by Hannah Sharpe
The first year of the IITM DPhil consists of three rotations in different labs, covering a broad range of disciplines and research. To summarise the outcomes of the first rotation, the students gave poster presentations to their peers and supervisors. Here is an outline of the achievements of each project.
Robert’s first rotation was in Simon Draper’s lab at the Jenner Institute, and contributed towards the development of a blood-stage vaccine for Plasmodium falciparum malaria infection. PfCyRPA forms part of a triple complex of proteins involved in Plasmodium merozoite erythrocyte invasion. Robert designed and made mutants of PfCyRPA through altering amino acids on the surface of PfCyRPA, and through addition of glycosylation sites. These mutants were tested in a growth inhibition assay using monoclonal and polyclonal anti-PfCyRPA antibodies.
Mari’s project was also related to malaria, and was conducted Dr Matt Higgins’ lab in the Biochemistry department. During her project, she investigated the widely-conserved HAP2 fusion protein involved in gametocyte fusion of Plasmodium, and how it could be used as a potential vaccine against malaria. During this project, Mari used Phyre and Pross modelling programmes to design structurally homologous proteins to HAP2, consisting of fused HAP2 alpha helical bundles and fusion loops, as well as whole HAP2 protein vaccines. These proteins were cloned into E. coli, and some of these protein constructs were successfully purified.
Lea worked in the lab of William James at the Dunn School. Here, she studied multinucleated fusion cells that arise during HIV-1 infection through the interaction of membrane-bound HIV-1 envelope proteins with CD4 and co-receptor molecules, and which can act as HIV-1 reservoirs in the brain. Lea showed that multinucleated macrophage-T cell fusion cells have similar gene expression pattern to macrophages but increased SAMHD1 phosphorylation. This was ascertained through development of a CEM T cell line expressing the HIV-1 Bal envelope protein, exposing them to macrophages, and using RNA expression analysis to measure gene expression.
Hannah’s first rotation project was with Professor Ellie Barnes, and aimed to characterise the T cell responses to rodent hepacivirus in infected and vaccinated rodents. Hannah used ELISpot assays to map the T cell epitopes in rats infected with RHV, and rats and mice vaccinated against RHV using a chimpanzee adenovirus-vectored vaccine. She then conducted intracellular cytokine staining to ascertain whether the epitopes elicited a CD4+ or CD8+ T cell response. The ultimate aim is to use RHV to develop an animal model of closely-related hepatitis C virus infection, in order to improve vaccine development and further research into HCV.
Sarah’s first rotation project took her to Hal Drakesmith’s lab at the Weatherall Institute of Molecular Medicine. The Drakesmith lab researches iron deficiency, and how it can affect vaccine efficacy. Here, Sarah investigated how iron deficiency in murine T cells and dendritic cells have impaired proliferation, DNA synthesis, and cytokine production. She demonstrated that iron deficiency alters differentiation of antigen specific CD8+ T cells, and impairs antigen-specific T cell metabolism by using a Seahorse XF analyser to measure glycolysis, and that it also reduces dendritic cell cytokine production through flow cytometry and ELISA assays.
Overall, the first rotations were a success, and the first years are currently enjoying the first few weeks of their second rotation!