Studies on mechanisms of Fcgamma receptor internalisation of antigen

Melville, Bryony Claire (1998) Studies on mechanisms of Fcgamma receptor internalisation of antigen. MSc(R) thesis, University of Glasgow.

Abstract

A differential pattern of tyrosine kinase phosphorylation for distinct FcgammaR signalling domains was defined in several ways. Firstly, Syk recruitment was observed in interferon gamma (IFNgamma) and dibutyl cAMP (DBC) treated U937 monocyte-like cells, upregulating FcgammaRI + gamma chain of the FcepsilonRI and FcgammaRII receptors respectively. IFNgamma cells showed peak Syk activation at 5-7 minutes post-activation, while DBC cells show an immediate increase in Syk recruitment. In addition, blots of post-activation lysates of the two cell types were probed with anti-phosphotyrosine antibody, revealing a strong band of 110kDa on both IFNgamma and DBC blots, and an unknown 50kDa band found exclusively on the DBC blot. Lastly, experiments designed to test whether or not the two cells types display alternative NO2 release patterns post-activation were established. In a second approach, a chimeric receptor was made by fusion of the extracellular region of FcgammaRI to the transmembrane and cytoplasmic tail of zeta chain. This chimeric receptor was then compared to other similarly constructed mutants, FcgammaRI-gamma and FcgammaRI-II for its ability to phagocytose sheep red blood cells (SRBC) in a well established phagocytosis experiment using COS-7 cells (a simian kidney fibroblast cell line). All mutants demonstrated relatively equivalent phagocytic ability in the COS-7 cell system. Select tyrosine kinases were then cotransfected with each mutant FcgammaR receptor. There was no marked difference in distinct tyrosine kinases hFyn, mLyn, and mSyk required by distinct internal tyrosine kinase activation motifs (ITAMs) for increasing phagocytic efficiency. However, it was discovered that the effect of the tyrosine kinase mSyk on FcgammaRI/gamma, FcgammaRI/zeta, and FcgammaRI/II internalisation was very significant, dramatically increasing both the number of COS-7 cells capable of phagocytosis and the number of SRBCs phagocytosed by each individual COS-7 cell.

• Item Type: Thesis (MSc(R))
• Qualification Level: Masters
• Keywords: Molecular biology
• Colleges/Schools: College of Medical Veterinary and Life Sciences
• Supervisor's Name: Supervisor, not known
• Date of Award: 1998
• Depositing User: Enlighten Team
• Unique ID: glathesis:1998-71307
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 10 May 2019 10:49
• Last Modified: 24 Oct 2022 08:54
• Thesis DOI: 10.5525/gla.thesis.71307
• URI: https://theses.gla.ac.uk/id/eprint/71307