{"gene":"PF3D7_1335400","run_date":"2026-09-08T14:19:06","timeline":{"discoveries":[{"year":2003,"finding":"Targeted gene disruption of PfRh2a in P. falciparum showed that unlike PfRh2b, PfRh2a does not mediate the novel sialic acid-independent, trypsin-resistant, chymotrypsin-sensitive invasion pathway, and phenotypic variation (variant expression) of the PfRh protein family allows exploitation of different erythrocyte surface receptors.","method":"Targeted gene disruption (knockout) with erythrocyte invasion assays using enzyme-treated RBCs","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean targeted gene disruption with defined invasion pathway phenotype, replicated across multiple parasite lines","pmids":["12606570"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2001,"finding":"RH2a (PF3D7_1335400) encodes a >300 kDa protein expressed in late schizonts and localized to the apical end of the merozoite; no binding to mature RBCs or reticulocyte-enriched cell preparations was detected in this study, and antibodies to the protein can inhibit merozoite invasion of RBCs.","method":"Protein expression analysis, immunofluorescence localization, erythrocyte binding assay, invasion inhibition assay with antibodies","journal":"Infection and immunity","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization and binding assay, single lab, multiple methods; binding result was negative","pmids":["11160005"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2002,"finding":"RH2a protein is expressed in late schizonts and merozoites and is located in apical organelles and on the apical surface; expression of RH2a is variably transcribed and translated across parasite lines (absent in FCB1), indicating phenotypic variation at the protein level.","method":"Western blot, immunofluorescence, RT-PCR across multiple parasite lines","journal":"Infection and immunity","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization by immunofluorescence, protein expression confirmed by western blot, multiple parasite lines tested in one study","pmids":["12228308"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":"3D7, T996, FCB1"}]},{"year":2006,"finding":"PfROM4 (a P. falciparum rhomboid intramembrane protease) cleaves RH2a in a heterologous processing assay, with cleavage dependent on the RH2a transmembrane domain; PfROM1 did not cleave RH2a efficiently in this assay.","method":"Heterologous rhomboid cleavage assay with substrate transmembrane domain swap experiments","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — enzymatic assay with domain swaps establishing substrate specificity, but performed in heterologous system (single lab)","pmids":["17040128"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2009,"finding":"Targeted gene disruption of PfRh2a and PfRh2b separately in W2mef parasites showed that both PfRh2b and PfRh2a contribute to efficient sialic acid-independent erythrocyte invasion, and there is strong selective pressure for reconstitution of PfRh2b expression at the expense of PfRh2a expression.","method":"Targeted gene disruption (knockout) of PfRh2a and PfRh2b individually with invasion assays using sialic acid-depleted erythrocytes","journal":"Molecular microbiology","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal gene disruptions with defined quantitative invasion phenotype, selective pressure documented in W2mef background","pmids":["19400777"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":"W2mef"}]},{"year":2010,"finding":"The differential ability of PfRh2a vs. PfRh2b to specify alternative erythrocyte invasion pathways is conferred by their divergent cytoplasmic domains, not by the ectodomain or transmembrane regions, as shown using viable PfRh2a/2b chimeric parasites.","method":"Generation of chimeric parasites (domain-swap constructs of PfRh2a and PfRh2b) with invasion pathway assays","journal":"Molecular microbiology","confidence":"High","confidence_rationale":"Tier 2 / Strong — domain-swap chimeric parasite approach directly maps functional diversification to cytoplasmic domain, clean genetic experiment with defined phenotypic readout","pmids":["20487292"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2010,"finding":"Loss of function of EBA-181 ablates the ability of PfRh2a/b antibodies to inhibit merozoite invasion, providing evidence that PfRh and EBL proteins functionally interact and work cooperatively during merozoite invasion.","method":"Single and double gene knockouts of eba genes combined with antibody invasion inhibition assays (growth inhibition assay)","journal":"Infection and immunity","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — epistasis via gene knockouts with functional antibody inhibition readout, single lab","pmids":["21149582"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2011,"finding":"Native PfRH2a/2b is proteolytically processed near the N-terminus yielding a 220 kDa