Allelic Dimorphism Research Articles

Abstract 2900: To develop a next generation NK therapy with deep and durable response by engineered CD16 enhancing NK cytotoxicity

Abstract Introduction: Allogeneic NK cell-based therapies have been demonstrated to elicit clinical efficacy against hematopoietic malignancies. Unlike T-cell therapies, allogeneic NK cells do not cause toxicities such as serious cytokine release syndrome, neurotoxicity, or graft-vs-host-disease, and are less expensive. We believe NK cell-based therapies provide an important platform for “off-shelf” allogeneic cell therapies, so we designed a panel of arming strategies to enhance NK cytotoxicity and elicit deep and durable response. CD16a is a transmembrane protein that co-localizes with CD3ζ and FcεRIγ on NK cells and binds to the Fc portion of IgG antibodies. Upon ligation, it induces a potent series of signals resulting in cytokine production and cytotoxic effector activity via ADCC. Here we use the term CD16 to refer to CD16a on NK cells. CD16 is a potent activating receptor and is necessary for NK cell cytotoxic effector, so engineered CD16 is the cornerstone of NK enhancing strategy. CD16 displays a functional allelic dimorphism generating allotypes with either a phenylalanine (F) or a valine (V) residue at amino acid position 158. CD16VV is associated with higher ADCC activity and clinical response than CD16VF and CD16FF. However, only 15% of the population is CD16VV that maximizes ADCC. Several studies reported that tumor microenvironment may cause the shedding of CD16 which suppresses ADCC though ADAM17-mediated proteolytic cleavage upon NK cell activation. Therefore, selective inhibition of ADAM17 can reduce the shedding of CD16 in NK cells. Method: We designed a library of CD16 point mutations in the cleavage region of ADAM17 to screen for ADAM17-resistant CD16 mutations to prevent CD16 shedding and improve the efficacy of NK therapy. NK92 cells lacking expression of endogenous CD16 were transduced to express wild type CD16 or CD16 mutations, then activated with PMA. The surface expression of CD16 on these cells was then examined using flow cytometry. Result: We found 3 mutations effectively blocked CD16 cleavage in cell-based assays, while wild-type CD16 and other CD16 mutations underwent a marked down-regulation of expression. We also demonstrated these 3 mutations were resistant to cleavage when expressed in the human primary NK cells and did not disrupt IgG binding. Conclusion: We developed a non-cleavable CD16 with high affinity to Fc receptor, which can be expressed in engineered NK cells to develop a next generation NK therapy to elicit more potent and durable cytotoxic response. Citation Format: Cuiqing Yang, Fuwei Jiang, Yifang Wang, Tingting Liu, Qingyang Wang, Qin Wang, Gang Ye, Renhong Tang, Zhuoxiao Cao. To develop a next generation NK therapy with deep and durable response by engineered CD16 enhancing NK cytotoxicity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2900.

Population genetic analysis of the Plasmodium falciparum erythrocyte binding antigen-175 (EBA-175) gene in Equatorial Guinea

BackgroundPlasmodium falciparum erythrocyte binding antigen-175 (PfEBA-175) is a candidate antigen for a blood-stage malaria vaccine, while various polymorphisms and dimorphism have prevented to development of effective vaccines based on this gene. This study aimed to investigate the dimorphism of PfEBA-175 on both the Bioko Island and continent of Equatorial Guinea, as well as the genetic polymorphism and natural selection of global PfEBA-175.MethodsThe allelic dimorphism of PfEBA-175 region II of 297 bloods samples from Equatorial Guinea in 2018 and 2019 were investigated by nested polymerase chain reaction and sequencing. Polymorphic characteristics and the effect of natural selection were analyzed using MEGA 7.0, DnaSP 6.0 and PopART programs. Protein function prediction of new amino acid mutation sites was performed using PolyPhen-2 and Foldx program.ResultsBoth Bioko Island and Bata district populations, the frequency of the F-fragment was higher than that of the C-fragment of PfEBA-175 gene. The PfEBA-175 of Bioko Island and Bata district isolates showed a high degree of genetic variability and heterogeneity, with π values of 0.00407 & 0.00411 and Hd values of 0.958 & 0.976 for nucleotide diversity, respectively. The values of Tajima's D of PfEBA-175 on Bata district and Bioko Island were 0.56395 and − 0.27018, respectively. Globally, PfEBA-175 isolates from Asia were more diverse than those from Africa and South America, and genetic differentiation quantified by the fixation index between Asian and South American countries populations was significant (FST > 0.15, P < 0.05). A total of 310 global isolates clustered in 92 haplotypes, and only one cluster contained isolates from three continents. The mutations A34T, K109E, D278Y, K301N, L305V and D329N were predicted as probably damaging.ConclusionsThis study demonstrated that the dimorphism of F-fragment PfEBA-175 was remarkably predominant in the study area. The distribution patterns and genetic diversity of PfEBA-175 in Equatorial Guinea isolates were similar another region isolates. And the levels of recombination events suggested that natural selection and intragenic recombination might be the main drivers of genetic diversity in global PfEBA-175. These results have important reference value for the development of blood-stage malaria vaccine based on this antigen.

