Blastomycosis is a progressive pulmonary infection of immune-competent and -compromised hosts, but the factors responsible for pathogenicity of the causative fungus Blastomyces dermatitidis are not well understood. We identified BAD1 (formerly WI-1), a surface antigen and adhesin on B. dermatitidis, and developed tools to genetically manipulate B. dermatitidis. On targeting and disrupting BAD1, we found that BAD1 is an essential virulence factor that suppresses host TNF-ct responses and promotes fungal evasion of immunity. BAD1 displays yeast-phase specific expression, which is regulated transcriptionally by a mechanism shared in B. dermatitidis and Histoplasma capsulatum. Strong preliminary data indicate that BAD I's tandem repeat contributes to virulence, and its C-terminal domain fixes the protein to the yeast surface. We hypothesize that BAD1 is a multi-domain protein whose structural motifs - a core, tandem repeat domain with EF hands for Ca _ binding and a C- terminal EGF-like domain - must be present and assembled properly to mediate pivotal biological functions in the host-pathogen interaction. We also postulate that upstream regulators conserved in B. dermatitidis and It. capsulatum govern expression of BAD1 and other yeast-phase genes. Our 3 specific aims are to: 1) _Eluci_te the fur!ctional role of BAD1 domains, BAD1 deletions and domain swaps will be engineered and expressed in bad1 null 13. dermatitidis. These strains and 6 HIS-tagged BAD1 derivatives purified from them, or their synthetic peptides, will be used to define the key domains in pathogenesis, and their modes of action. 2) Define critical features of B_I fijnctional domains. Structural bases for virulence and TNF-ct suppression due to the tandem repeats will be defined. Ca 2?binding by repeats will be studied for effects on BADI's structure and function. Mutations of the repeat EF hand will assess effects on BAD1 Ca 2+ binding, shape and function. 3) Idcr!ti_, re_lators of BADt transcription. Insertional mutants of a reporter strain with a PBADI-lacZ fusion will be screened for impaired PBAOtregulation due to insertion into regulatory genes. Regulators of BAD1, other co-regulated phase-specific genes and morphogenesis will be cloned and studied from among the mutated genes. Our work will resolve the molecular and structural bases of BIastomcyes virulence and define how BAD1 is regulated and controlled coordinately with other yeast-phase genes, which is a key event in fungal pathogenesis.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37AI035681-19
Application #
8415581
Study Section
Special Emphasis Panel (NSS)
Program Officer
Duncan, Rory A
Project Start
1995-01-01
Project End
2013-12-31
Budget Start
2013-01-01
Budget End
2013-12-31
Support Year
19
Fiscal Year
2013
Total Cost
$403,305
Indirect Cost
$131,719
Name
University of Wisconsin Madison
Department
Pediatrics
Type
Schools of Medicine
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Hernández-Santos, Nydiaris; Wiesner, Darin L; Fites, J Scott et al. (2018) Lung Epithelial Cells Coordinate Innate Lymphocytes and Immunity against Pulmonary Fungal Infection. Cell Host Microbe 23:511-522.e5
Nanjappa, Som G; Mudalagiriyappa, Srinivasu; Fites, J Scott et al. (2018) CBLB Constrains Inactivated Vaccine-Induced CD8+ T Cell Responses and Immunity against Lethal Fungal Pneumonia. J Immunol 201:1717-1726
Lorenzini, Jenna; Scott Fites, J; Nett, Jeniel et al. (2017) Blastomyces dermatitidis serine protease dipeptidyl peptidase IVA (DppIVA) cleaves ELR+ CXC chemokines altering their effects on neutrophils. Cell Microbiol 19:
Wiesner, Darin L; Klein, Bruce S (2017) Lung epithelium: barrier immunity to inhaled fungi and driver of fungal-associated allergic asthma. Curr Opin Microbiol 40:8-13
Hernández-Santos, Nydiaris; Klein, Bruce S (2017) Through the Scope Darkly: The Gut Mycobiome Comes into Focus. Cell Host Microbe 22:728-729
Sterkel, Alana K; Mettelman, Robert; Wüthrich, Marcel et al. (2015) The unappreciated intracellular lifestyle of Blastomyces dermatitidis. J Immunol 194:1796-805
Beaussart, Audrey; Brandhorst, Tristan; Dufrêne, Yves F et al. (2015) Blastomyces Virulence Adhesin-1 Protein Binding to Glycosaminoglycans Is Enhanced by Protein Disulfide Isomerase. MBio 6:e01403-15
Nanjappa, Som G; Klein, Bruce S (2014) Vaccine immunity against fungal infections. Curr Opin Immunol 28:27-33
Verma, Akash; Wüthrich, Marcel; Deepe, George et al. (2014) Adaptive immunity to fungi. Cold Spring Harb Perspect Med 5:a019612
Marty, Amber J; Wüthrich, Marcel; Carmen, John C et al. (2013) Isolation of Blastomyces dermatitidis yeast from lung tissue during murine infection for in vivo transcriptional profiling. Fungal Genet Biol 56:1-8

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