Hyphae Formation of Candida albicans Is Regulated by Polyamines
Biological and Pharmaceutical Bulletin
Candida albicans are opportunistic fungi which generally exist in the oral cavity, skin, vagina and intestinal organs, 1, 2) and are dimorphic fungi which undergo a transition from the yeast form to the hyphal form depending on the growth conditions. 3,4) They grow in yeast form at membranes and the surface of the skin, but in hyphae form at the deep-seated mycosis. Temperature, pH and Ca 2ϩ are known as external factors of the transformation. 5-7) The hyphae transformation is induced by the
... s induced by the activation of the MAP kinase and cAMP pathways, which are controlled by RAS protein. 8, 9) Polyamines are synthesized from ornithine by ornithine decarboxylase (ODC) and known as inducers of adenylate cyclase. 10) In this report, we discovered that polyamines participated in the transformation of C. albicans, and the activation of adenylate cyclase was related to the transformation. MATERIALS AND METHODS Organism and Growth Conditions C. albicans NIH A-207 strain was routinely grown in Sabouraud's medium (peptone 10 g, glucose 20 g and yeast extract 5 g, per liter). To induce hyphae formation, the cells were grown in RPMI1640 medium at 37°C. Reagents 1,4-Diamino-2-butanone (DAB) was purchased from Aldrich Chemical Co. (Mo, U.S.A.). cis-N-[2-Phenylcyclopentyl]-azacyclotridec-1-en-2-amine (MDL-12,330A) was purchased from Sigma Chemical Co. (Mo, U.S.A.). The Amount of Polyamines in C. albicans C. albicans cells (1ϫ10 5 cells/ml in RPMI1640) were incubated with DAB (0, 960 mM) in 5% CO 2 at 37°C for 1 and 3 h. Cells were washed twice with sterile water and disrupted with a Mini-Beadbeater (diameter of glass beads, 0.3 mm, Central Scientific Commerce, Inc.). Total proteins in the supernatant were analyzed at 280 nm and polyamines levels were determined using a Labosearch polyamine auto (A and T Corp.). Yeast Induction of C. albicans by Polyamine Synthesis Inhibitor C. albicans cells were adjusted to 1ϫ10 5 cells/ml in RPMI1640. DAB (0, 30, 60, 120, 240, 480, 960 mM) was 890 Candida albicans generally grows in hyphae form in RPMI1640 medium. However, addition of 1,4-diamino-2-butanone (DAB), a competitive inhibitor of ornithine decarboxylase, decreased the amount of polyamines in C. albicans, and induced the proliferation of the C. albicans yeast form. The expression of CYR1 mRNA was significantly inhibited by the addition of DAB compared with that of the control. The amount of intracellular cAMP was also decreased by the addition of DAB. A specific adenylate cyclase inhibitor, cis-N-[2-phenylcyclopentyl]azacyclotridec-1-en-2-amine (MDL-12,330A) promoted the growth of the yeast form. These results indicated that polyamines exist upstream of the adenylate cyclase-cAMP signal pathway and regulate the transformation of C. albicans.