fragment (sialic acid-independent, trypsin-resistant, chymotrypsin-sensitive erythrocyte binding) and an 80 kDa fragment (sialic acid-dependent, trypsin-resistant, chymotrypsin-sensitive erythrocyte binding); the erythrocyte binding domain was mapped to a conserved 40 kDa N-terminal region common to both PfRH2a and PfRH2b, and recombinant protein of this domain blocks sialic acid-independent invasion.","method":"Western blot of native protein processing, erythrocyte binding assays with enzyme-treated RBCs, recombinant protein binding, invasion inhibition with antibodies","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods (binding assay, inhibition assay, recombinant domain mapping), single lab","pmids":["21386888"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2011,"finding":"PfRh2a undergoes proteolytic cleavage events in the transmembrane region during invasion consistent with PfROM4 activity, releasing the ectodomain; PfRh2a and PfRh2b bind red blood cells and the erythrocyte-binding domain was defined to a 15 kDa N-terminal region of each protein; antibodies to this region block merozoite invasion.","method":"Proteolytic processing analysis (western blot), erythrocyte binding assay, antibody invasion inhibition assay","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct binding domain mapping, proteolysis characterization, and functional antibody blocking in single study, single lab","pmids":["21698217"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2011,"finding":"Using monoclonal antibodies against a unique region of PfRH2a, PfRH2a was shown to translocate from the rhoptry neck to the moving junction during merozoite invasion, independent of the invasion pathway used, suggesting a function beyond receptor binding; PfRH2a is processed both in the schizont and during invasion, producing fragments with different erythrocyte binding properties.","method":"Live and fixed immunofluorescence with anti-PfRH2a monoclonal antibodies, erythrocyte binding assays with processed fragments","journal":"Infection and immunity","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization during live invasion with monoclonal antibodies, pathway-independent movement established, single lab","pmids":["21628513"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2012,"finding":"Aldolase does NOT bind the cytoplasmic tail of RH2a (only RH2b binds aldolase among the two paralogs), indicating RH2a lacks the aromatic amino acid context required for connection to the actin-myosin motor via aldolase; this is consistent with the functional distinction that RH2b but not RH2a is required for erythrocyte invasion.","method":"Peptide binding assay, surface plasmon resonance (SPR), X-ray crystal structure of aldolase with RH2b cytoplasmic peptide","journal":"mBio","confidence":"High","confidence_rationale":"Tier 1 / Strong — negative result for RH2a confirmed by SPR and structural data with RH2b as comparator; mechanistically informative negative finding with rigorous biochemical methods","pmids":["22991428"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2013,"finding":"RH2a (as a member of the reticulocyte binding-like family) binds heparin, localizing to the apical tip of the merozoite surface; heparin affinity chromatography followed by immunoblot confirmed heparin binding, suggesting heparin masks the apical surface and blocks erythrocyte interaction.","method":"Heparin affinity chromatography, immunoblot with ligand-specific antibodies, immunofluorescence localization","journal":"Scientific reports","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single pulldown/affinity chromatography experiment, RH2a identified among multiple proteins without specific functional follow-up for this protein alone","pmids":["24212193"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2018,"finding":"Forward genetic linkage analysis of experimental genetic crosses identified the PfRh2a/PfRh2b locus as the single major determinant of phenotypic variation in two erythrocyte invasion pathways; CRISPR-Cas9 editing confirmed that variation in PfRh2a and PfRh2b expression specifies alternative invasion pathway use.","method":"Forward genetic screen, whole genome sequencing linkage analysis of genetic cross progeny, CRISPR-Cas9 gene editing, invasion pathway phenotyping","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 2 / Strong — forward genetic screen confirmed by CRISPR-Cas9 editing with quantitative invasion phenotyping, two orthogonal genetic approaches","pmids":["30496294"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2025,"finding":"Disruption of pfrh2a in P. falciparum had a less prominent effect on susceptibility to human inhibitory antibodies compared to loss of PfRH1 or PfRH2b, suggesting PfRH2a is a minor target of acquired growth-inhibitory antibodies and plays a lesser role in immune evasion through phenotypic variation relative to PfRH1 and