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Investigating the function of MACPF proteins using Drosophila melanogaster
thesis
posted on 2017-02-07, 03:25 authored by Lauren Forbes-BeadleThe membrane
attack complex/perforin-like (MACPF) protein superfamily contains members found
within all kingdoms of life. Many MACPF proteins play important roles in
mammalian immunity because of their inherent ability to insert into cell
membranes and form pores. The complement proteins of the membrane attack
complex and the perforin protein are well characterised MACPF proteins involved
in immunity that have the ability to form pores. Not all MACPF proteins form
pores however, and many have functions in development. These developmental
roles are less well understood, and in addition many MACPF proteins have
unknown functions.
To address this, here the Drosophila model system has been utilised to investigate the in vivo roles of a number of MACPF proteins. Torso-like is the sole Drosophila MACPF protein and functions in embryonic development in the terminal patterning pathway. This pathway controls the specification of the anterior and posterior termini of the developing embryo. This thesis first aimed to determine if and how Tsl acts in Drosophila immunity, and if this function is similar to mammalian MACPF proteins that function in the immune response. By infecting a tsl null mutant with a range of pathogens, it was found that Tsl is required for the Drosophila immune response. Unexpectedly tsl null mutants had a reduced number of immune cells indicating that Tsl may be required for the development of these cells rather than acting as an immune effector.
Secondly, to further understand how Tsl functions, MACPF proteins from distinct organisms such as mammals and the sea urchin were expressed in Drosophila to determine if they could functionally replace Tsl. This method was also used to discover novel functions for MACPF proteins by utilising the functional conservation of many biological pathways between Drosophila and other organisms. The MACPF proteins tested could not replace the function of Tsl in terminal patterning of the embryo. Unexpectedly however, a novel function in the Notch pathway was uncovered for two distantly related MACPF proteins, the protein Mpeg1 and the sea urchin protein Apextrin.
Overall this thesis demonstrates the diverse developmental roles of MACPF proteins and highlights the utility of using Drosophila as a model system to investigate gene function. Due to the evolutionary conservation of important signalling pathways, the study of MACPF proteins in Drosophila provides significant insight into the function of MACPF proteins in new developmental roles. The possible cellular effects of MACPF proteins are discussed in relation to these novel findings.
To address this, here the Drosophila model system has been utilised to investigate the in vivo roles of a number of MACPF proteins. Torso-like is the sole Drosophila MACPF protein and functions in embryonic development in the terminal patterning pathway. This pathway controls the specification of the anterior and posterior termini of the developing embryo. This thesis first aimed to determine if and how Tsl acts in Drosophila immunity, and if this function is similar to mammalian MACPF proteins that function in the immune response. By infecting a tsl null mutant with a range of pathogens, it was found that Tsl is required for the Drosophila immune response. Unexpectedly tsl null mutants had a reduced number of immune cells indicating that Tsl may be required for the development of these cells rather than acting as an immune effector.
Secondly, to further understand how Tsl functions, MACPF proteins from distinct organisms such as mammals and the sea urchin were expressed in Drosophila to determine if they could functionally replace Tsl. This method was also used to discover novel functions for MACPF proteins by utilising the functional conservation of many biological pathways between Drosophila and other organisms. The MACPF proteins tested could not replace the function of Tsl in terminal patterning of the embryo. Unexpectedly however, a novel function in the Notch pathway was uncovered for two distantly related MACPF proteins, the protein Mpeg1 and the sea urchin protein Apextrin.
Overall this thesis demonstrates the diverse developmental roles of MACPF proteins and highlights the utility of using Drosophila as a model system to investigate gene function. Due to the evolutionary conservation of important signalling pathways, the study of MACPF proteins in Drosophila provides significant insight into the function of MACPF proteins in new developmental roles. The possible cellular effects of MACPF proteins are discussed in relation to these novel findings.
