Histoplasma acquisition of calcium and expression of CBP1 during intracellular parasitism

Abstract

SummaryA highly adapted parasite of macrophages, the yeast phase of Histoplasma capsulatum, survives and proliferates within phagolysosomes, while the mycelial phase exists only as a saprophyte in the soil. We have shown previously that these two phases of Histoplasma differ in their calcium requirements for growth and in the production of a released calcium-binding protein (CBP). Cloning and sequencing the CBP1 gene revealed two introns, a putative signal peptide and potential calcium-binding sites. We also evaluated CBP1 expression by reverse transcription-polymerase chain reaction (RT-PCR) of yeasts grown in broth culture and within two host cell types, a macrophage-like cell line and respiratory epithelial cells. H. capsulatum yeasts expressed CBP1 in all of these settings. Splenocytes from mice immunized with H. capsulatum yeasts responded to purified CBP in proliferation assays, providing evidence for the production of CBP during the infection of mammalian hosts. In addition, after H. capsulatum yeasts were subjected to a calcium-free shock, exogenously added CBP allowed yeasts to incorporate more calcium than yeasts incubated without added CBP. These results suggest that CBP may function to provide yeasts with calcium when they are in a low-calcium environment, such as the phagolysosomal compartment within macrophages.

Phenotypic variation and intracellular parasitism by Histoplasma capsulatum

“…Further studies with reverse transcription-PCR demonstrated that CBP1 continues to be expressed while H. capsulatum yeasts grow inside P388D1 cells. This expression is consistent with experiments with mice inoculated with H. capsulatum; splenocytes harvested later from these mice respond to purified CBP in proliferation assays, implying that CBP is secreted during infection of mammalian hosts (17). All of these regulatory studies have used either mycelial or yeast cultures of H. capsulatum, but there has been no means to examine carefully the developmental programming of CBP1 expression during the transition between these two forms.…”

Fungal Morphogenesis

“…Therefore, morphological and physiological plasticity allows fungi to rapidly adapt to changing extracellular conditions. Species-specific signaling and morphological features appear to be a direct result of fungal attempts to survive as new microenvironments, and their particular cell stresses, were encountered (Hogan and Klein 1994;Newman et al 1995;Batanghari et al 1998;Sebghati et al 2000;Gow et al 2002;Brandhorst et al 2004;Rappleye et al 2004Rappleye et al , 2007Gantner et al 2005;Nemecek et al 2006;Gauthier and Klein 2008;Nather and Munro 2008;Mora-Montes et al 2011;. The concerted action of morphotype and physiological changes in the context of a particular environment are therefore critical for successful fungal adaptation (Butler et al 2009;O'Connor et al 2010).…”

“…Additionally, these different fungal morphotypes can display altered tolerance of certain host physiological conditions, as well as varying abilities to disseminate in different host tissues (Klein and Tebbets 2007). The pathogenic Histoplasma yeast cells, and not the saprophytic hyphal cells, secrete Cbp1 to acquire calcium to better survive within phagolysosomes (Batanghari et al 1998). Cbp1 supports the growth of intracellular yeast cells and is required for Histoplasma virulence in animal models (Sebghati et al 2000).…”

Coevolution of Morphology and Virulence in Candida Species

“…Consistent with this hypothesis, disruption of the H. capsulatum yeast-specific gene AGS1, encoding ␣-(1,3)-glucan synthase, reduces both pathogenesis and the ability of yeast to grow in macrophages in vitro (114). Additional yeast phase-specific virulence factors include B. dermatitidis BAD1, an adhesin that binds complement type 3 receptors, as well as CD14 on lung cells and macrophages, to manipulate the host immune response (12,98), and H. capsulatum CBP1, a calcium-binding protein required for pathogenicity, in addition to growth in macrophages in vitro (8,122).…”