PfRH2b.","method":"Gene disruption (pfrh2a knockout) parasites challenged with human immune sera in invasion inhibition assays","journal":"Frontiers in immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct gene disruption with functional invasion inhibition readout using human sera, single study","pmids":["40201183"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"current_model":"RH2a (PF3D7_1335400) is a >300 kDa apical merozoite protein that is proteolytically processed (including by PfROM4 at its transmembrane region) to yield fragments with differential erythrocyte binding specificities; its conserved 15–40 kDa N-terminal ectodomain mediates sialic acid-independent, chymotrypsin-sensitive erythrocyte binding, while its divergent cytoplasmic domain (which does not bind aldolase) is responsible for its functional distinction from the paralogous PfRh2b; RH2a translocates from the rhoptry neck to the moving junction during invasion in a pathway-independent manner, cooperates with EBL proteins and PfRh2b for efficient sialic acid-independent invasion, and its variant expression — controlled at the PfRh2a/PfRh2b locus as the primary determinant of invasion pathway phenotype — contributes to evasion of host immunity."},"narrative":{"mechanistic_narrative":"RH2a (PF3D7_1335400) is a >300 kDa apical merozoite protein of the reticulocyte binding-like family that participates in erythrocyte invasion by Plasmodium falciparum, contributing to the selection of alternative invasion pathways [PMID:12606570, PMID:30496294]. It is expressed in late schizonts and merozoites and localizes to apical organelles and the apical merozoite surface, with antibodies against it inhibiting invasion [PMID:11160005, PMID:12228308]. Unlike its paralog PfRh2b, disruption of RH2a does not by itself specify the sialic acid-independent, trypsin-resistant, chymotrypsin-sensitive invasion pathway, but both paralogs contribute to efficient sialic acid-independent invasion, and their variant expression at the PfRh2a/PfRh2b locus is the single major genetic determinant of invasion-pathway phenotype [PMID:12606570, PMID:19400777, PMID:30496294]. The two paralogs share a conserved N-terminal ectodomain — mapped to a 15–40 kDa region — that mediates erythrocyte binding, while their functional divergence is conferred by their divergent cytoplasmic domains rather than the ectodomain or transmembrane region [PMID:20487292, PMID:21386888, PMID:21698217]. RH2a is proteolytically processed both in the schizont and during invasion, including transmembrane cleavage consistent with the rhomboid protease PfROM4, yielding fragments with distinct erythrocyte binding specificities, and it translocates from the rhoptry neck to the moving junction during invasion independently of the pathway used [PMID:17040128, PMID:21698217, PMID:21628513]. The RH2a cytoplasmic tail does not bind aldolase — in contrast to RH2b — explaining why RH2a is not directly coupled to the actin-myosin motor and is functionally distinct from RH2b [PMID:22991428]. Relative to PfRH1 and PfRh2b, RH2a is a minor target of acquired human growth-inhibitory antibodies [PMID:40201183].","teleology":[{"year":2001,"claim":"Establishing where and when RH2a acts was the first requirement; its identification as a large apical schizont/merozoite protein whose antibodies block invasion placed it at the invasion interface.","evidence":"Protein expression analysis, immunofluorescence, erythrocyte binding and antibody invasion-inhibition assays in P. falciparum","pmids":["11160005"],"confidence":"Medium","gaps":["No direct erythrocyte binding detected in this assay","Receptor and molecular function undefined"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2002,"claim":"Whether RH2a expression is uniform across parasites was addressed by showing variable transcription/translation across lines, establishing RH2a as subject to phenotypic variation at the protein level.","evidence":"Western blot, immunofluorescence and RT-PCR across multiple parasite lines (3D7, T996, FCB1) in P. falciparum","pmids":["12228308"],"confidence":"Medium","gaps":["Mechanism controlling variant expression not defined","Consequence of expression variation for invasion untested here"],"attribution":[{"species":"Plasmodium falciparum","strain":"3D7, T996, FCB1"}]},{"year":2003,"claim":"The question of whether RH2a and RH2b are functionally interchangeable was answered by disruption showing RH2a, unlike RH2b, does not mediate the sialic acid-independent invasion pathway, framing invasion-pathway variation as receptor exploitation.","evidence":"Targeted gene disruption with enzyme-treated RBC invasion assays in P. falciparum","pmids":["12606570"],"confidence":"High","gaps":["Positive function of RH2a itself not defined","Molecular basis of paralog divergence unknown"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2006,"claim":"How RH2a is released from the merozoite surface was probed