Epigenetic switches in<i>clag3</i>genes mediate blasticidin S resistance in malaria parasites

Mira-Martínez, Sofía; Rovira-Graells, Núria; Crowley, Valerie M.; Altenhofen, Lindsey M.; Llinás, Manuel; Cortés, Alfred

doi:10.1111/cmi.12162

Cited by 56 publications

(129 citation statements)

References 37 publications

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“…in that they form a part of a transporter in the erythrocyte membrane (31). Again there is evidence that they may differ in function from the mutational differences between the copies (32).…”

Section: Discussionmentioning

confidence: 99%

Role of Plasmodium vivax Duffy-binding protein 1 in invasion of Duffy-null Africans

Gunalan

Hostetler

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

121

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The ability of the malaria parasite Plasmodium vivax to invade erythrocytes is dependent on the expression of the Duffy blood group antigen on erythrocytes. Consequently, Africans who are null for the Duffy antigen are not susceptible to P. vivax infections. Recently, P. vivax infections in Duffy-null Africans have been documented, raising the possibility that P. vivax, a virulent pathogen in other parts of the world, may expand malarial disease in Africa. P. vivax binds the Duffy blood group antigen through its Duffy-binding protein 1 (DBP1). To determine if mutations in DBP1 resulted in the ability of P. vivax to bind Duffy-null erythrocytes, we analyzed P. vivax parasites obtained from two Duffy-null individuals living in Ethiopia where Duffy-null and -positive Africans live side-by-side. We determined that, although the DBP1s from these parasites contained unique sequences, they failed to bind Duffy-null erythrocytes, indicating that mutations in DBP1 did not account for the ability of P. vivax to infect Duffy-null Africans. However, an unusual DNA expansion of DBP1 (three and eight copies) in the two Duffy-null P. vivax infections suggests that an expansion of DBP1 may have been selected to allow low-affinity binding to another receptor on Duffy-null erythrocytes. Indeed, we show that Salvador (Sal) I P. vivax infects Squirrel monkeys independently of DBP1 binding to Squirrel monkey erythrocytes. We conclude that P. vivax Sal I and perhaps P. vivax in Duffy-null patients may have adapted to use new ligand-receptor pairs for invasion.Plasmodium vivax | Duffy blood group antigen | Duffy-binding protein | DNA expansionI n 1975 we identified the failure of Plasmodium knowlesi to invade Duffy-null erythrocytes (1). We assumed that the Duffy blood group null phenotype, a common African phenotype, conferred resistance in Africans to P. vivax, a Plasmodium closely related to P. knowlesi. Subsequent studies demonstrated that African American volunteers who were Duffy-null were resistant to mosquito-transmitted P. vivax (2). In addition, African American soldiers in Vietnam who were infected with P. vivax were all Duffypositive (3). Furthermore, African Americans in a village in Honduras who were infected with P. vivax were all Duffy-positive whereas those infected with P. falciparum were both Duffy-null and -positive (4). We concluded that Duffy null was the basis of resistance to P. vivax by Africans.The molecular basis of Duffy null was a single point mutation in the GATA1-binding sequence in the promotor region 5′ to the Duffy blood group ORF (Fig. 1B) that led to Duffy-blood-groupnull erythrocytes (5). In vitro studies with P. knowlesi demonstrated that the invasive merozoites were able to bind and reorient apically with Duffy-null erythrocytes but could not form a junction as occurred in Duffy-positive erythrocytes, indicating that the Duffy blood group was required for P. vivax invasion (6). Later, in P. knowlesi parasite culture supernatants, the parasite ligand binding to Duffy blood group antigen was identi...

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Section: Discussionmentioning

confidence: 99%

Role of Plasmodium vivax Duffy-binding protein 1 in invasion of Duffy-null Africans

Gunalan

Hostetler

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

121

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“…Evidence has included genetic mapping with the small-molecule PSAC inhibitor ISPA-28 and quantitative effects of externally applied proteases (9,16), selection of recombinant parasites cultivated in nutrient-restricted media (10), and molecular studies to characterize blasticidin S-resistant parasites (11,24). Although these multiple lines of evidence implicate this gene family in nutrient uptake, the precise structural contribution of the encoded CLAG3 protein is uncertain.…”

