Cytokine-Glycosaminoglycan Interactions and the Role of Dermatan Sulphate-PG and TGF-β1 in the Regulation of Interferon-γ Production by Murine NK Cells. / Bavisi, Karishma.

2019. 291 p.

Research output: ThesisDoctoral Thesis

Unpublished

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@phdthesis{41cc1e2be31a4c1ca9f2b6c0fcc0d06a,
title = "Cytokine-Glycosaminoglycan Interactions and the Role of Dermatan Sulphate-PG and TGF-β1 in the Regulation of Interferon-γ Production by Murine NK Cells",
abstract = "Enhancing the understanding of the biological and functional role of specific cytokines in the regulation of Natural Killer (NK) cell activity is vital to cell-mediated immunity against infections and tumour development. Binding of cytokines with glycosaminoglycans (GAGs) and proteoglycans (PGs) have been known as a potential mechanism for regulating biological activity of many cytokines. In this thesis work, the binding of interleukins IL-11, IL-18 and IL-22 to heparin/HS was investigated using an ELISA approach, alongwith examination of primary sequences, 3-D protein structures and molecular docking calculations. Our results demonstrate that these interleukins do not bind to heparin/HS, which implies that the heparin-binding property should not be considered a generic characteristic for any class of cytokines. Further research focussed on examining the functional role of NK cell-surface PG, particularly dermatan sulphate (DS), in IL-12 signalled IFN-γ production using a PG synthesis inhibitor, p-nitrophenyl-β-xyloside. Our findings show partial inhibition of IFN-γ due to disrupted GAG/PG metabolism. Mechanistically, DS-PG interference was found independent of STAT-4 phosphorylation but suggestive of involvement essentially at the transcriptional or post-transcriptional level. In addition, our data implies that p-nitrophenyl-β-xyloside exerts cellular effects attenuating NK cell response to IL-12, independent of GAG-PGs. Finally, an interplay of molecular mechanisms between immunoregulatory factors, TGF-β1 and IL-12, in suppressing IFN-γ expression in mNK cells is studied. A clear evidence of partial inhibition of IL-12-induced IFN-γ production independent of NK cell proliferation is presented that does not directly involve key transcription factors of IFN-γ expression, STAT-4 and T-bet. However, promoter and early inhibition (3-6 hrs) studies suggested a possible role of TGF-β1 in chromatin remodelling of Ifng locus that regulates IFN-γ expression. Overall, identifying IL-heparin/HS interactions within different cytokine families and dissecting molecular events of cytokine signalling in NK cells are therapeutically important in recognising potential targets to manage cell-mediated immune responses.",
keywords = "Cytokine-Glycosaminoglycan, GAGs, GAG-binding, Heparin/Heparan sulphate, Dermatan Sulphate, Interleukin-11 (IL-11), Interleukin-18 (IL-18), Interleukin-22 (IL-22), Interleukin 12 (IL-12), Interferon-gamma, TGF-beta1, NK cells, STAT-4, T-bet, IL-12 signalling, Heparin-binding",
author = "Karishma Bavisi",
year = "2019",
language = "English",
school = "Royal Holloway, University of London",

}

RIS

TY - THES

T1 - Cytokine-Glycosaminoglycan Interactions and the Role of Dermatan Sulphate-PG and TGF-β1 in the Regulation of Interferon-γ Production by Murine NK Cells

