: Babesia bovis is a protozoan intraerythrocytic parasite that establishes persistent infections of extremely long duration. Although recognition of parasite antigens on the infected red blood cell (IRBC) membrane surface by antibodies may provide protection from disease, the host cannot develop immunity to infection. At least two mechanisms are used by this parasite to establish persistent infection: [1] cytoadhesion of IRBCs to the microvascular endothelium prevents mature parasite stages from circulating, thus avoiding the spleen; and [2] antigenic variation of the parasite components on the IRBC surface """"""""protects"""""""" the cytoadhesive ligand from immune recognition, allowing variant parasites to persist. Strong circumstantial evidence implicates the VESA1 antigen in cytoadhesion, identifying the VESA1 as a bifunctional virulence factor of immune evasion. Previously, we identified the B. bovis ves multi-gene family encoding the VESAla polypeptide, a major participant in antigenic variation on the IRBC surface, and found evidence this occurs through segmental gene conversion. Recently, we have discovered this gene family to be consistently organized as palindromic (head-to-head) gene pairs, without obvious differences in the upstream regulatory sequences of transcribed and non-transcribed copies. However, this organization is likely significant to the regulation of expression of this polypeptide. We therefore propose to determine the general mechanisms responsible for regulating expression of the variant antigens, by completion of the following aims:
Aim 1 : To detect and discriminate between allelic exclusion and post-transcriptional mechanisms in the control of VESAla expression.
Aim 2 : To determine whether higher-order in vivo structure in the 5' end of the vesla gene differs between sequence donor and actively expressed (sequence recipient) vesla genes, and to determine the identity of the """"""""pseudogenes"""""""" apposing many vesla genes.
Aim 3 : To determine the chromosomal location and define the full structure of a known vesla site of transcription. The outcomes of this project will define the gross mechanisms used by this parasite to survive, and will direct future work targeting the enzymatic machinery of this and other microbes using similar strategies as a means to limit parasite survival.
