William E. Goldman, Ph.D.

goldman@med.unc.edu 519A Mary Ellen Jones CB# 7290 Chapel Hill, NC 27599-7290 919.966.9580

Research

Successful respiratory pathogens must be able to respond swiftly to a wide array of sophisticated defense mechanisms in the mammalian lung. In histoplasmosis, macrophages — a first line of defense in the lower respiratory tract — are effectively parasitized by Histoplasma capsulatum. This process depends on virulence factors produced as this "dimorphic" fungus undergoes a temperature-triggered conversion from a saprophytic mold form to a parasitic yeast form. One such molecule is a calcium-binding protein (CBP) that is secreted preferentially by the yeast form and is essential for Histoplasma virulence. The experiments to unravel CBP structure and function have relied heavily on our development of a telomeric shuttle plasmid that has been used for complementation cloning, gene disruptions, RNA interference, and reporter gene constructs. In addition, random insertional mutagenesis and transcriptional profiling with microarrays are helping us identify and characterize genes involved in the regulation of CBP1 expression.

Another yeast phase-specific product of H. capsulatum is alpha-(1,3)-glucan, a cell wall polysaccharide that is associated with virulence in a variety of fungal pathogens. We have taken two approaches to study alpha-(1,3)-glucan: the first is a forward genetics strategy, using Agrobacterium-mediated insertional mutagenesis, to identify genes implicated in the regulation, synthesis, and processing of this polysaccharide. The second approach uses reverse genetics, combining fungal gene disruption with mammalian RNA-interference, to study the genes involved in production of and response to alpha-(1,3)-glucan. This work has revealed that alpha-(1,3)-glucan on the surface of Histoplasma yeasts masks recognition of the underlying beta-glucan by dectin-1, a macrophage pattern-recognition receptor that is critical in the innate immune response to fungi.

Yersinia pestis also displays two temperature-regulated lifestyles, depending on whether it is colonizing a flea or mammalian host. Inhalation by humans leads to a rapid and overwhelming disease, and we are trying to understand the development of pneumonic plague by studying genes that are activated during the stages of pulmonary colonization. We have generated an oligonucleotide-based microarray corresponding to all of the predicted genes of a pneumonic plague isolate, and this has been used for in vivo transcriptional profiling during experimental infection. We have also developed and characterized a mouse model for studying the pathological and immunological changes during the progression of pneumonic plague. This model system has revealed two sharply contrasting phases to the syndrome: the first phase of infection features rapid bacterial proliferation in the lung, but almost no inflammatory response, symptoms, or pathology; the second phase, starting at approximately 36 hours post-inoculation, is marked by inflammation and pneumonia that lead quickly to death. The utility of this model was highlighted in a recent study that demonstrated how a plasminogen-activating protease (encoded by the pla gene of Y. pestis) is essential for development of the second phase of this disease. We have now initiated a more comprehensive research plan, combining forward genetics with microarray technology, to determine what other bacterial genes are specifically required for the development of these two phases of pneumonic plague.

We are also continuing studies of one of the virulence factors of Bordetella pertussis: tracheal cytotoxin (TCT) is a released fragment of peptidoglycan that causes pulmonary inflammation in pertussis (whooping cough). TCT and endotoxin synergistically trigger respiratory epithelial production of nitric oxide, causing ciliated cell damage that corresponds to the well-known airway cytopathology of pertussis. A variety of host receptor systems have been shown to recognize peptidoglycan fragments, and some – such as the PGRP family – are evolutionarily conserved from flies to mammals. Depending on the relationship between bacteria and host, the results of exposure to TCT can be beneficial or pathological. Our current work is aimed at understanding host responses to TCT that include epithelial defense, cytopathology, and remodeling.

Publications

Rappleye, C.A., J.T. Engle, and W.E. Goldman. 2004. RNA interference in Histoplasma capsulatum demonstrates a role for a-(1,3)-glucan in virulence. Molecular Microbiology 53:153-165.

Koropatnick, T.A., J.T. Engle, M.A. Apicella, E.V. Stabb, W.E. Goldman, and M.J. McFall-Ngai. 2004. Microbial factor-mediated development in a host-bacterial mutualism. Science 306:1186-1188.

Kaneko, T., W.E. Goldman, P. Mellroth, H. Steiner, K. Fukase, S. Kusumoto, W. Harley, A. Fox, D. Golenbock, and N. Silverman. 2004. Monomeric and polymeric gram-negative peptidoglycan but not purified LPS stimulate the Drosophila IMD pathway. Immunity 20:637-649.

Mellroth, P., J. Karlsson, J. Håkansson, N. Schultz, W.E. Goldman, and H. Steiner. 2005. Ligand induced dimerization of Drosophila peptidoglycan recognition proteins. Proceedings of the National Academy of Sciences U.S.A. 102:6455-60.

Lathem, W.W., S.D. Crosby, V.L. Miller, and W.E. Goldman. 2005. Progression of primary pneumonic plague: A mouse model of infection, pathology, and bacterial transcriptional activity. Proceedings of the National Academy of Sciences U.S.A. 102:17786-17791.

