Progress 02/22/90 to 12/31/03
Outputs
By using chemically synthesized peptides and testing them in cell culture and in animal model systems, the expression and modulation of a number of diferent cytokines was studied, which was further utilized in developing therapies for different types of cancer and for viral infections. Specifically, different therapetic peptides for the treatment of smallpox, melanoma, prostate cancer, and experimental allergic encephalomyelitis (an animal model for multiple sclerosis) were developed. A peptide from interferon gamma, spanning the residues 95 to 132 was found to be a very effective antiviral peptide. It was active against smallpox family of viruses, where the intact host interferon gamma was rendered inactive. Furthermore, this peptide was able to thwart the viral spread even at late stage of infection, and was active given orally, which would aid further in developing therapeutics. A tyrosine kinase inhibitor (Tkip) peptide was effective at immunomodulation needed to
suppress the symptoms of multiple sclerosis in mouse models. In summary, our approaches have been successful at developing a number of useful therapies.
Impacts
Smallpox virus is one of the biological warfare agents that we need to be prepared against. In addition, this virus has the capacity to re-emerge in different forms. We have developed therapeutic peptides that are very effective at thwarting the spread of smallpox family of viruses, even at late stages of infection, and they are effective given orally, which would aid in developing effective therapeutics from these. A tyrosine kinase inhibitor peptide developed is able to rescue mice from severe paralysis of multiple sclerosis.
Publications
- Ahmed CMI, Burkhart MA, Subramaniam PS, Mujtaba MG, and Johnson HM (2005). Peptide mimetics of gamma interferon possess antiviral properties against vaccii avirus and other viruses in the presence of poxvirus B8R protein. J. Virol 79: 5632-5639.
- Ahmed CMI, Martin JP, and Johnson HM. (2007). IFN mimetic as a therapeutic for lethal vaccinia virus infection: Possible effects on innate and adaptive immune responses. Submitted.
- Mujtaba MG, Flowers LO, Patel CB, Patel RA, Haider MI, and Johnson HM (2005). Treatment of mice with suppressor of cytokine signaling mimetic peptide, tyrosine kinase inhibitor peptide, prevents development of the acute form of experimental allergic encephalomyelitis and induces stable remission in the chronic relapsing/remitting form. J. Immunol 175: 5077-5086.
- Mujtaba MG, Patel CB, Patel RA, Flowers LO, Burkhart MA. Waiboci, LW, Martin J, Haider MI, Ahmed CM, and Johnson HM (2006). The gamma interferon mimetic peptide IFN(95-132) prevents encephalomyocarditis virus infection both in tissue culture and in mice. Clin Vaccine Immunol 13: 944-952.
- Ahmed CMI, Burkhart MA, Mujtaba MG, Subramaniam PS, and Johnson HM (2003). The role of IFN nuclear localization sequence in intracellular function. J Cell Sci 116: 3089-3098.
- Ahmed CMI, and Johnson HM (2006). IFNg and its receptor subunit IFNGR1 are recruited to the IFNg-activated sequence element at the promoter site of IFNg-activated genes. J. Immunol. 177: 315-321.
- Johnson HM, Subramaniam PS, Olsnes S, and Jans DA (2004). Trafficking and signaling pathways of nuclear localizing protein ligands and their receptors. BioEssays 26: 993-1004.
- Flowers LO, Johnson HM, Mujtaba MG, Ellis MR, Haider Sm, and Subramaniam PS (2004). Characterization of a peptide inhibitor of Janus kinase 2 that mimics suppressor of cytokine signaling-1 function. J. Immunol. 172: 7510-7518.
- Mujtaba MG, Soos J, and Johnson HM (1997). CD4 T cells mediate protection against experimental allergic encephalomyelitis. J. Neuroimmunol 75: 35-42.
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Progress 10/01/02 to 10/01/03
Outputs
We are currently studying a mimetic peptide of IFN-gamma (IFNg)for therapeutics of cancer and for smallpox and other respiratory viral infection. The C-terminus of IFNg contains a nuclear localization sequence (NLS) required for the activation and nuclear translocation of the transcription factor STAT1-alpha and induction of IFNg-activated genes. Based on this and other studies, we developed a peptide mimetic of IFNg that possesses the IFNg function of antiviral activity and upregulation of MHC class II molecules. The mimetic also shares with IFNg the ability to induce the activation and nuclear translocation of STAT1-alpha and the IFNGR-1 subunit. The mimetic, IFNg(95-132), is a peptide that consists of the C-terminal residues 95-132 of murine IFNg, and contains a required alpha helical domain and the NLS of IFNg. In this project we determined the mechanism of the intracellular action at the level of signal transduction. We showed that the mimetic mediates the
nuclear transport of IFNGR-1 through its interaction with IFNGR-1 cytoplasmic region 253-287 via both the helical region and the NLS of IFNg(95-132). Alanine substitutions of the NLS of the mimetic showed that the NLS was required for nuclear translocation, and that the nuclear transport properties of the mimetic correlated with its ability to bind IFNGR-1. These data further show that the NLS of IFNg(95-132) can interact simultaneously with IFNGR-1 and the nuclear import machinery. We found that in in-vitro nuclear transport assays tyrosine phosphorylated STAT1 alpha failed to undergo nuclear translocation in the presence of nuclear import factors, but was transported to nucleus in the presence of IFNg(95-132) and JAK2-rphosphorylated IFNGR-1, to which STAT1-alpha binds, as a complex of IFNg(95-132)/IFNGR-1/STAT1-alpha. Thus, the mimetic, which possesses IFNg function, is directly involved as a chaperone in the nuclear transport of STAT1-alpha, and shares this mechanism of action
with that previously desvrived for IFNg. The mimetic, like IFNGg, is able to upregulate the tumor suppressor p21WAF1/CIP1, a direct target of STAT1-alpha, and this ability requires the NLS of the mimetic. However, unlike IFNg, the mimetic is unable to downregulate c-myc and hence does not inhibit the cycling of cells. This suggests that IFNg has additional functions that are not tied directly to the nuclear translocation of STAT1-alpha.
Impacts
These studies will help define immunotherapeutic approaches for the treatment of cancer and viral infections, including smallpox and other upper respiratory viral infections.
Publications
- Torres BA, Perrin GQ, Mujtaba MG, Subramaniam PS, Anderson A, Johnson HM. Superantigen enhancement of specific immunity: antibody production and signaling pathways. Journal of Immunology 2002; 169(6):2907-14.
- Soos JM, Mujtaba MG, Schiffenbauer J, Torres BA, and Johnson HM. Intramolecular epitope spreading induced by staphylococcal enterotoxin superantigen reactivation of experimental allergic encephalomyelitis. Journal of Neuoimmunology 2002; 123:30-34.