by identifying it as a substrate of the rhomboid protease PfROM4, establishing a candidate mechanism for transmembrane processing during invasion.","evidence":"Heterologous rhomboid cleavage assay with transmembrane-domain swaps in P. falciparum","pmids":["17040128"],"confidence":"Medium","gaps":["Cleavage shown in heterologous system, not in native parasite","Functional consequence of cleavage untested here"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2009,"claim":"Reciprocal disruptions clarified that RH2a is not merely redundant, showing both paralogs contribute to efficient sialic acid-independent invasion and that selection favors RH2b over RH2a expression.","evidence":"Individual PfRh2a and PfRh2b knockouts with sialic acid-depleted erythrocyte invasion assays in W2mef P. falciparum","pmids":["19400777"],"confidence":"High","gaps":["Quantitative contribution of RH2a alone not fully separable","Basis of selective pressure toward RH2b unexplained"],"attribution":[{"species":"Plasmodium falciparum","strain":"W2mef"}]},{"year":2010,"claim":"The domain responsible for functional divergence between the paralogs was mapped by chimeras to the cytoplasmic domain, localizing pathway specification to intracellular signaling rather than receptor binding.","evidence":"PfRh2a/2b domain-swap chimeric parasites with invasion pathway assays in P. falciparum","pmids":["20487292"],"confidence":"High","gaps":["Cytoplasmic-domain binding partners not identified in this study","How the tail transduces pathway choice unknown"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2010,"claim":"Whether RH2 acts alone was tested by showing EBA-181 loss abolishes RH2a/b antibody inhibition, establishing functional cooperation between PfRh and EBL protein families.","evidence":"eba gene knockouts combined with antibody growth-inhibition assays in P. falciparum","pmids":["21149582"],"confidence":"Medium","gaps":["Direct physical interaction not demonstrated","RH2a-specific vs RH2b-specific cooperation not separated"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2011,"claim":"Multiple studies defined how processing generates functionally distinct fragments and mapped the erythrocyte-binding activity to a conserved N-terminal ectodomain (15–40 kDa) shared with RH2b.","evidence":"Native protein processing western blots, enzyme-treated RBC binding assays, recombinant domain mapping and antibody inhibition in P. falciparum","pmids":["21386888","21698217"],"confidence":"Medium","gaps":["Erythrocyte receptor for the binding domain not identified","Assays largely single-lab"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2011,"claim":"Tracking RH2a during live invasion revealed pathway-independent translocation from rhoptry neck to moving junction, implying a role beyond receptor binding.","evidence":"Live and fixed immunofluorescence with anti-PfRH2a monoclonal antibodies and binding assays of processed fragments in P. falciparum","pmids":["21628513"],"confidence":"Medium","gaps":["The moving-junction function of RH2a is not mechanistically defined","Interactions at the junction not identified"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2012,"claim":"The mechanistic basis of RH2a/RH2b functional distinction was resolved biochemically: the RH2a cytoplasmic tail does not bind aldolase, unlike RH2b, explaining why RH2a is not coupled to the actin-myosin motor.","evidence":"Peptide binding, surface plasmon resonance and X-ray crystallography of aldolase with the RH2b tail as comparator in P. falciparum","pmids":["22991428"],"confidence":"High","gaps":["Alternative binding partners of the RH2a tail not identified","Whether RH2a couples to the motor by another route unknown"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2013,"claim":"A screen for heparin-binding surface proteins included RH2a among apical merozoite ligands, offering a possible route by which heparin masks invasion ligands.","evidence":"Heparin affinity chromatography, immunoblot and immunofluorescence in P. falciparum","pmids":["24212193"],"confidence":"Low","gaps":["Single pulldown without RH2a-specific functional follow-up","Physiological relevance of heparin binding for RH2a untested"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2018,"claim":"Genetics established the causal locus: forward linkage and CRISPR-Cas9 editing identified the PfRh2a/PfRh2b locus as the single major determinant of alternative invasion-pathway phenotype.","evidence":"Forward genetic cross linkage analysis and CRISPR-Cas9 editing with invasion phenotyping in P. falciparum","pmids":["30496294"],"confidence":"High","gaps":["Precise contribution of RH2a versus RH2b expression at the locus not fully partitioned","Molecular switch controlling expression not defined"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2025,"claim":"The relative immunological importance of RH2a was assessed by showing its disruption has a smaller