Amphipods in estuaries: the sibling species low salinity switch hypothesis

A novel low salinity switch hypothesis is proposed to account for the speciation of an obligate estuarine (oligohaline) amphipod, Orchestia aestuarensis, from a closely-related one, Orchestia mediterranea, found in both estuarine and marine conditions (euryhaline). The underlying genetic mechanisms could involve: 1. A dimorphic allele, or linked set of alleles, carried by the euryhaline amphipod which controls the ability to breed in low salinity conditions in estuaries and which is selected for in these conditions, producing the oligohaline amphipod. 2. A genetically-assimilated gene or genes, controlling the ability to breed in low salinity conditions in estuaries, which is/are “switched on” by low salinity conditions. 3. Allopatric speciation from a euryhaline to an oligohaline amphipod species where low salinity conditions is the selective switch. It is possible that other estuarine, sibling, amphipod pairs have evolved by salinity switching. In the North Atlantic coastal region, this could include: Gammarus tigrinus/G. daiberi and G. salinus/G. zaddachi (Amphipoda, Gammaridae).

Temporal changes in Plasmodium falciparum reticulocyte binding protein homolog 2b (PfRh2b) in Senegal and The Gambia

BackgroundThe Plasmodium falciparum reticulocyte binding protein homolog 2b (PfRh2b) is an important P. falciparum merozoite ligand that mediates invasion of erythrocytes by interacting with a chymotrypsin-sensitive “receptor Z”. A large deletion polymorphism is found in the c-terminal ectodomain of this protein in many countries around the world, resulting in a truncated, but expressed protein. The varying frequencies by region suggest that there could be region specific immune selection at this locus. Therefore, this study was designed to determine temporal changes in the PfRh2b deletion polymorphism in infected individuals from Thiès (Senegal) and Western Gambia (The Gambia). It was also sought to determine the selective pressures acting at this locus and whether prevalence of the deletion in isolates genotyped by a 24-SNP molecular barcode is linked to background genotype or whether there might be independent selection acting at this locus.MethodsInfected blood samples were sourced from archives of previous studies conducted between 2007 and 2013 at SLAP clinic in Thiès and from 1984 to 2013 in Western Gambia by MRC Unit at LSHTM, The Gambia. A total of 1380 samples were screened for the dimorphic alleles of the PfRh2b using semi-nested Polymerase Chain Reaction PCR. Samples from Thiès were previously barcoded.ResultsIn Thiès, a consistent trend of decreasing prevalence of the PfRh2b deletion over time was observed: from 66.54% in 2007 and to 38.1% in 2013. In contrast, in Western Gambia, the frequency of the deletion fluctuated over time; it increased between 1984 and 2005 from (58.04%) to (69.33%) and decreased to 47.47% in 2007. Between 2007 and 2012, the prevalence of this deletion increased significantly from 47.47 to 83.02% and finally declined significantly to 57.94% in 2013. Association between the presence of this deletion and age was found in Thiès, however, not in Western Gambia. For the majority of isolates, the PfRh2b alleles could be tracked with specific 24-SNP barcoded genotype, indicating a lack of independent selection at this locus.ConclusionPfRh2b deletion was found in the two countries with varying prevalence during the study period. However, these temporal and spatial variations could be an obstacle to the implementation of this protein as a potential vaccine candidate.