AU - Bavisi, Karishma

PY - 2019

Y1 - 2019

N2 - Enhancing the understanding of the biological and functional role of specific cytokines in the regulation of Natural Killer (NK) cell activity is vital to cell-mediated immunity against infections and tumour development. Binding of cytokines with glycosaminoglycans (GAGs) and proteoglycans (PGs) have been known as a potential mechanism for regulating biological activity of many cytokines. In this thesis work, the binding of interleukins IL-11, IL-18 and IL-22 to heparin/HS was investigated using an ELISA approach, alongwith examination of primary sequences, 3-D protein structures and molecular docking calculations. Our results demonstrate that these interleukins do not bind to heparin/HS, which implies that the heparin-binding property should not be considered a generic characteristic for any class of cytokines. Further research focussed on examining the functional role of NK cell-surface PG, particularly dermatan sulphate (DS), in IL-12 signalled IFN-γ production using a PG synthesis inhibitor, p-nitrophenyl-β-xyloside. Our findings show partial inhibition of IFN-γ due to disrupted GAG/PG metabolism. Mechanistically, DS-PG interference was found independent of STAT-4 phosphorylation but suggestive of involvement essentially at the transcriptional or post-transcriptional level. In addition, our data implies that p-nitrophenyl-β-xyloside exerts cellular effects attenuating NK cell response to IL-12, independent of GAG-PGs. Finally, an interplay of molecular mechanisms between immunoregulatory factors, TGF-β1 and IL-12, in suppressing IFN-γ expression in mNK cells is studied. A clear evidence of partial inhibition of IL-12-induced IFN-γ production independent of NK cell proliferation is presented that does not directly involve key transcription factors of IFN-γ expression, STAT-4 and T-bet. However, promoter and early inhibition (3-6 hrs) studies suggested a possible role of TGF-β1 in chromatin remodelling of Ifng locus that regulates IFN-γ expression. Overall, identifying IL-heparin/HS interactions within different cytokine families and dissecting molecular events of cytokine signalling in NK cells are therapeutically important in recognising potential targets to manage cell-mediated immune responses.

AB - Enhancing the understanding of the biological and functional role of specific cytokines in the regulation of Natural Killer (NK) cell activity is vital to cell-mediated immunity against infections and tumour development. Binding of cytokines with glycosaminoglycans (GAGs) and proteoglycans (PGs) have been known as a potential mechanism for regulating biological activity of many cytokines. In this thesis work, the binding of interleukins IL-11, IL-18 and IL-22 to heparin/HS was investigated using an ELISA approach, alongwith examination of primary sequences, 3-D protein structures and molecular docking calculations. Our results demonstrate that these interleukins do not bind to heparin/HS, which implies that the heparin-binding property should not be considered a generic characteristic for any class of cytokines. Further research focussed on examining the functional role of NK cell-surface PG, particularly dermatan sulphate (DS), in IL-12 signalled IFN-γ production using a PG synthesis inhibitor, p-nitrophenyl-β-xyloside. Our findings show partial inhibition of IFN-γ due to disrupted GAG/PG metabolism. Mechanistically, DS-PG interference was found independent of STAT-4 phosphorylation but suggestive of involvement essentially at the transcriptional or post-transcriptional level. In addition, our data implies that p-nitrophenyl-β-xyloside exerts cellular effects attenuating NK cell response to IL-12, independent of GAG-PGs. Finally, an interplay of molecular mechanisms between immunoregulatory factors, TGF-β1 and IL-12, in suppressing IFN-γ expression in mNK cells is studied. A clear evidence of partial inhibition of IL-12-induced IFN-γ production independent of NK cell proliferation is presented that does not directly involve key transcription factors of IFN-γ expression, STAT-4 and T-bet. However, promoter and early inhibition (3-6 hrs) studies suggested a possible role of TGF-β1 in chromatin remodelling of Ifng locus that regulates IFN-γ expression. Overall, identifying IL-heparin/HS interactions within different cytokine families and dissecting molecular events of cytokine signalling in NK cells are therapeutically important in recognising potential targets to manage cell-mediated immune responses.

KW - Cytokine-Glycosaminoglycan

KW - GAGs

KW - GAG-binding

KW - Heparin/Heparan sulphate

KW - Dermatan Sulphate

KW - Interleukin-11 (IL-11)

KW - Interleukin-18 (IL-18)

KW - Interleukin-22 (IL-22)

KW - Interleukin 12 (IL-12)

KW - Interferon-gamma

KW - TGF-beta1

KW - NK cells

KW - STAT-4

KW - T-bet

KW - IL-12 signalling

KW - Heparin-binding

M3 - Doctoral Thesis

ER -