Two genes encoding unique proliferating-cell-nuclear-antigens are expressed in Toxoplasma gondii

Complete cDNA sequences encoding two novel proliferating-cell-nuclear-antigens (designated TgPCNA1 and 2) were isolated from a Toxoplasma gondii tachyzoite cDNA library, and Southern analysis using cDNA probes confirmed the presence of two PCNA genes in T. gondii genomic DNA. Expressed-sequence-tags were identified in the T. gondii database that matched each TgPCNA cDNA and closely related PCNA coding regions (designated PfPCNA1 and 2) were discovered in sequence data obtained from chromosome 12 and 13 of Plasmodium falciparum. TgPCNA1 and PfPCNA1 were found to share the highest amino acid identity at 49% compared to TgPCNA2 and PfPCNA2 (37% identity) whereas intraspecies PCNAs were determined to be less similar (27-30% identity). Phylogenetic analysis suggests the two apicomplexan PCNAs are the result of a gene duplication in the common ancestor of these parasites. Antibodies specific for TgPCNA1 ( approximately 40 kDa) or TgPCNA2 ( approximately 37 kDa) detected single antigen species in tachyzoite extracts that were expressed at similar levels in isolates representative of the T. gondii Type I, II and III strains. TgPCNA1-specific cDNA probes detected multiple mRNA species on Northern blots, which when combined, were expressed 5-7 fold higher than the single species of mRNA detected by the TgPCNA2 probe. The difference in the number of mRNA species and comparative mRNA levels suggests each TgPCNA gene is independently controlled, although in light of the nearly equal levels of protein a post-transcriptional mechanism may be responsible for equalizing protein expression.