High-Complexity Plasmodium falciparum Infections, North Central Nigeria, 2015-2018.

The mass migration that occurred during 2009–2013 and after the insurgency in northeastern Nigeria could have increased malaria incidence and Plasmodium falciparum genetic diversity in North Central Nigeria. To determine P. falciparum sequence diversity in this region, we screened 282 samples collected in regional clinics during 2015–2018 for Plasmodium spp. and, with positive samples, determined P. falciparum infection complexity and allele diversity using PCR. Of 34 P. falciparum–positive samples, 39 msp1, 31 msp2, and 13 glurp alleles were detected, and 88% of infections were polyclonal. We identified trimorphic and dimorphic allele combinations in a high percentage of samples, indicative of a high infection complexity in the study population. High genetic diversity is a catalyst for the evolution of drug-resistant alleles. Improved measures (e.g., better drug quality, diagnostics) are needed to control P. falciparum transmission and reduce the potential for the emergence of drug resistance in Nigeria.

Parallel evolution of Batesian mimicry supergene in two Papilio butterflies, P. polytes and P. memnon.

Batesian mimicry protects animals from predators when mimics resemble distasteful models. The female-limited Batesian mimicry in Papilio butterflies is controlled by a supergene locus switching mimetic and nonmimetic forms. In Papilio polytes, recent studies revealed that a highly diversified region (HDR) containing doublesex (dsx-HDR) constitutes the supergene with dimorphic alleles and is likely maintained by a chromosomal inversion. In the closely related Papilio memnon, which exhibits a similar mimicry polymorphism, we performed whole-genome sequence analyses in 11 butterflies, which revealed a nearly identical dsx-HDR containing three genes (dsx, Nach-like, and UXT) with dimorphic sequences strictly associated with the mimetic/nonmimetic phenotypes. In addition, expression of these genes, except that of Nach-like in female hind wings, showed differences correlated with phenotype. The dimorphic dsx-HDR in P. memnon is maintained without a chromosomal inversion, suggesting that a separate mechanism causes and maintains allelic divergence in these genes. More abundant accumulation of transposable elements and repetitive sequences in the dsx-HDR than in other genomic regions may contribute to the suppression of chromosomal recombination. Gene trees for Dsx, Nach-like, and UXT indicated that mimetic alleles evolved independently in the two Papilio species. These results suggest that the genomic region involving the above three genes has repeatedly diverged so that two allelic sequences of this region function as developmental switches for mimicry polymorphism in the two Papilio species. The supergene structures revealed here suggest that independent evolutionary processes with different genetic mechanisms have led to parallel evolution of similar female-limited polymorphisms underlying Batesian mimicry in Papilio butterflies.

Determination of multiple-clone infection at allelic dimorphism site of Plasmodium vivax merozoite surface protein-1 in the Republic of Korea by pyrosequencing assay

Allelic diversity leading to multiple gene polymorphisms of vivax malaria parasites has been shown to greatly contribute to antigenic variation and drug resistance, increasing the potential for multiple-clone infections within the host. Therefore, to identify multiple-clone infections and the predominant haplotype of Plasmodium vivax in a South Korean population, P. vivax merozoite surface protein-1 (PvMSP-1) was analyzed by pyrosequencing. Pyrosequencing of 156 vivax malaria-infected samples yielded 97 (62.18%) output pyrograms showing two main types of peak patterns of the dimorphic allele for threonine and alanine (T1476A). Most of the samples evaluated (88.66%) carried multiple-clone infections (wild- and mutant-types), whereas 11.34% of the same population carried only the mutant-type (1476A). In addition, each allele showed a high frequency of guanine (G) base substitution at both the first and third positions (86.07% and 81.13%, respectively) of the nucleotide combinations. Pyrosequencing of the PvMSP-1 42-kDa fragment revealed a heterogeneous parasite population, with the mutant-type dominant compared to the wild-type. Understanding the genetic diversity and multiple-clone infection rates may lead to improvements in vivax malaria prevention and strategic control plans. Further studies are needed to improve the efficacy of the pyrosequencing assay with large sample sizes and additional nucleotide positions.

