Diversity and evolution of the rhoph1/clag multigene family of Plasmodium falciparum

Abstract

A complex of high molecular mass proteins (PfRhopH) of the human malaria parasite Plasmodium falciparum induces host protective immunity and therefore is a candidate for vaccine development. Understanding the level of polymorphism and the evolutionary processes is important for advancements in both vaccine design and knowledge of the evolution of cell invasion in this parasite. In the present study, we sequenced the entire open reading frames of seven genes encoding the proteins of the PfRhopH complex (rhoph2, rhoph3, and five rhoph1/clag gene paralogs). We found that four rhoph1/clag genes (clag2, 3.1, 3.2, and 8) were highly polymorphic. Amino acid substitutions and indels are predominantly clustered around amino acid positions 1000-1200 of these four rhoph1/clag genes. An excess of nonsynonymous substitutions over synonymous substitutions was detected for clag8 and 9, indicating positive selection. The McDonald-Kreitman test with a Plasmodium reichenowi orthologous sequence also supports positive selection on clag8. Based on the ratio of interspecific genetic distance to intraspecific distance, the time to the most recent common ancestor of the clag2 and 8 polymorphisms was estimated to be 1.89 and 0.87 million years ago, respectively, assuming divergence of P. falciparum and P. reichenowi 6 million years ago. In addition to a copy number polymorphism, gene conversion events were detected for the rhoph1/clag genes on chromosome 3, which likely play a role in increasing the diversity of each locus. Our results indicate that a high diversity of the PfRhopH1/Clag multigene family is maintained by diversifying selection forces over a considerably long period.

A CLAG3 Mutation in an Amphipathic Transmembrane Domain Alters Malaria Parasite Nutrient Channels and Confers Leupeptin Resistance

“…The A1210T mutation in HB3-leuR1 CLAG3 is near the protein's C terminus, distal to a 10-to 30-residue motif that varies between P. falciparum lines (28,29). Functional proteolysis studies found that this variable motif is exposed at the erythrocyte surface and that the CLAG3 C terminus is intracellular (16), implicating one or more transmembrane domains within this segment (Fig.…”

“…As with most P. falciparum lines (28,36), the transfectant parasite carries two clag3 genes. Individual parasites in a culture express only one of these two genes and silence the other through epigenetic mechanisms (13,37).…”

An Epigenetic Antimalarial Resistance Mechanism Involving Parasite Genes Linked to Nutrient Uptake

“…Although most plasmodial species have only two or three clag genes in their genomes (27,28), P. falciparum has undergone significant expansion to have five members in this gene family. To explore whether the other paralogs contribute to PSAC, we performed phylogenetic analysis of clag genes from laboratory lines representing Africa, South America, and Southeast Asia and found that each paralog has primary sequence elements that distinguish it from all other clag products, with the clag9 group being the most divergent (Fig.…”

Expression of the Plasmodium falciparum Clonally Variant clag3 Genes in Human Infections

“…The 2 clag3 genes, clag3.1 (PF3D7_0302500) and clag3.2 (PF3D7_0302200), are separated by only 10 kb and show 95% sequence coincidence. The clag3.1 or clag3.2 identity is determined by the relative position of each gene in the chromosome and by the conserved clag3.1-or clag3.2-specific flanking regions [13]. These genes are part of the 5-member clag family, which encodes the CLAG/RhopH1 component of the RhopH complex.…”