- Ahmed CMI, Burkhart MA, Mujtaba MG, Subramaniam PS, Johnson HM. The role of IFN gamma nuclear localization sequence in intracellular function. J cell Science 2003: 116:8089-8098.
- Ahmed CMI and Johnson HM. Interferon gene therapy for the treatment of cancer and viral infections. Drugs of Today 2003: 39, 763-766.
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Progress 10/01/01 to 10/01/02
Outputs
Gamma Interferon: We have previously shown that gamma interferon (IFN-g) localizes to the nucleus via a nuclear localization sequence (NLS). Further, this NLS causes the localization of receptor alpha chain as well as associated transcriptions factors (STATs). We are currently studying this NLS peptide (region 95-133) as being a mimetic/agonist for interferon gamma in vivo in mice and in tissue culture. Also, we have expressed an intracellular non-secreted form of IFN-g in order to further define the intracellular events involved in gamma interferon action. This interferon will allow us to define events that occur upon binding of IFN-g to the intracellular domain of the receptor. Tau interferon: IFN-tau, a novel type I IFN that possesses immunomodulatory properties, lacks toxicity normally associated with other type I IFNs. We have shown that oral administration of IFN-tau promotes a Th2 bias and enhances suppression of autoimmune encephalomyelitis by oral glatiramer
acetate. Furthermore, we have found that superantigen enhancement of specific antibody produced is inhibited by IFN-tau induced cytokines. Finally, IFN-tau expressed intracellularly in a nonsecreted form possesses similar biological and signaling activity as extracellularly added IFN-tau. Superantigens: Recently, we showed that superantigens (SAgs) can protect mice against lethal melanoma using superantigens, likely due to expansion of tumor-specific cytolytic T cells. We have now shown that SAg can enhance the humoral (antibody) responses against soluble antigens such as bovine serum albumin (BSA). As in the melanoma studies, SAgs were found to significantly enhance humoral responses. Antibody titers climbed rapidly with SAgs, and antibodies were produced in greater quantities and titers were slower to decline than in the absence of SAgs. Further, interleukin 10, which has been shown by us to block T cell activation by superantigens, blocked SEB-induced phosphorylation of the MAP
kinase protein, ERK and blocked the perinuclear localization of another MAP kinase protein, MNK, which is an important component of the transcription complex. We now are currently studying superantigen mimetics and the cytokine profile that different superantigens elicit via doing microarray experiments.
Impacts
These studies will help define immunotherapeutic approaches for the treatment of cancer, allergies, viral infection, and autoimmune diseases in humans and animals.
Publications
- Subramaniam, P.S., Torres, B.A., and Johnson H.M. (2001). So many ligands, so few transcription factors: A new paradigm for STAT signaling. Cytokine, 15:175-187.
- Soos JM, Stuve O., Johnson HM, Weiner HL, Zamvil SS. (2002). Oral type I IFN - tau promotes Tr1 bias and enhances suppression of encephalomyelitis by oral glatiramer acetate. J. Immunol. 169(5)2231-5.
- Subramanaim, PS and Johnson HM. (2002). Lipid microdomains are required sites for the selective endocytosis and nuclear translocation of IFN gamma, its receptor cleave, IFNGR-1, and the phosphorylation and nuclear translocation of STAT1alpha. J. Immunol. 169(4):1959-69.
- Subramaniam PS, Mujtaba MG, Johnson HM. 2002. In vitro nuclear translocation of STAT1a occurs via an NLS in IFNg. (Submitted).
- Ahmed CMI, Burkhart MA, Subramaniam PS, Johnson HM. 2002. Intracellular expression of IFNg: Role of a nuclear localization sequence and IFNGR1 subunit endocytosis from within in STAT1 activation and nuclear translocation. (Submitted).
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Progress 10/01/00 to 10/01/01
Outputs
Gamma Interferon: We have shown that gamma interferon (IFN-g) localizes to the nucleus via a polybasic region that functions as a nuclear localization sequence (NLS). Further, this NLS causes the localization of receptor alpha chain as well as associated transcriptions factors (STATs). We are currently studying mutant IFN-g receptor molecules that have the extracellular domain of eryrthopoeitin receptor and the intracellular domain of IFN-g receptor, and vice versa. Also we are pursuing studies on the nature of the endosomal compartments that are involved in gamma interferon receptor internalization. Finally, as another approach to defining the intracellular events involved in gamma interferon action, we have expressed an intracellular non-secreted form of IFN-g. This interferon will allow us to define events that occur upon binding of IFN-g to the intracellular domain of the receptor, namely STAT activation and nuclear localization. Tau interferon: A chimeric
interferon was produced that has the first (1-27) amino acids of ovine IFN-t and the 28-166 of human interferon alpha D. The rationale for producing this interferon was that ovine IFN-t is significantly less toxic than human IFNaD, and the region of tau involved in lack of toxicity is the first 27 amino acids. This chimeric was tested in mice and was shown to have similar activity as native IFN-t in preventing autoimmune disease. Also, like the IFN-g studies mentioned above, a non-secreted form of IFN-t has been produced. This interferon will help further in the understanding of the lack of toxicity of IFN-t. Superantigens: We previously showed that superantigens (SAgs) can protect mice against lethal melanoma using superantigens, likely due to expansion of tumor-specific cytolytic T cells. We have looked into the effects of SAgs on antibody responses to soluble antigens, both T-dependent and T-independent. As in the melanoma studies, SAgs were found to significantly enhance humoral
responses. Antibody titers climbed rapidly with SAgs, and antibodies were produced in greater quantities and titers were slower to decline than in the absence of SAgs. Further, antibody titers to Type I T-independent antigens such as the pneumococcal polysaccharides were also positively affected by SAgs whereas no effect was seen on antibody production to Type II T-independent antigens such as lipopolysaccharide. The intracellular events associated with SAg activation of CD4+ T cells was further studied. SEB, a staphylococcal enterotoxin SAg, was shown to activate the mitogen arm of the MAP kinase signaling pathway, whereas T cell activators such as the mitogen concanavalin A function via the p38 stress component of the MAP kinase pathway. Further, interleukin 10, which has been shown by us to block T cell activation by superantigens, blocked SEB-induced phosphorylation of the MAP kinase protein, ERK and blocked the perinuclear localization of another MAP kinase protein, MNK, which is
an important component of the transcription complex.
Impacts
These studies will help further define immunotherapeutic apporaches for the treatment of cancer, allergies, viral infections, and autoimmune dieases in animals and humans.