Section: Discussionmentioning

confidence: 99%

“…Alternatively, CLAG3 may have only a modulatory role, such as through enzymatic activation of quiescent channels in the host membrane (19). Distinguishing between these models is an important step in understanding the structural basis of solute and nutrient transport at the host membrane and in determining whether the channel can be targeted for antimalarial drug development (10,20).One way to address this uncertainty involves molecular studies with available PSAC mutants that have been generated through in vitro selection with toxins that require channel-mediated uptake (11,(21)(22)(23)(24). A leupeptin-resistant clone, HB3-leuR1, carries a nonsynonymous mutation in the clag3.2 gene.…”

mentioning

confidence: 99%

“…One way to address this uncertainty involves molecular studies with available PSAC mutants that have been generated through in vitro selection with toxins that require channel-mediated uptake (11,(21)(22)(23)(24). A leupeptin-resistant clone, HB3-leuR1, carries a nonsynonymous mutation in the clag3.2 gene.…”

mentioning

confidence: 99%

“…The clag multigene family, also conserved in and restricted to malaria parasites (8), has recently been linked to PSAC activity (9)(10)(11). Two paralogs on parasite chromosome 3, known as clag3.1 and clag3.2, appear to play a critical role, as identified by genetic mapping experiments with ISPA-28, an isolate-specific PSAC antagonist that blocks channels on the Dd2 laboratory clone but is ineffective against channels from other lines.…”

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confidence: 99%

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

et al. 2015

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Erythrocytes infected with malaria parasites have increased permeability to ions and nutrients, as mediated by the plasmodial surface anion channel (PSAC) and recently linked to parasite clag3 genes. Although the encoded protein is integral to the host membrane, its precise contribution to solute transport remains unclear because it lacks conventional transmembrane domains and does not have homology to ion channel proteins in other organisms. Here, we identified a probable CLAG3 transmembrane domain adjacent to a variant extracellular motif. Helical-wheel analysis revealed strict segregation of polar and hydrophobic residues to opposite faces of a predicted ␣-helical transmembrane domain, suggesting that the domain lines a water-filled pore. A single CLAG3 mutation (A1210T) in a leupeptin-resistant PSAC mutant falls within this transmembrane domain and may affect pore structure. Allelic-exchange transfection and site-directed mutagenesis revealed that this mutation alters solute selectivity in the channel. The A1210T mutation also reduces the blocking affinity of PSAC inhibitors that bind on opposite channel faces, consistent with global changes in channel structure. Transfected parasites carrying this mutation survived a leupeptin challenge significantly better than a transfection control did. Thus, the A1210T mutation contributes directly to both altered PSAC activity and leupeptin resistance. These findings reveal the molecular basis of a novel antimalarial drug resistance mechanism, provide a framework for determining the channel's composition and structure, and should guide the development of therapies targeting the PSAC.T he human malaria parasite Plasmodium falciparum remodels its host erythrocyte by exporting many proteins, generating membranous structures in the host cytosol, and increasing erythrocyte permeability to many solutes. Studies by multiple groups have determined that anions, sugars, purines, organic cations, and some vitamins have increased permeability after infection (1-4). The increase in permeability is primarily mediated by a parasitederived ion and nutrient channel known as the plasmodial surface anion channel (PSAC) (5). Importantly, both PSAC single-channel properties and the relative increases in solute permeabilities are conserved in divergent malaria parasites (6). Because Babesia parasites do not induce PSAC-like activity in erythrocytes that they invade (7), this channel is thought to be restricted to the genus Plasmodium.The clag multigene family, also conserved in and restricted to malaria parasites (8), has recently been linked to PSAC activity (9-11). Two paralogs on parasite chromosome 3, known as clag3.1 and clag3.2, appear to play a critical role, as identified by genetic mapping experiments with ISPA-28, an isolate-specific PSAC antagonist that blocks channels on the Dd2 laboratory clone but is ineffective against channels from other lines. These genes undergo epigenetic switching to encode a single protein that is initially packaged in specialized organelles known as rh...

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Epigenetic Gene Regulation: Key to Development and Survival of Malaria Parasites

Fraschka

Bártfai

2016

Comprehensive Analysis of Parasite Biology: From Metabolism to Drug Discovery

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Epigenetic switches inclag3genes mediate blasticidin S resistance in malaria parasites

Cited by 56 publications

References 37 publications

Role of Plasmodium vivax Duffy-binding protein 1 in invasion of Duffy-null Africans

Role of Plasmodium vivax Duffy-binding protein 1 in invasion of Duffy-null Africans

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

Epigenetic Gene Regulation: Key to Development and Survival of Malaria Parasites

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