Evolution of Fseg/Cseg dimorphism in region III of the Plasmodium falciparum eba-175 gene

The 175-kDa erythrocyte binding antigen (EBA-175) of the malaria parasite Plasmodium falciparum is important for its invasion into human erythrocytes. The primary structure of eba-175 is divided into seven regions, namely I to VII. Region III contains highly divergent dimorphic segments, termed Fseg and Cseg. The allele frequencies of segmental dimorphism within a P. falciparum population have been extensively examined; however, the molecular evolution of segmental dimorphism is not well understood. A comprehensive comparison of nucleotide sequences among 32 P. falciparum eba-175 alleles identified in our previous study, two Plasmodium reichenowi, and one P. gaboni orthologous alleles obtained from the GenBank database was conducted to uncover the origin and evolutionary processes of segmental dimorphism in P. falciparum eba-175. In the eba-175 nucleotide sequence derived from a P. reichenowi CDC strain, both Fseg and Cseg were found in region III, which implies that the original eba-175 gene had both segments, and deletions of F- and C-segments generated Cseg and Fseg alleles, respectively. We also confirmed the presence of allele with Fseg and Cseg in another P. reichenowi strain (SY57), by re-mapping short reads obtained from the GenBank database. On the other hand, the segmental sequence of eba-175 ortholog in P. gaboni was quite diverged from those of the other species, suggesting that the original eba-175 dimorphism of P. falciparum can be traced back to the stem linage of P. falciparum and P. reichenowi. Our findings suggest that Fseg and Cseg alleles are derived from a single eba-175 allele containing both segments in the ancestral population of P. falciparum and P. reichenowi, and that the allelic dimorphism of eba-175 was shaped by the independent emergence of similar dimorphic lineage in different species that has never been observed in any evolutionary mode of allelic dimorphism at other loci in malaria genomes.

The pdm3 Locus Is a Hotspot for Recurrent Evolution of Female-Limited Color Dimorphism in Drosophila

Sex-limited polymorphisms are an intriguing form of sexual dimorphism that offer unique opportunities to reconstruct the evolutionary changes that decouple male and female traits encoded by a shared genome. We investigated the genetic basis of a Mendelian female-limited color dimorphism (FLCD) that segregates in natural populations of more than 20 species of the Drosophila montium subgroup. In these species, females have alternative abdominal color morphs, light and dark, whereas males have only one color morph in each species. A comprehensive molecular phylogeny of the montium subgroup supports multiple origins of FLCD. Despite this, we mapped FLCD to the same locus in four distantly related species-the transcription factor POU domain motif3 (pdm3), which acts as a repressor of abdominal pigmentation in D.melanogaster. In D.serrata, FLCD maps to a structural variant in the first intron of pdm3; however, this variant is not found in the three other species-D.kikkawai, D.leontia, and D.burlai-and sequence analysis strongly suggests the pdm3 alleles responsible for FLCD originated independently at least three times. We propose that cis-regulatory changes in pdm3 form sexually dimorphic and monomorphic alleles that segregate within species and are preserved, at least in one species, by structural variation. Surprisingly, pdm3 has not been implicated in the evolution of sex-specific pigmentation outside the montium subgroup, suggesting that the genetic paths to sexual dimorphism may be constrained within a clade but variable across clades.

Merozoite surface protein-1 genetic diversity in Plasmodium malariae and Plasmodium brasilianum from Brazil.