Publications
- Subramaniam PS, Green MM, Larkin J 3rd, Torres BA, Johnson HM. 2001. Nuclear Translocation of IFN-gamma Is an Intrinsic Requirement for Its Biologic Activity and Can Be Driven by a Heterologous Nuclear Localization Sequence. J Interferon Cytokine Res 21:951-959.
- Kominsky SL, Torres BA, Hobeika AC, Lake FA, Johnson HM. 2001. Superantigen enhanced protection against a weak tumor-specific melanoma antigen: Implications for prophylactic vaccination against cancer. Int J Cancer 94:834-841.
- Subramaniam PS, Torres BA, Johnson HM. 2001. So many ligands, so few transcription factors: a new paradigm for signaling through the stat transcription factors. Cytokine 15:175-187.
- Larkin J 3rd, Subramaniam PS, Torres BA, Johnson HM. 2001. Differential properties of two putative nuclear localization sequences found in the carboxyl-terminus of human IFN-gamma. J Interferon Cytokine Res 21:341-348.
- Torres BA, Kominsky S, Perrin GQ, Hobeika AC, Johnson HM. 2001. Superantigens: the good, the bad, and the ugly. Exp Biol Med 226:164-176.
- Subramaniam PS, Green MM, Larkin III J, Torres BA, Johnson HM. 2001. Nuclear translocation of interferon-gamma is an intrinsic requirment for its biological activity and can be driven by a heterologous nuclear localization sequence. J Interferon Cytokine Res 21:951-959.
- Soos JM, Mujtaba MG, Schiffenbauer J, Torres BA, Johnson HM. 2001. Intramolecular epitope spreading induced by staphylococcal enterotoxin superantigen reactivation of experimental allergic encephalomyelitis. J Neuroimmunol, In Press.
- Torres BA, Perrin GQ, Mujtaba MG, Subramaniam PS, Johnson HM. 2001. Superantigen enhancement of specific immunity: antibody production and signaling pathways. Submitted.
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Progress 10/01/99 to 09/30/00
Outputs
Gamma Interferon: We recently showed that the nuclear localization of IFN gamma is mediated by a polybasic nuclear localization sequence (NLS) in its C terminus. This NLS is required for the full expression of biological activity of IFN gamma, both extracellularly and intracellularly. We now show that this NLS plays an integral intracellular role in the nuclear translocation of the transcription factor STAT1 alpha activated by IFN gamma. The ligand and STAT1 translocate to the nucleus along with the receptor alpha chain, which acts as a `scaffolding' for these proteins. The beta chain remains on the surface of the cell. The intracellular vesicles for nuclear localization of ligand/receptor/STAT are caveolae, a unique form of endosome. Tau Interferon: We have successully produced and purified large quantities of the chimeric IFNtau(1-27)/IFNalpha(28-166). The chimeric is highly purified and is now ready for x-ray crystallography studies and NMR solution structure
studies. The chimeric will be tested in an animal model for multiple sclerosis. Superantigens: We are continuing studies on amplification of immune responses by superantigens. We previously reported on the ability of SEA/SEB to induce protective anti-tumor responses against melanoma. We further studied the effects of superantigens on humoral immune responses to soluble antigens such as bovine serum albumin (BSA). Mice given superantigens produced higher levels of anti-BSA antibodies as well production occurring more rapidly and more prolonged. Thus, superantigens may be useful in augmenting antibody responses against weak vaccines. Superantigens bind to the T cell receptor (TCR) and induce T cells to proliferate and become activated. This occurs via TCR-linked signal transduction pathways such as the MAP kinase pathway. Proliferation induced by superantigens can be blocked or inhibited by a TH2 cytokine, interleukine-10 (IL-10). We have been investigating the site(s) in the MAP kinase
pathway at which IL-10 exerts its inhibitory effects.
Impacts
These studies will help define immunotherapeutic approaches for the treatment of cancer, allergies, viral infections, and autoimmune diseases in humans and animals.
Publications
- Kominsky SL, Hobeika AC, Lake FA, Torres BA, Johnson HM. (2000). Downregulation of neu/HER-2 by interferon gamma in prostate cancer cells. Cancer Res 60:3904-3908.
- Tanabe T, Kominsky SL, Subramaniam PS, Johnson HM, Torres BA. 2000. Cell cycle inhibition by type I IFNs occurs at both the G1 and S phase and correlates with upregulation of p21WAF1. J Neuro-Oncol 48:225-232.
- Larkin III J, Subramaniam PS, Torres BA, Johnson HM. 2000. Differential properties of two putative nuclear localization sequences found in the carboxyl terminus of human IFN gamma. J Interferon Cytokine Res, In Press.
- Kominsky SL, Torres BA, Hobeika AC, Lake FA, Johnson HM. 2000. Vaccination with irradiated tumor cells by superantigen administration elicits a potent and protective anti-tumor immune response in C57Bl/6 mice. Submitted.
- Torres BA, Kominsky SL, Johnson HM. 2000. Superantigens can significantly enhance antibody production to soluble antigens: a method to augment immune responses to weak vaccines. Submitted.
- Perrin GQ, Subramaniam PS, Johnson HM. 2000. Interleukin 10 suppresses activation of the Erk MAP kinase pathway, inhibits Erk nuclear localization and inhibits Mnk perinuclear localization in SEB-stimulated human CD4+ T cells. Submitted.
- Torres BA, Soos JM, Perrin GQ, Johnson HM. (2000). Microbial superantigens and immunological deregulation. In Persistent Bacterial Infections. (Cunningham-Ruddles S, ed), ASM Press, Washington, D.C., p. 183-197.
- Kominsky SL, Torres BA, Subramaniam PS, Johnson HM. (2000). Inhibitory effects of IFN gamma and acyclovir on the glioblastoma cell cycle. J Interferon Cytokine Res, 20:463-469.
- Larkin III J, Johnson HM, Subramaniam PS. (2000). Differential endocytosis and nuclear translocation of alpha and beta subunits of IFN gamma activated receptor. J Interferon Cytokine Res, 20:565-576.
- Subramaniam PS, Larkin III J, Johnson HM. (2000). The COOH terminal nuclear localization sequence of interferon gamma regulates STAT1alpha nuclear translocation at an intracellular site. J Cell Sci 113:2771-2781.
