T cell antigens and immune evasion genes from the parasitic nematode Brugia malayi

• Main Author: Zang, Xingxing
• Published: University of Edinburgh 1999
• Subjects: 591.35
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.664211

The majority of this thesis deals with the identification, gene cloning, protein expression, molecular function and vaccine development of major T cell antigens in <I>B. malayi. </I>Immunization of microfilariae (Mf) proteins with different adjuvants selectively induced antigen-specific Th1 and Th2 responses <I>in vivo. </I>Mf-specific T cells, including Th1 and Th2 cells, were used to identify Mf protein fractions separated by fast protein liquid chromatography and SDS-polyacrylamide gel electrophoresis elution. This revealed a restricted number of major T cell antigens from Mf, a minority of which were insoluble. A novel serine proteinase inhibitor (serpin) gene (<I>Bm-spn-</I>2) was cloned by screening an Mf cDNA library with antisera against one Mf protein fraction which was highly potent at inducing antigen-specific T cell proliferation and cytokine production, while only the paramyosin gene was cloned using antisera against whole Mf proteins. <I>Bm-spn-</I>2, a single copy gene consisting of seven exons, was abundantly (>2% of total mRNA) and exclusively expressed by the Mf stage. This sequence analysis has been broadened into a genome data bank-based review of intron and exon sequences in <I>B. malayi</I>, which revealed an extended and conserved 3' splice site in all recorded <I>B. malayi </I>genes and relatively large introns compared with <I>Caenorhabditis elegans. Bm-spn-2</I> contains 428 amino acids with a putative signal peptide and native protein was 47.5 kDa, one of the largest of the 93 known serpins. The recombinant Bm-SPN-2 protein was expressed in bacteria and purified to determine biological function. A panel of mammalian serine proteinases, which are involved in a wide range of biological processes, were screened and Bm-SPN-2 protein found to specifically inhibit enzymatic activities of human neutrophil cathepsin G and human neutrophil elastase, but not a range of other serine proteinases. This suggests that Bm-SPN-2 may directly interfere with human immune function. Human neutrophil cathepsin G and elastase mediate multiple host defence functions such as interleukin processing, chemokinetic stimulation for T lymphocytes and chemoattraction for monocytes. We hypothesize that neutrophils, which represent 55% of human blood leukocytes, would be the primary cell type to interact with <I>B. malayi</I> Mf, and that release of Bm-SPN-2 neutralizes the immune-stimulating properties of neutrophil serine proteinases.