BackgroundThe merozoite surface protein 1 (MSP1) gene encodes the major surface antigen of invasive forms of the Plasmodium erythrocytic stages and is considered a candidate vaccine antigen against malaria. Due to its polymorphisms, MSP1 is also useful for strain discrimination and consists of a good genetic marker. Sequence diversity in MSP1 has been analyzed in field isolates of three human parasites: P. falciparum, P. vivax, and P. ovale. However, the extent of variation in another human parasite, P. malariae, remains unknown. This parasite shows widespread, uneven distribution in tropical and subtropical regions throughout South America, Asia, and Africa. Interestingly, it is genetically indistinguishable from P. brasilianum, a parasite known to infect New World monkeys in Central and South America.MethodsSpecific fragments (1 to 5) covering 60 % of the MSP1 gene (mainly the putatively polymorphic regions), were amplified by PCR in isolates of P. malariae and P. brasilianum from different geographic origin and hosts. Sequencing of the PCR-amplified products or cloned PCR fragments was performed and the sequences were used to construct a phylogenetic tree by the maximum likelihood method. Data were computed to give insights into the evolutionary and phylogenetic relationships of these parasites.ResultsExcept for fragment 4, sequences from all other fragments consisted of unpublished sequences. The most polymorphic gene region was fragment 2, and in samples where this region lacks polymorphism, all other regions are also identical. The low variability of the P. malariae msp1 sequences of these isolates and the identification of the same haplotype in those collected many years apart at different locations is compatible with a low transmission rate. We also found greater diversity among P. brasilianum isolates compared with P. malariae ones. Lastly, the sequences were segregated according to their geographic origins and hosts, showing a strong genetic and geographic structure.ConclusionsOur data show that there is a low level of sequence diversity and a possible absence of allelic dimorphism of MSP1 in these parasites as opposed to other Plasmodium species. P. brasilianum strains apparently show greater divergence in comparison to P. malariae, thus P. malariae could derive from P. brasilianum, as it has been proposed.Electronic supplementary materialThe online version of this article (doi:10.1186/s12879-015-1238-8) contains supplementary material, which is available to authorized users.

The Plasmodium gaboni genome illuminates allelic dimorphism of immunologically important surface antigens in P. falciparum

In the deadly human malaria parasite Plasmodium falciparum, several major merozoite surface proteins (MSPs) show a striking pattern of allelic diversity called allelic dimorphism (AD). In AD, the vast majority of observed alleles fall into two highly divergent allelic classes, with recombinant alleles being rare or not observed, presumably due to repression by natural selection (recombination suppression, or RS). The three AD loci, merozoite surface proteins (MSPs) 1, 2, and 6, along with MSP3, which also exhibits RS among four allelic classes, can be collectively called AD/RS. The causes of AD/RS and the evolutionary history of allelic diversity at these loci remain mysterious. The few available sequences from a single closely related chimpanzee parasite, P. reichenowi, have suggested that for 3/4 loci, AD/RS is an ancient state that has been retained in P. falciparum since well before the P. falciparum–P. reichenowi ancestor. On the other hand, based on comparative sequence analysis, we recently suggested that (i) AD/RS P. falciparum loci have undergone interallelic recombination over longer evolutionary times (on the timescale of recent speciation events), and thus (ii) AD/RS may be a recent phenomenon. The recent publication of genomic sequencing efforts for P. gaboni, an outgroup to P. falciparum and P. reichenowi, allows for improved reconstruction of the evolutionary history of these loci. In this work, I report genic sequence for P. gaboni for all four AD/RS P. falciparum loci (MSP1, 2, 3, and 6). Comparison of these sequences with available P. falciparum and P. reichenowi data strengthens the evidence for interallelic recombination over the evolutionary history of these species and also strengthens the case that AD/RS at these loci is ancient. Combined with previous results, these data provide evidence that AD/RS at different loci has evolved at several different times in the evolutionary history of P. falciparum: (i) before the P. gaboni–P. falciparum divergence, for much of MSP1 and MSP3; (ii) between the P. gaboni–P. falciparum and P. reichenowi–P. falciparum divergences, for the 5′ end of the AD region of MSP6 and block 3 of MSP1; (iii) near the P. reichenowi–P. falciparum divergence, for the 3′ end of the AD region of MSP6; and (iv) after the P. reichenowi–P. falciparum divergence, for MSP2. Based on these results, I suggest a new hypothesis for long-term evolutionary maintenance of AD/RS by recombination within allelic groups.