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Progress 10/01/98 to 09/30/99
Outputs
Gamma Interferon: It is known that the transcription factors, STATs, do not possess nuclear localization sequences, even though they must get into the nucleus in order to function. We have shown that the C-terminal domain of the gamma interferon (gamma IFN) molecule possesses nuclear localization activity via a nuclear localization sequence (NLS). A C-terminal peptide, IFN(95-133), is sufficient for function, including the nuclear localization activity. We have shown that IFN(95-133) co-localizes to the nucleus along with the STATs and the alpha chain of the gamma IFN receptor. We hypothesize that gamma IFN acts as a nuclear chaperone for molecules such as the gamma IFN receptor alpha chain and the STATs, which require nuclear localization and yet do not have an NLS. Along these lines, we have shown that the NLS of gamma IFN can be replaced with the NLS of the SV40 T antigen, the prototypic NLS. In cell cycle studies on glioblastomas, we have shown that suboptimal
doses of gamma IFN and acyclovir, a nucleoside analog, block growth of these cancer cells. Further, gamma IFN was shown to induce the production of the tumor suppressor, p21, which acts in part by binding to PCNA, an integral member of the transcriptional machinery. Tau Interferon: We have shown that tau IFN blocks the induction of atopic allergy in animal studies, and blocks the production of IgE from myeloma cells. In structure/function studies, we were able to successfully label tau IFN with 15N for future NMR studies. In cell cycle studies, we have shown that type I IFNs blocks glioblastoma cell growth by the upregulation of the tumor suppressor, p21. We are commencing studies on nuclear translocation of tau IFN and other type I IFNs. We are starting studies on the effects of tau IFN on the MAP kinase pathway. Superantigens: We are currently finishing work on modulation of anti-tumor immunity by superantigens in a melanoma animal model. Briefly, mice are vaccinated with inactive
tumor cells and, one week later, are given a dose of superantigens (SEA/SEB). Then mice are challenged with a lethal dose of tumor cells, which normally causes death within two weeks. We have had a significant number of mice survive as long as 150 days after tumor challenge. This project is the subject of a patent application by UF.
Impacts
These studies will help define immune-based approaches for the treatment of human and animal diseases such as cancer, allergies, viral infections, and autoimmune diseases.
Publications
- Kominsky SL, Bryan G, Hobeika AC, Tanabe T, Subramaniam PS, Johnson HM, Torres BA. (1998). IFNg inhibition of cell growth in glioblastomas is accompanied by induction of the cyclin-dependent kinase inhibitor, p21. Oncogene, 17:2973-2979.
- Perrin GQ, Johnson HM, Subramaniam PS. (1999). Mechanism of IL-10 inhibition of T-helper cell activation by superantigens at the level of the cell cycle. Blood, 93:208-219.
- Subramaniam PS, Mujtaba MG, Paddy MR, Johnson HM. (1999). The C-terminus of interferon-g contains a functional polybasic nuclear localization sequence. J Biol Chem, 274:403-407.
- Soos JM, Johnson HM. (1999). Interferon-t: Prospects for clinical use in autoimmune disorders. BioDrugs 11:125-135.
- Johnson TM, Holaday SK, Sun Y, Subramaniam PS, Johnson HM. (1999). Expression, purification, and characterization of functionally active 15N-labeled interferon-t in Pichia pastoris. J Interferon Cytokine Res, 19:631-636.
- Mujtaba MG and Johnson HM. (1999). IFNt inhibits IgE production in a murine model of allergy and in a IgE-producing human myeloma cell line. J Allergy Clin Immunol. 104:1037-1044.
- Johnson HM, Torres BA, Subramaniam PS. (1999). Interferons. In Encyclopedia of Life Sciences. MacMillan Reference Ltd, In Press.
- Torres BA, Soos JM, Perrin GQ, Johnson HM. (1999). Microbial superantigens and immunological deregulation. In Persistent Bacterial Infections. (Cunningham-Ruddles S, ed), ASM Press, In Press.
- Soos JM, Mujtaba MG, Schiffenbauer, Johnson HM. (1999). Intramolecular epitope spreading induced by staphylococcal enterotoxin superantigen reactivation of experimental allergic encephalomyelitis. J Neuroimmunol, In Press.
- Subramaniam PS, Larkin III J, Johnson HM. (1999). The COOH terminal nuclear localization sequence of interferon g regulates STAT1a nuclear translocation at an intracellular site. Submitted.
- Larkin III J, Johnson HM, Subramaniam PS. (1999). Differential endocytosis and nuclear translocation of a and b subunits of IFNg activated receptor. Submitted.
- Kominsky SL, Torres BA, Subramaniam PS, Johnson HM. (1999). Inhibitory effects of IFNg and acyclovir on the glioblastoma cell cycle. Submitted.
- Kominsky SL, Hobeika AC, Lake FA, Torres BA, Johnson HM. (1999). Downregulation of neu/HER-2 by interferon g in prostate cancer cells. Submitted.
- Tanabe T, Kominsky SL, Subramaniam PS, Johnson HM, Torres BA. (1999) Cell cycle inhibition by type I IFNs occurs ar both the G1 and S phase and correlates with upregulation of p21WAF1. Submitted.
- Hobeika AC, Etienne W, Torres BA, Johnson HM, Subramaniam PS. (1999). IFNg induction of p21WAF1 is required for cell cycle inhibition and suppression of apoptosis. J Interferon Cytokine Res, In Press.
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Progress 10/01/97 to 09/30/98
Outputs
Gamma Interferon: We have shown that the C-terminal domain of gamma interferon (gamma IFN) has a nuclear localization sequence that targets the cytokine to the nucleus, and shuttles important transcription factors along with it, thereby playing an important role in signal transduction. We have been successful in expression of the human gamma IFN receptor cytoplasmic domain, and have characterized the binding of human gamma IFN to this domain of the receptor. We are in the process of substituting the nuclear localization sequence of human gamma IFN with that of SV40 T antigen. We have examined the antitumor potential of gamma IFN in human prostate cancer cells, glioblastoma cells and breast cancer cells. Gamma IFN induces the expression of p21WAF1 in prostate cancer cells, which causes alteration in the phenotype and invasive potential of the cells. Further cell cycle studies show that gamma IFN inhibits the cell growth of several clinically-isolated glioblastomas and
inhibition is accompanied by induction of the cyclin-dependent kinase inhibitor, p21. We are looking onto the possible synergistic effects of gamma IFN with the nucleoside analog, acyclovir. We are in the process of expressing the beta chain of the gamma IFN receptor complex and this protein will help us determine the interaction of gamma IFN with the entire receptor complex. Further, we have made peptides encompassing the entire length of the beta chain and antibodies to these peptides have been generated. These reagents will be used to determine the role of the beta chain in the activity of gamma IFN on human cells. Tau Interferon: As far as autoimmune disease, we have shown that tau interferon (tau IFN) suppresses both the autoreactive humoral and cellular immune responses and induces stable remission in mice with chronic experimental allergic encephalomyelitis, the animal model for multiple sclerosis. Work we have performed on the anticancer activity of that tau IFN and other type
I IFNs shows that induction of the tumor suppressor gene product, p21WAF1, in Daudi cells is accompanied by ordered G1 arrest, differentiation, and apoptosis. Further, differential induction of apoptosis was found by the type I IFNs. We have also shown that cell cycle inhibition by type I IFNs occurs at both the G1 and S phase and correlates with upregulation of p21WAF1. We are beginning studies on the nuclear translocation of that tau IFN akin to the studies described above for gamma IFN. Further, we are characterizing the binding of tau IFN to the type I IFN receptor using peptides encompassing both the extracellular and cytoplasmic domains of the beta chain of the type I IFN receptor. Superantigens: We have developed a novel method for detecting Vbeta T cell expansion by weak superantigens. This has enabled us to further characterize the superantigen activity of an HIV protein, Nef. Studies are being carried out on the ability of antisera from HIV-infected patients to neutralize
Nef activity in vitro. We are also in the process of characterizing superantigens from bacterial sources that may be involved in human periodontal disease and in neuropsychiatric disorders.
