondrial respiration (stage three) and completion of the morphological transition. Mitochondria isolated from cells in the second stage show no respiration with NADH, succinate, or other substrates unless they are first incubated with cysteine. In addition, a novel, cytosolic cysteine oxidase appears during the latter part of the second stage. In stage three, the respiration rate rises, intracellular concentrations offree cysteine and other amino acids increase to levels characteristic of yeast, and the morphological transition is completed. The results support the idea that alterations in cysteine metabolism play a key role in this differentiation process.Histoplasma capsulatum is a dimorphic pathogenic fungus that grows as a multicellular mycelium in soil and as a yeast in the tissues of infected humans and animals (1). Factors that affect morphogenesis and maintenance of one phase of the organism are of considerable interest because growth of the yeast phase appears to be required for pathogenicity (2). In culture, the transition from mycelial to yeast phase is triggered by a shift in temperature from 25°C to 37°C. Growth and maintenance of the yeast phase in culture are also dependent upon a low oxidation-reduction potential (3) and on the presence of cysteine in the culture medium (2, 4). Thus far, however, there has been little insight into the biochemical basis of any of these observations.We have shown that cysteine stimulates oxygen consumption in the yeast phase but not in the mycelial phase (5). We reasoned that this observation might provide a good starting point for the investigation of biochemical mechanisms in phase determination and morphogenesis ofH. capsulatum. In the present work, we show that the cysteine stimulation of oxygen consumption is due to a cytosolic cysteine oxidase activity, which is present only in yeast-phase cells. In addition, experiments monitoring cellular respiration, cysteine levels, and cysteine oxidase activity during the temperature-induced mycelial-to-yeast phase transition support the idea that alterations in cysteine metabolism play a key role in the differentiation process. They also demonstrate the occurrence of novel regulatory mechanisms involving the effect ofcysteine on the mitochondrial respiratory chain. On the basis of our results and work by others, we propose a model that outlines biochemical events at specific stages of the morphological transition.MATERIALS AND METHODS Growth of H. capsulatum. The wild-type Down's strain of H. capsulatum mating type (-) was used in this study. Cells were grown in the defined m...

Involvement of an Alternative Oxidase in Oxidative Stress and Mycelium-to-Yeast Differentiation in Paracoccidioides brasiliensis

“…This might occur by a decreased overflow of electrons in the mitochondrial respiratory chain and/or in association with the external alternative NADH-ubiquinone oxidoreductase involving the oxidation of cytosolic NADH that is generated in the glyoxylate cycle or in fermentative pathways, which subsequently balance the cytosolic redox potential, as we previously suggested (40). It had been reported that H. capsulatum, Blastomyces dermatitidis, and P. brasiliensis possess three different respiratory phases during morphogenesis (37,42). After the temperature shift, there is an uncoupling of oxidative phosphorylation and a progressive decrease of respiration (stage 1).…”

“…During stage 2, cysteine and other sulfhydryl compounds are able to induce the shunt pathways utilizing cytochrome oxidase and alternative oxidase, which is inhibited by cyanide and SHAM but is resistant to antimycin A inhibition (57). It is speculated that the inactivation of mitochondrial respiration during stage 2 results from a temperature-induced increase in oxidation-reduction potential; under this condition, the sulfhydryl compounds would act as reducing agents, which may be required for maintenance of the activity of the remaining mitochondrial components (37). In accordance with these previous reports, our findings show that there is an increase in the carbonylated proteins (oxidative stress) during M-to-Y transition, which is correlated with the upregulation of Pbaox.…”

Effect of Extracellular Factors on Growth and Dimorphism of Rhizopus oryzae with Multiple Enzyme Synthesizing Ability

“…Peptone and yeast extract acted as best growth supplementing nitrogen sources and among the amino acids tested, only cysteine HCl could restore the filamentous form even at higher temperature (Table 2), which might be due to restoration of mitochondrial respiration that would other wise stop during transformation to yeast phase [5,32]. Sub culturing the strain in cysteine HCl supplemented media successfully prevented the appearance of yeast form both at higher temperature and during the normal transition of mycelia-yeast form.…”

“…This is an active approach to survive in a changing environment, which simply shift their thallic organization to adapt and thrive in the new conditions. Dimorphism is studied in many fungi, Histoplasma capsulatum [5], blastomyces dermatitidis, and Sporothrix schenckii [6], Ceratocystis ulmi [7], Paracoccidioides brasiliensis [8], Candida albicans [9], Penicillium marneffei [10], Rhizopus oryzae ATCC 20344 [11]. Mucor dimorphism has interested microbiologists since the time of Pasteur [12].…”

Expression of alpha- and beta-tubulin genes during dimorphic-phase transitions of Histoplasma capsulatum.

“…Three distinct stages, characterized by discrete physiological and biochemical patterns, have been delineated during this morphological transition (15 smaller masses, and the tips of mycelial filaments become constricted ( Fig. 1G to H).…”