We are studying the function of host extracellular matrix components in the pathogenesis of Candida albicans. Using in vitro assays to quantify adhesion of C. albicans to immobilized fibronectin and its proteolytic and recombinant fragments and to evaluate binding of soluble fibronectin to C. albicans in suspension, we found that interactions of fibronectin with this organism are not mediated by the Arg-Gly-Asp integrin recognition sequence of fibronectin. High affinity binding, observed following grown in complex medium, is primarily mediated by the collagen-binding domain of fibronectin. A 30 kDa fragment of fibronectin containing the collagen binding domain is as active as intact fibronectin for binding to C. albicans. Expression of fibronectin receptors is tightly regulated by growth conditions. C. albicans grown in defined media do not bind fibronectin. We identified hemoglobin as a highly specific activator of receptor expression, which reversible induces fibronectin binding when added to defined growth medium. This binding is of lower affinity and is mediated by the cell-binding domain of fibronectin. Hemoglobin-inducible binding was observed in all clinical isolates of C. albicans and in other members of the Candida genus. The response to hemoglobin is mediated by a low affinity hemoglobin receptor on C. albicans. This activation may play an important role in pathogenesis, since only pathogenic strains of C. albicans express hemolytic activity. Hemoglobin may therefore be a host environmental cue that triggers extracellular matrix receptor expression at a septic site. Inhibitors of this activation process may decrease the pathogenicity of C. albicans. The second, high affinity fibronectin receptor was found to be induced following inhibition of b1,3 glucan synthetase using subclinical doses of MK0991 (Caspofungin) in a defined medium. Characteristic of the high affinity receptor, binding of 125I-fibronectin was specifically inhibited by a proteolytic collagen-binding domain of fibronectin. Macromolecular complexes (Mr > 500,000) sheared from the cell walls after culture with MK0991 retained high affinity fibronectin binding. The fibronectin binding activity of the complexes was sensitive to inactivation by proteases, disulfide reduction, or periodate oxidation followed by alkaline b-elimination. A mutant strain of C. albicans that lacks b1,6 glucans, Kre9, did not express the high affinity fibronectin receptor but retained hemoglobin-induced fibronectin binding. Induction of a high affinity fibronectin receptor by low doses of MK0991 suggests that defining the pharmacokinetics and tissue distribution of the drug may be critical to maximize its efficacy. We have identified ESTs for several genes whose expression is induced by hemoglobin. Northern analysis showed that these genes differ in their time-dependence induction by hemoglobin. Genomic clones corresponding to three of the genes were obtained and mapped. One of these encodes a gene related to YDL166c in S. cerevisiae. Disruption of this gene in C. albicans was conditional lethal for growth.

Agency
National Institute of Health (NIH)
Institute
Division of Clinical Sciences - NCI (NCI)
Type
Intramural Research (Z01)
Project #
1Z01SC009173-14
Application #
6756862
Study Section
(LP)
Project Start
Project End
Budget Start
Budget End
Support Year
14
Fiscal Year
2002
Total Cost
Indirect Cost
Name
Clinical Sciences
Department
Type
DUNS #
City
State
Country
United States
Zip Code
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Pendrak, Michael L; Roberts, David D (2007) Hemoglobin is an effective inducer of hyphal differentiation in Candida albicans. Med Mycol 45:61-71
Pendrak, Michael L; Rodrigues, Rui G; Roberts, David D (2007) Induction of a high affinity fibronectin receptor in Candida albicans by caspofungin: requirements for beta (1,6) glucans and the developmental regulator Hbr1p. Med Mycol 45:157-68
Pendrak, Michael L; Yan, S Steve; Roberts, David D (2004) Sensing the host environment: recognition of hemoglobin by the pathogenic yeast Candida albicans. Arch Biochem Biophys 426:148-56
Pendrak, Michael L; Yan, S Steve; Roberts, David D (2004) Hemoglobin regulates expression of an activator of mating-type locus alpha genes in Candida albicans. Eukaryot Cell 3:764-75
Pendrak, Michael L; Chao, Mark P; Yan, S Steve et al. (2004) Heme oxygenase in Candida albicans is regulated by hemoglobin and is necessary for metabolism of exogenous heme and hemoglobin to alpha-biliverdin. J Biol Chem 279:3426-33
Pendrak, M L; Krutzsch, H C; Roberts, D D (2000) Structural requirements for hemoglobin to induce fibronectin receptor expression in Candida albicans. Biochemistry 39:16110-8
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