Impacts
(N/A)
Publications
- Schiffenbauer, J., Soos, J.M., and Johnson, H,M. 1997. The role of superantigens in the pathogenesis of autoimmune disorders. Immunol. Today 19:117-120.
- Khan, O.A., Jiang, H., Subramaniam, P.S., Johnson, H.M., and Dhib-Jalbut, S.S. 1998. Immunomodulating functions of recombinant ovine interferon tau: potential for therapy in multiple sclerosis and autoimmune disorders. Mult. Scler. 4:63-69.
- Subramaniam, P.S., Cruz, P.E., Hobeika, A.C., and Johnson HM. 1998. Type I interferon induction of Cdk-inhibitor p21WAF1 is accompanied by ordered G1 arrest, differentiation, and apoptosis of the Daudi B-cell line. Oncogene 16:1885-1890.
- Green, M.M., Larkin, J. 3rd, Subramaniam, P.S., Szente, B.E., and Johnson, H.M. 1998. Human IFNgamma receptor cytoplasmic domain: expression and interaction with HuIFNgamma. Biochem. Biophys. Res. Comm.243:170-176.
- Johnson, H.M., Torres, B.A., Green, M.M., Szente, B.E., Siler, K.I., Larkin, J. 3rd, and Subramaniam, P.S. 1998. Cytokine-receptor complexes as chaperones for nuclear translocation of signal transducers. Biochem. Biophys. Res. Comm. 244:607-614.
- Hobeika, A.C., Etienne, W., Cruz, P.E., Subramaniam, P.S., and Johnson, H.M. 1998. IFNgamma induction of p21WAF1 in prostate cancer cells: role in cell cycle and alteration of phenotype and invasive potential. Int. J. Cancer 77:138-145.
- Johnson, H.M., Torres, B.A., Green, M.M., Szente, B.E., Siler, K.I., Larkin, J. 3rd, and Subramaniam, P.S. 1998. Hypothesis: ligand/receptor-assisted nuclear translocation of STATs. Proc. Soc. Exp. Biol. Med. 218:149-155.
- Mujtaba, M.G., Streit, W.J., and Johnson, H.M. 1998. IFNtau suppresses both the autoreactive humoral and cellular immune responses and induces stable remission in chronic experimental allergic encephalomyelitis. Cell. Immunol. 186:94-102.
- Torres, B.A, and Johnson, H.M. 1998. Modulation of disease by superantigens. Current Opin. Immunol. 10:465-470.
- Torres, B.A., Tanabem T., Subramaniamm P.S., Yamamotom J.K., and Johnson, H.M. 1998. Mechanism of HIV pathogenesis: role of superantigens in disease. Alcoholism: Clin. Exp. Res. 22:188S-192S.
- Kominsky, S.L., Bryan, G., Hobeika, A.C., Tanabe, T., Subramaniam, P.S., Johnson, H.M., and Torres, B.A. 1998. IFNgamma inhibition of cell growth in glioblastomas is accompanied by induction of the cyclin-dependent kinase inhibitor, p21WAF1. Oncogene, In Press.
- Perrin, G.Q., Johnson, H.M., and Subramaniam, P.S. 1998. Mechanism of IL-10 inhibition of T-helper cell activation by superantigens at the level of the cell cycle. Blood, In Press.
- Subramaniam, P.S., Mujtaba, M.G., Paddy, M.R., and Johnson, H.M. 1998. The C-terminus of interferon-gamma contains a functional polybasic nuclear localization sequence. J. Biol. Chem., In Press.
- Hobeika, A.C., Etienne, W., Johnson, H.M., and Subramaniam, P.S. 1998. Induction of p21WAF1 is required for cell cycle inhibition and suppression of apoptosis. Submitted.
- Tanabe, T., Kominsky, S.L., Subramaniam, P.S., Johnson, H.M., and Torres, B.A. 1998. Cell cycle inhibition by type I IFNs occurs at both the G1 and S phase and correlates with upregulation of p21WAF1. Manuscript in preparation.
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Progress 10/01/96 to 09/30/97
Outputs
IFN : We have shown that the C-terminal domain of IFN has a nuclear localization sequence that targets the cytokine to the nucleus, and shuttles important transcription factors along with it, thereby playing an important role in signal transduction. We have been successful in expression of the human IFN receptor cytoplasmic domain, and have characterized the binding of huIFN to this domain of the receptor. We have also examined the antitumor potential of IFN in human prostate cancer cells. IFN induces the expression of p21WAF1 in prostate cancer cells, which causes alteration in the phenotype and invasive potential of the cells. Further cell cycle studies show that IFN inhibits the cell growth of several clinically-isolated glioblastomas and inhibition is accompanied by induction of the cyclin-dependent kinase inhibitor, p21. IFN#: As far as autoimmune disease, we have shown that IFN# suppresses both the autoreactive humoral and cellular immune responses and induces
stable remission in mice with chronic experimental allergic encephalomyelitis, the animal model for multiple sclerosis. Work we have performed on the anticancer activity of IFN shows that induction of the tumor suppressor gene product, p21WAF1, in Daudi cells is accompanied by ordered G1 arrest, differentiation, and apoptosis. Further, differential induction of apoptosis was found by the type I interferons. We have also shown that cell cycle inhibition by type I IFNs occurs at both the G1 and S phase and correlates with upregulation of p21WAF1. Superantigens: We have developed a novel method for detecting V T cell expansion by weak superantigens. This has enabled us to further characterize the superantigen activity of an HIV protein, Nef.
Impacts
(N/A)
Publications
- Subramaniam PS, Johnson HM: A role for cyclin-dependent kinase inhibitor p21 in the G1 cell cycle arrest mediated by type I interferons. J Interfeon Cytokine Res 1997, 17:11-15.