Swaminathan, C.P., P.H. Brown, A. Roychowdhury, Q. Wang, R. Guan, N. Silverman, W.E. Goldman, G.-J. Boons, and R.A. Mariuzza. 2006. Dual strategies for peptidoglycan discrimination by peptidoglycan recognition proteins (PGRPs). Proceedings of the National Academy of Sciences U.S.A. 103:684-689.

Kaneko, T., T. Yano, K. Aggarwal, J.H. Lim, K. Ueda, Y. Oshima, C. Peach, D. Erturk-Hasdemir, W.E. Goldman, B.H. Oh, S. Kurata and N. Silverman. 2006. PGRP-LC and PGRP-LE play essential yet distinct roles in the Drosophila immune response to monomeric DAP-type peptidoglycan. Nature Immunology 7:715-723.

Rappleye, C.A., and W.E. Goldman. 2006. Defining virulence genes in the dimorphic fungi. Annual Review of Microbiology 60:281-303.

Mielcarek, N., A.-S. Debrie, D. Raze, J. Bertout, A. Ben Younes, J. Engle, W.E. Goldman, and C. Locht. 2006. Live attenuated Bordetella pertussis as a highly efficient single-dose mucosal vaccine against whooping cough. PLoS Pathogens 2:e65.

Cathelyn, J., S.D. Crosby, W.W. Lathem, W.E. Goldman, and V.L. Miller. 2006. RovA, a global regulator of Yersinia pestis, specifically required for bubonic plague. Proceedings of the National Academy of Sciences U.S.A. 103:13514-13519.

Marion, C.L., C.A. Rappleye, J.T. Engle, and W.E. Goldman. 2006. An alpha-(1,4)-amylase is essential for alpha-(1,3)-glucan production and virulence in Histoplasma capsulatum. Molecular Microbiology 62:970-983.

Cloud-Hansen, K.A., S.B. Peterson, E.V. Stabb, W.E. Goldman, M.J. McFall-Ngai, and J. Handelsman. 2006. Breaching the great wall: Peptidoglycan and microbial interactions. Nature Reviews Microbiology 4:710-716.

Almeida, A.J., J.A. Carmona, C. Cunha, A. Carvalho, C.A. Rappleye, W.E. Goldman, P.J. Hooykaas, C. Leão, P. Ludivico, and F. Rodrigues. 2007. Towards a molecular genetic system for the pathogenic fungus Paracoccidioides brasiliensis. Fungal Genetics and Biology 44:1387-1398.

Rappleye, C.A., L.G. Eissenberg, and W.E. Goldman. 2007. Histoplasma capsulatum alpha-(1,3)-glucan blocks recognition by the macrophage beta-glucan receptor. Proceedings of the National Academy of Sciences U.S.A. 104:1366-1370.

Lathem, W.W., P.A. Price, V.L. Miller, and W.E. Goldman. 2007. A plasminogen-activating protease specifically controls the development of primary pneumonic plague. Science 26:509-513.

Rappleye, C.A., and W.E. Goldman. 2008. Fungal stealth technology. Trends in Immunology 29:18-24.

Beck, M.R., G.T. DeKoster, D.M. Hambly, M.L. Gross, D.P. Cistola, and W.E. Goldman. 2008. Structural features responsible for biological stability of Histoplasma’s virulence factor CBP. Biochemistry 47:4427-4438.

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Accomplishments

Honors and Leadership Activities
Burroughs Wellcome Fund Scholar Award in Molecular Pathogenic Mycology 1996 - 2001
Director, Washington University Graduate Program in Molecular Microbiology 1998 - 2007
Chair of FASEB Research Conference on Microbial Pathogenesis 2000
Chair of Gordon Research Conference on Microbial Toxins and Pathogenicity 2002
Fellow of the American Academy of Microbiology (AAM) 2002 - present

Other Professional Activities
Editorial Board, Infection & Immunity 1987 - 2001
Reviewer, NIH Biological Sciences Study Section, Subcommittee 3 1990 - 1994
Ad hoc reviewer, NIH Bacteriology and Mycology Study Section, Subcommittees 1 & 2 1991 - 2002
Editorial Boards: Cell. Microbiol., Curr. Opin. Microbiol., Trends Microbiol. 1997 - present
Ad hoc member, NIAID Board of Scientific Counselors 1998
Editor, Molecular Microbiology 2001 - 2008
"Faculty of 1000" Section Co-Head (Cellular Microbiology and Pathogenesis) 2001 - present
AAM Colloquia on new directions/issues in microbiology and genomics 2003 - 2004
ASM Meetings & Conferences Committee 2004 - present
ASM-NIH Workshop on Basic Bacterial Research 2005
Advisory Committee, Burroughs Wellcome Fund 2005 - 2008
Program Committee, ASM Biodefense and Emerging Diseases Research Meeting 2007 - present