Investigation of the cir multi-gene family of Plasmodium chabaudi.
Doctoral thesis, UCL (University College London).
The pir genes comprise the largest multi-gene family in Plasmodium, with members found in P. vivax, P. knowlesi and rodent malaria species. Despite their almost universal presence, little is known about the functions of the PIR proteins. To investigate the role PIR proteins play during the erythrocytic stages of infection, the P. chabaudi model was chosen, where this gene family is termed cir. 198 cir genes were identified in the P. chabaudi genome, 86% of which clustered to form two major sub-families on the basis of sequence similarity. Quantitative RT-PCR and Illumina RNA sequencing were used to investigate cir transcription during P. chabaudi infection. Both methods detected many cir genes transcribed at low levels, as shown previously for other pirs. Three of the transcribed cir genes were selected for recombinant protein expression in the yeast Pichia pastoris: PCHAS_000100, PCHAS_070130 and PCHAS_040110. Soluble PCHAS_000100 was used for measurements of CIR secondary structure. Conserved and sub-family specific peptides were also synthesized. Antibodies present in the sera of P. chabaudi immune mice recognized all CIR proteins and peptides. Polyclonal antibodies were used to determine CIR localization by confocal microscopy and flow cytometry. Whilst most CIRs were located within the parasites, some CIRs were also found on the infected erythrocyte surface of mature trophozoites. In addition, CIRs were detected at the apical end of merozoites. These results imply that CIR proteins are exposed to the immune system during P. chabaudi infection and are antigenic, yet immunization with most CIR proteins and peptides did not protect mice from P. chabaudi infection. Upon P. chabaudi challenge of mice immunized with CIR sub-family specific reagents, increased levels of cir transcripts belonging to the other major sub-family were detected. This may explain why few differences in parasitaemia were observed. The exception was observed during P. chabaudi challenge of mice immunized with PCHAS_000100, which were able to clear parasitaemia earlier than controls.
|Title:||Investigation of the cir multi-gene family of Plasmodium chabaudi|
|Open access status:||An open access version is available from UCL Discovery|
|UCL classification:||UCL > School of Life and Medical Sciences > Faculty of Medical Sciences > Infection and Immunity (Division of)|
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