- Schiffenbauer J, Johnson HM, Soos JM: Superantigens in autoimmunity: Their role as etiologic and therapeutic agents. In Superantigens: Molecular Biology, Immunology, and Relevance to Human Disease. (Leung D, Huber B, and Schlievert P, eds), Marcel Dekker, Inc., 1997.
- Hobeika AC, Subramaniam PS, Johnson HM: IFNalpha induces the expression of the cyclin- dependent kinase inhibitor p21 in human prostate cancer cells. Oncogene 1997, 14:1165-1170.
- Tanabe T, Torres BA, Subramaniam PS, Johnson HM: Broad range V activation by HIV Nef protein: Detection by a simple amplification procedure. Biochem Biophys Res Comm 1997, 230:509-513.
- Soos JM, Schiffenbauer J, Johnson HM, Pontzer CH: Interferon tau: A potential treatment for inflammatory central nervous system disease. In Interferon Therapy of Multiple Sclerosis". (Reder AT, ed) Marcel Dekker, Inc, 1997 , pp 479-498.
- Soos JM, Schiffenbauer J, Torres BA, Johnson HM: Superantigens as virulence factors in autoimmune and immunodeficiency diseases. Medical Hypothesis 1997, 48:253-259.
- Soos JM, Mujtaba MG, Subramaniam PS, Streit WJ, Johnson HM: Chronic relapsing experimental allergic encephalomyelitis prevented by oral feeding of interferon tau. J Neuroimmunol 1997, 75:43-50.
- Mujtabe MG, Soos JM, Johnson HM: CD4 T cells mediate protection against experimental allergic encephalomyelitis J Neuroimmunol 1997,
- Pontzer CH, Yamamoto JK, Bazer FW, Johnson HM: Potent anti-FIV and anti-HIV effect of interferon tau. J Immunol 1997, 158:4351-4357.
- Jarpe MA, Pontzer CH, Szente BE, Johnson HM: Structure and functional studies of interferon: a solid foundation for rational drug design. In Structure-Based Drug Design. (Veerapandian P, ed), Marcel Dekker,
- Torres BA, Johnson HM: Neuroendocrine peptide hormone regulation of immunity. From "Neuroimmunoendocrinology" volume of Progress in Allergy (Blalock JE, ed), 1997, pp 155-184.
- Jablonsky MJ, Subramaniam PS, Johnson HM, Russell JK, Krishna NR: Proton NMR assignments and solution structure of a class II MHC protein binding domain. Biochem Biophys Res Comm 1997, 234:600-665.
- Schiffenbauer J, Soos JM, Johnson HM: The role of superantigens in the pathogenesis of autoimmune disorders. Immunol. Today, 1997, In
- Subramaniam PS, Cruz PE, Hobeika AC, Johnson HM: Type I interferon induction of Cdk-inhibitor p21WAF1 is accompanied by ordered G1 arrest, differentiation, and apoptosis of the Daudi B-cell line.
- Torres BA, Tanabe T, Subramaniam PS, Yamamoto JK, Johnson HM: Mechanism of HIV pathogenesis: role of superantigens in disease. From "Proceedings of Alcohol-Induced Immunopathology", New Orleans meeting of Oct. 17-18, 1997 (Spitzer JE, ed), 1997, In Press.
- Cruz PE, Subramaniam PS, Hobeika AC, Torres BA, Johnson HM: Differential induction of apoptosis by type I interferons. 1997,
- Hobeika AC, Etienne W, Cruz PE, Subramaniam PS, Johnson HM: IFN induction of p21WAF1 in prostate cancer cells: role in cell cycle and alteration of phenotype and invasive potential. 1997, Submitted.
- Mujtaba MG, Streit WJ, Johnson HM: IFN# induces stable remission in chronic experimental allergic encephalomyelitis. 1997, Submitted.
- Johnson HM, Torres BA, Green MM, Szente BE, Siler KI, Larkin J, Subramaniam PS: Direct involvement of cytokines in JAK/STAT signal transduction. 1997, Submitted.
- Green MM, Larkin J, Subramaniam PS, Szente BE, Johnson HM: Human IFN receptor cytoplasmic domain: Expression and interaction with HuIFN .
- Kominsky SL, Torres BA, Bryan G, Hobeika AC, Tanabe T, Subramaniam PS, Johnson HM: IFN inhibition of cell growth in glioblastomas is accompanied by induction of the cyclin-dependent kinase inhibitor,
- Tanabe T, Kominsky SL, Subramaniam PS, Johnson HM, Torres BA: Cell cycle inhibition by type I IFNs occurs at both the G1 and S phase and correlates with upregulation of p21WAF1. 1997, Submitted.
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Progress 10/01/95 to 09/30/96
Outputs
IFNy. We have shown that the C-terminus of IFNy is sufficient for its antiviral and immunoregulatory properties. This has been demonstrated with the immune IFNy peptide IFNy (95-133) and with its human IFNy counterpart. The peptides must be internalized in order to be active. We have also established the structural basis of the C-terminus activity, which requires an intact helical structure as well as a polycationic region. Current studies are in progress to show the structural basis for C-terminus binding to cyto plasmic receptor. IFNY. Ovine IFN# has potential for treatment of multiple sclerosis (MS) in humans. We have shown the ovine 1FN# administered orally can protect mice against paralysis in an animal model for MS. Further, ovine 1FN# modulates human T-cell function in tissue culture. These findings will serve as the basis for phase I clinical trials by others. Additionally, we have shown that 1FN# and other type I IFNs induce the tumor suppressor gene product
p21 in cells that results in blockage of cell progress from G1 to S. Thus, we have discovered one mechanism by which IFNs inhibit cell growth, including cancer cells. Superantigens. We have shown that the Nef superantigen can induce blast transformation in primary human CD4 T-cells, and that this renders the cell susceptible to HIV replication. Antibodies to Nef protects the cells, so the Nef protein is a viable candidate for vaccination against AIDS.
Impacts
(N/A)
Publications
- Johnson, H.M., B.A. Torres, and J.M. Soos. 1996 Superantigens: Structure and relevance to human disease. Proc. Sci. Exp. Biol. Med. 212:99-109.
- Hobeika, A.C. and H.M. Johnson. 1996. A neutralizing epitope of the superantigenSEA has agonist activity on T cells. Biochem. Biophys. Res. Comm. 223:565-571.
- Torres, B.A., T. Tanabe, and H.M. Johnson. 1996. Replication of HIV-1 in human peripheral blood mononuclear cells activated by exogenous Nef. AIDS 10:1042-1043.
