Abstract

This proposal represents a continuation of our investigations into the structure, function and physiological importance of perforin. Perforin is a component of the cytolytic mechanism of cytotoxic T cells (CTL) and natural killer (NK) cells. It is contained in cytoplasmic granules that are secreted during the cytolytic mechanism. Although it has been established that perforin and granules can play an important in cytolysis, it is still controversial whether perforin is essential for killing or even represents a major cytolytic component. Research in this area has been impeded previously by the unavailability of sensitive assays for perforin. Our success in the isolation and analysis of cDNA clones for both murine and human perforin opens new avenues into the exploration of the crucially important effector mechanisms of CTL and NK. Accordingly, this continuation application will focus on several new areas of research intimately related to perforin and cytotoxicity. The genomic structure of perforin will be investigated and compared to C9. The regulation of the induced expression of perforin will be studied and transcriptional or translational control mechanisms uncovered. The cell biology of the expression of perforin by transfection will be analyzed. In particular, the influence of granule factors on stability and cytotoxicity of perforin will be investigated. The requirement for a secretory apparatus for the mediation of cytotoxicity in transfected cells will be determined, In anti-sense transfection experiments, it will be determined whether perforin is essential for the cytolysis of target cells. In addition to its cytotoxicity, factors that protect cells from cytotoxicity by perforin (perforin inhibitor) will be isolated and analyzed. The role of these factors in self protection of CTL and NK will be investigated. The perofrin inhibitor will be purified and characterized. Its structure will be determined by obtaining and isolating cDNA clones from a human LAK cDNA library. Cumulatively, these studies will allow an unambiguous assessment of perforin's role in cell mediated cytotoxicity and the role of perforin inhibitors in killer cell self protection.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA039201-08
Application #
3177962
Study Section
Experimental Immunology Study Section (EI)
Project Start
1988-05-15
Project End
1994-03-31
Budget Start
1991-04-01
Budget End
1992-03-31
Support Year
8
Fiscal Year
1991
Total Cost
Indirect Cost

  Institution

Name
University of Miami School of Medicine
Department
Type
Schools of Medicine
DUNS #
City
Miami
State
FL
Country
United States
Zip Code
33146

  Publications

McCormack, Ryan M; Lyapichev, Kirill; Olsson, Melissa L et al. (2015) Enteric pathogens deploy cell cycle inhibiting factors to block the bactericidal activity of Perforin-2. Elife 4:
McCormack, Ryan M; de Armas, Lesley R; Shiratsuchi, Motoaki et al. (2015) Perforin-2 is essential for intracellular defense of parenchymal cells and phagocytes against pathogenic bacteria. Elife 4:
McCormack, Ryan; de Armas, Lesley R; Shiratsuchi, Motoaki et al. (2013) Inhibition of intracellular bacterial replication in fibroblasts is dependent on the perforin-like protein (perforin-2) encoded by macrophage-expressed gene 1. J Innate Immun 5:185-94
Fields, K A; McCormack, R; de Armas, L R et al. (2013) Perforin-2 restricts growth of Chlamydia trachomatis in macrophages. Infect Immun 81:3045-54
McCormack, Ryan; de Armas, Lesley; Shiratsuchi, Motoaki et al. (2013) Killing machines: three pore-forming proteins of the immune system. Immunol Res 57:268-78
Fang, Lei; Adkins, Becky; Deyev, Vadim et al. (2008) Essential role of TNF receptor superfamily 25 (TNFRSF25) in the development of allergic lung inflammation. J Exp Med 205:1037-48
Xiao, Yanping; Motomura, Seiichi; Podack, Eckhard R (2008) APRIL (TNFSF13) regulates collagen-induced arthritis, IL-17 production and Th2 response. Eur J Immunol 38:3450-8
Oizumi, Satoshi; Deyev, Vadim; Yamazaki, Koichi et al. (2008) Surmounting tumor-induced immune suppression by frequent vaccination or immunization in the absence of B cells. J Immunother 31:394-401
Podack, Eckhard R; Deyev, Vadim; Shiratsuchi, Motoaki (2007) Pore formers of the immune system. Adv Exp Med Biol 598:325-41
Oizumi, Satoshi; Strbo, Natasa; Pahwa, Savita et al. (2007) Molecular and cellular requirements for enhanced antigen cross-presentation to CD8 cytotoxic T lymphocytes. J Immunol 179:2310-7

Showing the most recent 10 out of 62 publications