Two Plasmodium 6-Cys family-related proteins have distinct and critical roles in liver-stage development

The 10 Plasmodium 6-Cys proteins have critical roles throughout parasite development and are targets for antimalaria vaccination strategies. We analyzed the conserved 6-cysteine domain of this family and show that only the last 4 positionally conserved cysteine residues are diagnostic for this domain and identified 4 additional “6-Cys family-related” proteins. Two of these, sequestrin and B9, are critical to Plasmodium liver-stage development. RT-PCR and immunofluorescence assays show that B9 is translationally repressed in sporozoites and is expressed after hepatocyte invasion where it localizes to the parasite plasma membrane. Mutants lacking B9 expression in the rodent malaria parasites P. berghei and P. yoelii and the human parasite P. falciparum developmentally arrest in hepatocytes. P. berghei mutants arrest in the livers of BALB/c (100%) and C57BL6 mice (>99.9%), and in cultures of Huh7 human-hepatoma cell line. Similarly, P. falciparum mutants while fully infectious to primary human hepatocytes ab...