- Torres, B.A., T. Tanabe, and H.M. Johnson. 1996. Characterization of Nef-induce
- Torres, B.A., T. Tanabe, J.K. Yamamoto, and H.M. Johnson. 1996. HIV encodes for its own CE4 T cell superantigen mitogen. Biochem. Biophys. Res. Comm. 225:672-678.
- Szente, B.E., I.J. Weiner, M.J. Jablonsky, N.R. Krishna, B.A. Torres, and H.M. Johnson. 1996. Structural requirements for agonist activity of a murine interferon gamma peptide. J. Interferon Cytokine Res. 16:813-817.
- Subramaniam, P.S. and H.M. Johnson. 1996. A role for cyclin-dependent Kinase inhibitor p21 in the G1 cell cycle arrest mediated by type I interferons. J. Interferon Cytokine Res. In Press.
- Hobeika, A.C., P.S. Subramaniam. And H. M. Johnson. 1996. IFNalpha induces the expression of the cyclin-dependent kinase inhibitor p21 in human prostate cancer cells. Oncogene. In Press.
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Progress 10/01/94 to 09/30/95
Outputs
IFN . We have established a unique interaction between IFN and its receptor. TheN-terminus of IFN binds to the extracellular domain of the receptor, while the C-terminus binds to a site on the cytoplasmic domain of the receptor. The cytoplasmic binding occurs after IFN -receptor internalization. The cytoplasmic binding initiates a tyrosine tyrosine protein kinase cascade that is responsible for some of the biological effects of IFN . IFN#. We have shown that IFN# is effective in treatment of experimental allergic encephalomyelitis (EAE) in mice, an animal model for multiple sclerosis. The key finding is that IFN# can abrogate disease without toxicity such as bone marrow suppression. This has potential for IFN# treatment of a variety of diseases, including AIDS and cancer. Superantigens. We have found a superantigen in the human immunodeficiency virus (HIV), which is the causative agent of AIDS. The superantigen is the Nef protein. Inhibition of Nef induced V -specific
expansion of CD4 T cells by specific antibodies blocks HIV infection of CD4 T cells in tissue culture. This finding is important for understanding the mechanism of the disease, and for designing a strategy for treatment of AIDS.
Impacts
(N/A)
Publications
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Progress 10/01/93 to 09/30/94
Outputs
(1)A putative superantigen has been identified for the AIDS virus. It is the "regulatory" protein Nef. Nef is presented to T cells via the MHC Class II molecules of antigen-presenting cells and causes polyclonal proliferation of T-helper cells. This could result in providing blast cells for virus replication and ultimately drive the T cells to depletion and anergy or non- responsiveness; responses that are characteristic of AIDS. Controlling Nef superantigen activity has potential for prevention/treatment of AIDS.(2)Superantigens can exacerbate autoimmune conditions such as experimental allergic encephalomyelitis (EAE) in mice, an animal model for multiple sclerosis in humans. Superantigens aggravate EAE by causing a non-restricted polyclonal expansion of autoreactive T cells. The result is a much more aggressive disease that is more closely related to events that occur in multiple sclerosis in humans.(3)We have determined that gamma interferon (IFN ) binds to its
receptor at both the extracellular and intracellular domains. The N-terminus binds to the extracellular domain, while the C-terminus binds to the intracellular domain. Endocytosis of IFN is required for intracellular binding, and it is this binding that initiates the signal cascade of tyrosine kinases called Janus kinases and their associated transcription factor substrates. This is a newly discovered mechanism of ligand- receptor interactions for signal transduction.
Impacts
(N/A)
Publications
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Progress 10/01/92 to 09/30/93
Outputs
We have made the following progress with out interferon studies: 1. The solutionstructure of mouse gamma interferon (IFN(alpha)) N-terminal receptor-binding domain has been determined using 2-D nuclear magnetic resonance spectroscopy. 2. The receptor interacting functional regions of interferon tau (IFN(gamma)) has been determined using the synthetic peptide approach. Preliminary modeling shows that all of the regions are on the same face of the molecule. 3. Preliminary studies show that IFN(gamma) can inhibit function of cells involved in autoimmune disease. 4. We have identified the extracellular binding site on the IFN(alpha) receptor. We have made the following progress with our superantigen studies: 1. Bacterial superantigens can exacerbate autoimmune diseases such as experimental allergic encephalomyelitis, an animal model of multiple sclerosis. 2. The region on the viral superantigen of mouse mammary tumor virus that interacts with the MNC Class II receptor
has been identified.
Impacts
(N/A)
Publications
- JARPE, M.A. and JOHNSON, H.M. 1993. Stable conformation of IFN(gamma) receptor binding peptide in water is required for IFN(gamma) antagonist activity. J. Interferon Res. 13:99-103.
- SOOS, J.M., SCHIFFENBAUER, J. and JOHNSON, H.M. 1993. Treatment of PL/J mice with the superantigen, staphylococcal enterotoxin B, prevents development of experimental allergic encepholomyelitis. J. Neuroimmunol. 43:39-44.
- PONTZER, C.H., GRIGGS, N.D. and JOHNSON, H.M. 1993. Agonist properties of a microbial superantigen peptide. Biochem. Biophys. Research Commun. 193:1191-1197.
- SCHIFFENBAUER, J., SOOS, J., JOHNSON, H. and WEGRZYN, L. 1993. Staphylococcal enterotoxin can re-activate experimental allergic encephalomyelitis. Proc. Natl. Acad. Sci. USA. 90:8543-8546.
- SOOS, J.M., RUSSELL, J.K., JARPE, M.A., PONTZER, C.H. and JOHNSON, H.M. 1993. Regions of interaction between superantigen toxic shock syndrome toxin-1 and class II MHC molecules. Biochem. Biophys. Research Commun. 191:1211-1217.
- SAKAI, T.T., JABLONSKY, M.J., DEMUTH, P.A., KRISHNA, N.R., JARPE, M.A. and JOHNSON, H.M. 1993. Proton NMR sequence specific assignments and secondary structure of a receptor binding domain of mouse (gamma)-interferon. Biochemistry 32:5650-.
- TORRES, B.A., GRIGGS, N.D. and JOHNSON, H.M. 1993. Bacterial and retroviral sup.
