Clues to evolution of the SERA multigene family in 18 Plasmodium species

Abstract

SERA gene sequences were newly determined from 11 primate Plasmodium species including two human parasites, P. ovale and P. malariae, and the evolutionary history of SERA genes was analyzed together with 7 known species. All have one each of Group I to III cysteine-type SERA genes and varying number of Group IV serine-type SERA genes in tandem cluster. Notably, Group IV SERA genes were ascertained in all mammalian parasite lineages; and in two primate parasite lineages gene events such as duplication, truncation, fragmentation and gene loss occurred at high frequency in a manner that mimics the birth-and-death evolution model. Transcription profile of individual SERA genes varied greatly among rodent and monkey parasites. Results support the lineage-specific evolution of the Plasmodium SERA gene family. These findings provide further impetus for studies that could clarify/provide proof-of-concept that duplications of SERA genes were associated with the parasites' expansion of host range and the evolutionary conundrums of multigene families in Plasmodium.

Figure 1. The organization of the SERA gene family in 18 Plasmodium species.

SERA genes are arrayed onto a solid horizontal line for each parasite species. Gene arrangement follows the genomic organization in each species. Individual SERA genes are clustered between a conserved hypothetical gene and the iron-sulfur assembly protein gene. Pfa-SERA9, Pre-SERA9 and PgaSERA3 have aberrant locations. SERA genes were categorized to Groups I to IV and Clades 1 to 6 reflecting orthologous gene groups as inferred from phylogenetic analyses shown in Figure 2 and Figure S3. SERA genes of Groups I to III (cysteine-type SERA gene) and those of Group IV (serine-type SERA gene) are shown in green and blue, respectively. TSERA denotes truncated SERA genes shown in yellow. SERA genes are drawn to scale, but other genes and intergenic regions are not. Dashed lines and/or dark gray boxes denote orthologous relationships. A generally accepted consensus phylogenetic tree of Plasmodium species is shown in right. The abbreviations for species names are: P. falciparum (Pfa), P. vivax (Pvi), P. malariae (Pma), P. ovale (Pov), P. reichenowi (Pre), P. gonderi (Pgo), P. fragile (Pfr), P. coatneyi (Pco), P. knowlesi (Pkn), P. inui (Pin), P. fieldi (Pfi), P. simiovale (Pso), P. cynomolgi (Pcy), P. hylobati (Phy), P. yoelii (Pyo), P. berghei (Pbe), P. chabaudi (Pch), and P. gallinaceum (Pga). The SERA gene family has common exon/intron structure: four exons and three introns, with some exceptions. Group I SERA genes have six exons and five introns structure, except for Pfa-SERA8 and Pvi-SERA12, which lack one intron. SERA genes of Group IV Clade 2 and Pma-SERA1 have no third intron and consist of three exons and two introns. Group I SERA genes of three rodent parasites have an extra intron near the 5′-end. Pkn-SERA1 gene in Clade 1 contains three stop codons, causing truncation of the cysteine-rich conserved domain; but since this gene was expressed, we consider this is a SERA gene. TSERA1 genes have truncations of the protease domain, variable domain 2 and cysteine-rich conserved domain. Pco-TSERA2, Pkn-TSERA2 and Pfr-TSERA2 lack a long amino acid region (498 residues) including the enzyme domain (Figure S4), of which Pfr-TSERA2 seems to be a putative pseudogene because its 2nd exon contains two stop codons.

Figure 2. The maximum likelihood (ML) phylogenetic tree of Plasmodium SERA genes.

This unrooted tree was constructed from 115 SERA genes (encompassing Groups II to IV, see Figure 1) using 570 amino acid positions under the JTT + Γ (eight categories) model (α = 1.15) with 500 heuristic replicates. Bootstrap proportions >50% are shown along nodes. Groups II - III and Group IV are cysteine-type and serine-type SERA genes, respectively. Note that Pga-SERA1 (ga1) is an offshoot of Groups II and III SERA genes, suggesting the occurrence of a common ancestor, leading to Pga-SERA1 (ga1) and a common ancestor of Group II and Group III. In P. vivax and P. vivax-related monkey malaria parasite species, the six clades are color-boxed. Pgo-SERA1 (go1) and Pgo-SERA5 (go5) were grouped, despite low bootstrap values, into Clade 2 and Clade 5 respectively, because these genes showed common features to each clade in exon/intron structure and/or gene array.

Figure 3. Primary structure and inter-species sequence variation of the Plasmodium SERA gene.

The putative domain structure of the gene family is shown in (A). In (B) are sequence variations in Groups I to IV SERA domains using amino acid sequence similarity for three parasite lineages: (i) P. vivax and P. vivax-related monkey malaria parasite species, (ii) three rodent parasite species, and (iii) P. falciparum and P. reichenowi. Percent (%) similarity is color coded.

Figure 4. The SERA gene organization of P. vivax and P. vivax-related monkey malaria parasites.

Six monophylic clades of Group IV SERA genes, Clade 1 to Clade 6 are designated in colored boxes. SERA genes, truncated SERA genes and SERA gene fragments are shown by light gray circle, square, and triangle, respectively. The conserved hypothetical protein gene at 5′-end and the iron-sulfur assembly protein gene at the 3′-end are shown by dark gray circle and open circle, respectively.

Figure 5. Transcription of the SERA gene family of rodent parasite P. berghei, and three monkey parasites, P. cynomolgi, P. coatneyi and P. knowlesi.

Parasites at the late trophozoite to schizont stages were used for analyses. Relative amounts of transcribed SERA genes were standardized relative to β-tubulin (TBB), set at 1.0. Cysteine-type SERA genes (Groups I, II and III) and serine-type SERA genes (Group IV) are arrayed in gray shaded areas and unshaded areas, respectively. SERA genes of three monkey parasites are separated into six clades, as designated on top of the P. cynomolgi panel.

Figure 6. Inferred gene duplication events in the evolution of the Plasmodium SERA gene family.

Each colored circle represents cysteine-type SERA gene (Groups I to III, green circle) and serine-type SERA genes (Group IV, blue circle), and is placed alongside parasite lineages of a generally accepted phylogenetic tree of Plasmodium, with Theileria used as an outgroup.