A new model for the origins of allelic dimorphism in Plasmodium falciparum

In his landmark 1987 study of the merozoite surface protein-1 locus in Plasmodium falciparum, Kazuyuki Tanabe and coauthors introduced the phenomenon of allelic dimorphism, in which antigenic diversity is arranged into two maximally diverged haplotypes. Further work has extended this finding to other loci in P. falciparum. Each of the loci at which allelic dimorphism is observed encodes major surface antigens of blood-stage malaria parasites, and is consequently a major vaccine target, thus understanding the origins and implications of allelic dimorphism is of crucial importance. Here we examine the essential features of allelic dimorphism in dimorphic malarial surface antigens. From sequence analysis, we conclude that the ancestral population may have been recombining/multimorphic rather than dimorphic. We hypothesize a pathway to allelic dimorphism in which an ancestral allele-rich recombining population could have undergone a severe population bottleneck, putatively caused by the lateral transfer of P. falciparum from apes to humans. This bottleneck produced a reduction in allelic diversity, favoring the survival of the most divergent alleles, which in turn led to recombination suppression by strong natural selection against recombinants.

Multiple episodic evolution events in V1R receptor genes of East-African cichlids.

Fish use olfaction to detect a variety of nonvolatile chemical signals, and thus, this sense is key to survival and communication. However, the contribution of the olfactory sense to social—especially reproductive—interactions in cichlids is still controversial. To obtain insights into this issue, we investigated the genes encoding V1Rs—possible candidates for reproductive pheromone receptors—among East-African cichlids. Interestingly, we found an excess of nonsynonymous over synonymous substitutions in four of six V1R genes in multiple cichlid lineages. First, we found that highly dimorphic V1R2 allele groups were shared among the cichlids inhabiting all East-African Great Lakes emerged through the episodic accumulation of the nonsynonymous substitutions prior to the radiation of the Lake Tanganyika species flock. We further detected such episodic events in V1R1 of the tribe Tropheini, and in V1R3 and V1R6 of the tribe Trematocarini. The excess of nonsynonymous substitutions in these examples were indicated as dN/dS > 1, which were all statistically significant by Fisher’s exact test. Furthermore, we speculate that the amino acid changes in these episodic events are likely functional switch because they occurred in the putative ligand-binding pocket. Our finding of the occurrence of multiple episodic events and the unexpected gene diversity in one unique gene family is suggestive of the contribution of the V1R to the species diversification and the social interaction in cichlids.

Genotyping of Erythrocyte Binding Antigen 175 KD (eba-175) Alleles of Plasmodium Falciparum-Malaria Using Nested-PCR in South-East of Iran

The Erythrocyte Binding Antigen-175 (EBA-175) on Plasmodium falciparum (P.falciparum) merozoites mediates sialic acid dependent binding to glycophorin a on host erythrocytes and, therefore, plays a crucial role in cell invasion. Dimorphic allele segments have been found in its encoding gene with in FCR-3 strains (F-segment) and CAMP strains (C-segment). This study was designed to determine the distribution of EBA-175 alleles of P.falciparum in the South-East of Iran. Nested Polymerase Chain Reaction method used with specific primers, which improves the two fragments of the EBA-175 gene. Ninety-four microscopically positive blood samples were collected from the infected P.falciparum malaria patients from four different districts. The 88 out of 94 confirmed P.falciparum samples were successfully scored for EBA-175. The allelic genotyping exhibited CAMP strains (714 bp) and FCR-3 strains (795 bp) in 31(32.97%) and 49(52.12%) cases, respectively and 8 cases (8.51%) showed mixed allelic CAMP/FCR-3 infections. The two fragments of dimorphic EBA-175 gene were observed and the FCR-3 allele was more prevalent in South-East of Iran. This distributional pattern should be considered in designing to control P. falciparum malaria in the region.