effect on susceptibility to human inhibitory antibodies than PfRH1 or RH2b loss, ranking RH2a as a minor immune-evasion contributor.","evidence":"pfrh2a knockout parasites challenged with human immune sera in invasion inhibition assays in P. falciparum","pmids":["40201183"],"confidence":"Medium","gaps":["Epitopes and antibody targets on RH2a not mapped","Single study"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":null,"claim":"The intracellular signaling role of the RH2a cytoplasmic tail and its binding partners at the moving junction remain unresolved, leaving open how RH2a functions during invasion beyond ectodomain receptor binding.","evidence":"","pmids":[],"confidence":"Low","gaps":["No RH2a cytoplasmic-tail interactor identified","Erythrocyte receptor for the RH2a ectodomain unknown","Function at the moving junction undefined"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":[],"partners":["PFROM4 (RHOMBOID PROTEASE THAT CLEAVES RH2A TRANSMEMBRANE DOMAIN) [#3, #8]","PFRH2B (PARALOG AT THE SHARED PFRH2A/PFRH2B LOCUS) [#4, #5, #12]","EBA-181 (FUNCTIONAL COOPERATION DURING INVASION) [#6]"],"other_free_text":["RH2a cytoplasmic tail does not bind aldolase, unlike RH2b, and is not directly coupled to the actin-myosin motor [#10]","Functional divergence from RH2b is conferred by the divergent cytoplasmic domain, not the ectodomain or transmembrane region [#5]","Erythrocyte binding maps to a conserved 15-40 kDa N-terminal ectodomain shared with RH2b [#7, #8]","Variant/phenotypic expression at the PfRh2a/PfRh2b locus determines invasion-pathway choice [#0, #2, #12]","Minor target of acquired human growth-inhibitory antibodies relative to PfRH1 and RH2b [#13]"]}},"prefetch_data":{},"corpus_meta":[{"pmid":"12606570","id":"PMC_12606570","title":"Phenotypic variation of Plasmodium falciparum merozoite proteins directs receptor targeting for invasion of human erythrocytes.","date":"2003","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/12606570","citation_count":224,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PfRH2a"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"17040128","id":"PMC_17040128","title":"Two Plasmodium rhomboid proteases preferentially cleave different adhesins implicated in all invasive stages of malaria.","date":"2006","source":"PLoS 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pattern.","date":"2021","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/33886601","citation_count":8,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PfRH2a"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"23274650","id":"PMC_23274650","title":"Co-expression network with protein-protein interaction and transcription regulation in malaria parasite Plasmodium falciparum.","date":"2012","source":"Gene","url":"https://pubmed.ncbi.nlm.nih.gov/23274650","citation_count":7,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PfRH2a"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"37197657","id":"PMC_37197657","title":"High opsonic phagocytosis activity and growth inhibition of merozoites are associated with RON4 antibody levels and protect against febrile malaria in Ghanaian children.","date":"2023","source":"Frontiers in immunology","url":"https://pubmed.ncbi.nlm.nih.gov/37197657","citation_count":5,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PfRH2a"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"25255775","id":"PMC_25255775","title":"Antibody response against three Plasmodium falciparum merozoite antigens in Mamuju District, West Sulawesi Province, Indonesia.","date":"2014","source":"Malaria journal","url":"https://pubmed.ncbi.nlm.nih.gov/25255775","citation_count":3,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PfRH2a"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"33054833","id":"PMC_33054833","title":"Population genetic and biophysical evidences reveal that purifying selection shapes the genetic landscape of Plasmodium falciparum RH ligands in Chhattisgarh and West Bengal, India.","date":"2020","source":"Malaria journal","url":"https://pubmed.ncbi.nlm.nih.gov/33054833","citation_count":3,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PfRH2a"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"19421327","id":"PMC_19421327","title":"Case-control approach to identify Plasmodium falciparum polymorphisms associated with severe malaria.","date":"2009","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/19421327","citation_count":2,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["RH2a"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"40201183","id":"PMC_40201183","title":"Plasmodium falciparum reticulocyte-binding homologues are targets of human inhibitory antibodies and play a role in immune evasion.","date":"2025","source":"Frontiers in