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Progress 10/01/91 to 10/30/92
Outputs
Gamma interferon (IFN(gamma)) is an essential immunoregulatory lymphokine for a variety of immunological functions. The solution structure of a receptor-binding domain of murine IFN(gamma) has been determined using multidimensional nuclear magnetic resonance (NMR). This will facilitate the rational design of agonists and antagonists of IFN(gamma) activity, since function can be directly correlated with structural changes in the receptor-binding domain of IFN(gamma). Mutants of a synthetic IFN(gamma) gene have been expressed. The mutations are in position 14, which contains a tyrosine. Conservative aromatic changes do not affect IFN(gamma) activity, but nonaromatic substitutions result in significant lost of activity. Ovine trophoblast protein (oTP-1) is the interferon a variant (call interferon tau) with potent antiviral activity and/or toxicity that is responsible for maternal recognition of pregnancy in sheep. oTP-1 is a potent inhibitor of replication of human AIDS
(HIV) and feline AIDS (FIV) viruses in tissue culture without the toxicity to cells that is associated with high concentrations of interferon (alpha). Further, oTP-1 shows significant antitumor activity in mouse models of cancer. Thus, oTP-1 is a potent antiviral and antitumor cytokine without the toxicity associated with high concentrations or doses of interferon.
Impacts
(N/A)
Publications
- JOHNSON, H.M., J.K. RUSSELL, and C.H. PONTZER. 1992. Superantigens in human disease. Scientific American. April.
- RUSSELL, J.K., M.A. JARPE and H.M. JOHNSON. 1992. Evidence for the (alpha)-helicity of class II MHC molecular binding sites for the superantigen staphylococcal enterotoxin A. Biochem. Biophys. Res. Comm. 182:1016-1024.
- GRIGGS, N.D., C.H. PONTZER, M.A. JARPE and H.M. JOHNSON. 1992. Mapping of multiple binding domains of the superantigen SEA for HLA. J. Immunol. 148:2516-2521.
- GRIGGS, N.D., M.A. JARPE, J.L. PACE, S.W. RUSSELL, and H.M. JOHNSON.1992. The N-terminus and C-terminus of Interferon-(gamma) are binding domains for cloned soluble IFN(gamma)I receptor. J. Immunol. 149:517-520.
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Progress 10/01/90 to 09/30/91
Outputs
Gamma interferon (IFN(gamma)) is an essential immunoregulatory lymphokine for a variety of immunological functions. Using a soluble form of the murine IFN(gamma) receptor, we have further determined the binding regions on the IFN(gamma) molecule. The N-terminus as well as the C-terminus compete with IFN(gamma) for binding to receptor. This has facilitated the formulation of a novel model for receptor-ligand interaction. A synthetic gene for IFN(gamma) has been constructed and recombinant protein has been expressed in bacteria for the purpose of site directed mutagenesis for further structure/function studies and NMR analysis. Ovine trophoblast protein (oTP-1) is the interferon (alpha) (IFN(gamma)) variant with potent antiviral activity and low toxicity that is responsible for maternal recognition of pregnancy in sheep. Overlapping synthetic peptides which encompass the entire oTP-1 molecule have been used to examine functionally important regions of oTP-1 involved in
its antiviral, antiproliferative, and antileuteolytic activity. Preliminary results indicate that the carboxy-terminal peptide antagonizes oTP-1 antileuteolytic activity, and more importantly, the amino-terminal peptide may have agonist properties. Through maximization of recombinant oTP-1 expression, an adequate supply of oTP-1 is now available for continuing structural analysis.
Impacts
(N/A)
Publications
- JOHNSON, H.M., J.K. RUSSELL and C.H. PONTZER. 1991 Staphylococcal enterotoxin microbial superantigens. FASEB J., 5:2706-2712.
- JOHNSON, H.M., J.K. RUSSELL, and C.H. PONTZER. 1991. Staphylococcal enterotoxin superantigens. Proc. Soc. Exptl. Biol. Med., in-press.
- JOHNSON, H.M., J.K. RUSSELL, and C.H. PONTZER. 1991. Superantigens. Sci. American, in press.
- PONTZER, C.H., RUSSELL, J.K. and H.M. JOHNSON. 1990. Structural basis for differential binding of staphylococcal enterotoxin A and toxic shock syndrome toxin-one to class II major histocompatibility molecules. Proc. Natl. Acad. Sci. (USA).
- PONTZER, C.H., J.K. RUSSELL, M.A. JARPE and H.M. JOHNSON. 1991. Site of nonrestrictive binding of SEA to class II MHC antigens. Int. Arch. Allergy Appl. Immunol. 93:107-112.
- JOHNSON, H.M., C.H. PONTZER and J.K. RUSSELL. 1991. Staphylococcal microbial superantigens: MHC class II and T-cell receptor interactions. Progress in NeuroEndocrinImmunol. 4(1):10-19.
- RUSSELL, J.K., C.H. PONTZER and H.M. JOHNSON. 1991. Both a-helices along the MHC binding cleft are required for staphylococcal enterotoxin A function. Proc. Natl. Acad. Sci. (USA) 88:7228-7232.
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Progress 10/01/89 to 09/30/90
Outputs
Gamma interferon (IFN gamma) is an essential immunoregulatory lymphokine for a variety of immunologic functions. Using overlapping synthetic peptides that encompass the entire IFN gamma molecule and antibodies to the peptides, the receptor binding domains of IFN gamma were found to involve both the N-terminal and C-terminal domains of the molecule. Synthetic peptide IFN gamma(1-39) blocked binding of IFN gamma in receptor competition, and antibodies to IFN gamma(1-39) and IFN(95-133) neutralized IFN gamma activity. Using this and other data, a three dimensional model of IFN gamma was constructed that consists of a four-alpha-helical bundle along with closely associated N- and C-terminal receptor binding helices. Ovine trophoblast protein (oTP-1) is the interferon alpha (INA) variant with potent antiviral activity and low toxicity that is responsible for maternal recognition of pregnancy in sheep. Using structure prediction modeling, synthetic peptides and site
specific antibodies produced to the peptides, oTP-1 and IFN alpha are predicted to possess similar C-terminal structure but different N-terminal structure. Further, the C-terminal regions of oTP-1 and IFN alpha bind to a common site on the IFN alpha receptor, while the N-terminal region binds to a site unique for the particular IFN alpha.
Impacts
(N/A)
Publications
- JARPE, M.A. and JOHNSON, H.M. 1990. Structure of an epitope in an immunodominant region of the interferon-Y molecule that is involved in receptor interaction. J. Interferon Res. 10:243-252.
- PONTZER, C.H., OTT, T.L., BAZER, F.W. and JOHNSON, H.M. 1990. Localization of an antiviral site on the pregnancy recognition hormone, ovine trophoblast protein-one. Proc. Natl. Acad. Sci. USA 87:5945-5949.
- RUSSELL, J.K., PONTZER, C.H. and JOHNSON, H.M. 1990. The 1-A(B)_b region (65-85) is a binding site for the superantigen staphylococcal enterotoxin A. Biochem. Biophys. Res. Comm. 168:696-701.
- JARPE, M.A. and JOHNSON, H.M. 1990. Topology of receptor binding domains of mouse IFN(Y). J. Immunol. 145:3304-3309.
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