Hybridization between previously isolated ancestors may explain the persistence of exactly two ancient lineages in the genome of the oyster parasite Perkinsus marinus

Theory predicts that neutral genetic variation accumulates within populations to a level determined by gains through mutation and losses by genetic drift. This balance results in a characteristic distribution of allelic variation with the maximum allelic difference determined by effective population size. Here, we report a striking departure from these expectations in the form of allelic dimorphism, observed at the majority of seven loci examined in Perkinsus marinus, an important oyster parasite that causes Dermo disease. DNA sequences were collected from five loci flanking microsatellite repeats and two loci coding for superoxide dismutase enzymes that may mediate the parasite’s interaction with its host. Based on 474 sequences, sampled across 5000km of the eastern United States coastline, no more than two alleles were observed at each locus (discounting singletons). Depending on the locus, the common allele ranged in overall frequency from 72% to 92%. At each locus the two alleles differed substantially (3.8% sequence difference, on average), and the among-locus variance in divergences was not sufficient to reject a simultaneous origin for all dimorphisms using approximate Bayesian methods. Dimorphic alleles were estimated to have diverged from a common ancestral allele at least 0.9 million years ago. Across these seven loci, only five other alleles were ever observed, always as singletons and differing from the dimorphic alleles by no more than two nucleotides. Free recombination could potentially have shuffled these dimorphisms into as many as 243 multilocus combinations, but the existence of only ten combinations among all samples strongly supports low recombination frequencies and is consistent with the observed absence of intragenic recombination. We consider several demographic and evolutionary hypotheses to explain these patterns. Few can be conclusively rejected with the present data, but we advance a recent hybridization of ancient divergent lineages scenario as the most parsimonious.

Distribution of erythrocyte binding antigen 175 (EBA-175) gene dimorphic alleles in Plasmodium falciparum field isolates from Sudan.

BackgroundThe Erythrocyte Binding Antigen (EBA) 175 has been considered as one of the most important Plasmodium falciparum (P. falciparum) merozoite ligands that mediate invasion of the erythrocytes through their sialated receptor: Glycophorin A (GPA). The effect of the EBA 175 dimorphic alleles (F and C) on the severity of the disease is not yet fully understood. Therefore this study was designed to assess the distribution of the divergent dimorphic alleles of P. falciparum EBA-175 (F and C) in three different geographical areas in Sudan and the possible association of this dimorphism with the severity of the disease.MethodsA sum of 339 field isolates of P. falciparum obtained from patients in three different geographical areas in Sudan were screened for the dimorphic alleles (F, C) of the EBA-175 using nested PCR.ResultsThe percentage of F, C, and mixed F/C alleles were; 41%, 51%, and 8% respectively. F and C alleles showed significantly different distributions in the various geographic areas (p = 0.00). There was no significant association between malaria clinical manifestation and P. falciparum EBA-175 F and C alleles frequencies.ConclusionsThis study showed a significant differential distribution of F and C alleles in different geographical malaria endemic areas. No significant association was observed between F and C alleles and different malaria phenotypes.

Dimorphisms of the proteasome subunit beta type 8 gene (PSMB8) of ectothermic tetrapods originated in multiple independent evolutionary events

The proteasome subunit beta type 8 gene (PSMB8) encodes one of the beta subunits of the immunoproteasome responsible for the generation of peptides presented by major histocompatibility complex class I molecules. Dimorphic alleles of the PSMB8 gene, termed A and F types, based on the deduced 31st amino acid residue of the mature protein have been reported from various vertebrates. Phylogenetic analysis revealed the presence of dichotomous ancient lineages, one comprising the F-type PSMB8 of basal ray-finned fishes, and the other comprising the A-type PSMB8 of these animals and both the F- and A-type PSMB8 of Xenopus and acanthopterygians, indicating that evolutionary history of the PSMB8 dimorphism was not straightforward. We analyzed the PSMB8 gene of five reptile and one amphibian species and found both the A and F types from all six. Phylogenetic analysis indicated that the PSMB8 F type was apparently regenerated from the PSMB8 A type at least five times independently during tetrapod evolution. Genomic typing of wild individuals of geckos and newts indicated that the frequencies of the A- and F-type alleles are not highly biased in these species. Phylogenetic analysis of each exon of the reptile PSMB8 gene suggested interallelic sequence homogenization as a possible evolutionary mechanism for the apparent recurrent regeneration of PSMB8 dimorphism in tetrapods. An extremely strong balancing selection acting on PSMB8 dimorphism was implicated in an unprecedented pattern of allele evolution.

Long-Lived Dichotomous Lineages of the Proteasome Subunit Beta Type 8 (PSMB8) Gene Surviving More than 500 Million Years as Alleles or Paralogs

On an evolutionary time scale, polymorphic alleles are believed to have a short life, persisting at most tens of millions of years even under long-term balancing selection. Here, we report highly diverged trans-species dimorphism of the proteasome subunit beta type 8 (PSMB8) gene, which encodes a catalytic subunit of the immunoproteasome responsible for the generation of peptides presented by major histocompatibility complex (MHC) class I molecules, in lower teleosts including Cypriniformes (zebrafish and loach) and Salmoniformes (trout and salmon), whose last common ancestor dates to 300 Ma. Moreover, phylogenetic analyses indicated that these dimorphic alleles share lineages with two shark paralogous genes, suggesting that these two lineages have been maintained for more than 500 My either as alleles or as paralogs, and that conversion between alleles and paralogs has occurred at least once during vertebrate evolution. Two lineages termed PSMB8A and PSMB8F show an A(31)F substitution that would probably affect their cleaving specificity, and whereas the PSMB8A lineage has been retained by all analyzed jawed vertebrates, the PSMB8F lineage has been lost by most jawed vertebrates except for cartilaginous fish and basal teleosts. However, a possible functional equivalent of the PSMB8F lineage has been revived as alleles within the PSMB8A lineage at least twice during vertebrate evolution in the amphibian Xenopus and teleostean Oryzias species. Dynamic evolution of the PSMB8 polymorphism through long-term persistence, loss, and regaining of dimorphism and conversion between alleles and paralogs implies the presence of strong selective pressure for functional polymorphism of this gene.

Plasmodium falciparum merozoite surface protein 1 blocks the proinflammatory protein S100P

The malaria parasite, Plasmodium falciparum, and the human immune system have coevolved to ensure that the parasite is not eliminated and reinfection is not resisted. This relationship is likely mediated through a myriad of host-parasite interactions, although surprisingly few such interactions have been identified. Here we show that the 33-kDa fragment of P. falciparum merozoite surface protein 1 (MSP1(33)), an abundant protein that is shed during red blood cell invasion, binds to the proinflammatory protein, S100P. MSP1(33) blocks S100P-induced NFκB activation in monocytes and chemotaxis in neutrophils. Remarkably, S100P binds to both dimorphic alleles of MSP1, estimated to have diverged >27 Mya, suggesting an ancient, conserved relationship between these parasite and host proteins that may serve to attenuate potentially damaging inflammatory responses.

Highly divergent dimorphic alleles of the proteasome subunit beta type-8 (PSMB8) gene of the bichir Polypterus senegalus: implication for evolution of the PSMB8 gene of jawed vertebrates

The proteasome subunit beta type-8 (PSMB8) gene encodes a catalytic subunit of the immunoproteasome, which is involved in the generation of peptides presented by MHC class I molecules. To date, highly diverged dichotomous alleles of PSMB8 have been reported in Oryzias species (actinopterygian teleosts) and Xenopus species (sarcopterygian amphibians). These dimorphic alleles share a similar substitution (A/V(31)F/Y) at the 31st position of the mature protein, which is most probably involved in formation of the S1 pocket. This substitution likely confers different cleavage specificities on the dimorphic PSMB8s. In addition, two paralogous PSMB8 genes possessing the A and F residues at the 31st position have been reported in sharks. Phylogenetic analysis indicated that the two types of PSMB8 of Oryzias, Xenopus, and sharks arose by independent evolutionary events. Here, we identified another pair of dimorphic alleles of PSMB8, which have the A and F residues at the 31st position of the mature protein, from bichir, Polypterus senegalus, a basal actinopterygian. The sequences of the mature proteins-encoding region of the dimorphic alleles of bichir PSMB8, the A and F types, showed only 72.7% and 77.5% identities at the nucleotide and the deduced amino acid levels, respectively. Their intronic sequences show almost no similarity, indicating that the dimorphic alleles of bichir PSMB8 have a very ancient origin. However, phylogenetic analysis showed that the dimorphisms of PSMB8 of bichir, Xenopus, and Oryzias arose by independent evolutionary events, suggesting the presence of a strong selective pressure for possessing the dimorphism.