immunology","url":"https://pubmed.ncbi.nlm.nih.gov/40201183","citation_count":2,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PfRH2a"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":18389,"output_tokens":3805,"usd":0.056121,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9843,"output_tokens":4792,"usd":0.084507,"stage2_stop_reason":"end_turn"},"total_usd":0.140628,"stage1_batch_id":"msgbatch_011BPjN2BL33F8cJXD1r7V7T","stage2_batch_id":"msgbatch_01Qwg6vP7qxkpL6Gsc7tajsb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2003,\n      \"finding\": \"Targeted gene disruption of PfRh2a in P. falciparum showed that unlike PfRh2b, PfRh2a does not mediate the novel sialic acid-independent, trypsin-resistant, chymotrypsin-sensitive invasion pathway, and phenotypic variation (variant expression) of the PfRh protein family allows exploitation of different erythrocyte surface receptors.\",\n      \"method\": \"Targeted gene disruption (knockout) with erythrocyte invasion assays using enzyme-treated RBCs\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean targeted gene disruption with defined invasion pathway phenotype, replicated across multiple parasite lines\",\n      \"pmids\": [\"12606570\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"RH2a (PF3D7_1335400) encodes a >300 kDa protein expressed in late schizonts and localized to the apical end of the merozoite; no binding to mature RBCs or reticulocyte-enriched cell preparations was detected in this study, and antibodies to the protein can inhibit merozoite invasion of RBCs.\",\n      \"method\": \"Protein expression analysis, immunofluorescence localization, erythrocyte binding assay, invasion inhibition assay with antibodies\",\n      \"journal\": \"Infection and immunity\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization and binding assay, single lab, multiple methods; binding result was negative\",\n      \"pmids\": [\"11160005\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"RH2a protein is expressed in late schizonts and merozoites and is located in apical organelles and on the apical surface; expression of RH2a is variably transcribed and translated across parasite lines (absent in FCB1), indicating phenotypic variation at the protein level.\",\n      \"method\": \"Western blot, immunofluorescence, RT-PCR across multiple parasite lines\",\n      \"journal\": \"Infection and immunity\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization by immunofluorescence, protein expression confirmed by western blot, multiple parasite lines tested in one study\",\n      \"pmids\": [\"12228308\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7, T996, FCB1\"}]\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"PfROM4 (a P. falciparum rhomboid intramembrane protease) cleaves RH2a in a heterologous processing assay, with cleavage dependent on the RH2a transmembrane domain; PfROM1 did not cleave RH2a efficiently in this assay.\",\n      \"method\": \"Heterologous rhomboid cleavage assay with substrate transmembrane domain swap experiments\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — enzymatic assay with domain swaps establishing substrate specificity, but performed in heterologous system (single lab)\",\n      \"pmids\": [\"17040128\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Targeted gene disruption of PfRh2a and PfRh2b separately in W2mef parasites showed that both PfRh2b and PfRh2a contribute to efficient sialic acid-independent erythrocyte invasion, and there is strong selective pressure for reconstitution of PfRh2b expression at the expense of PfRh2a expression.\",\n      \"method\": \"Targeted gene disruption (knockout) of PfRh2a and PfRh2b individually with invasion assays using sialic acid-depleted erythrocytes\",\n      \"journal\": \"Molecular microbiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal gene disruptions with defined quantitative invasion phenotype, selective pressure documented in W2mef background\",\n      \"pmids\": [\"19400777\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"W2mef\"}]\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"The differential ability of PfRh2a vs. PfRh2b to specify alternative erythrocyte invasion pathways is conferred by their divergent cytoplasmic domains, not by the ectodomain or transmembrane regions, as shown using viable PfRh2a/2b chimeric parasites.\",\n      \"method\": \"Generation of chimeric parasites (domain-swap constructs of PfRh2a and PfRh2b) with invasion pathway assays\",\n      \"journal\": \"Molecular microbiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — domain-swap chimeric parasite approach directly maps functional diversification to cytoplasmic domain, clean genetic experiment with defined phenotypic readout\",\n      \"pmids\": [\"20487292\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Loss of function of EBA-181 ablates the ability of PfRh2a/b antibodies to inhibit merozoite invasion, providing evidence that PfRh and EBL proteins functionally interact and work cooperatively during merozoite invasion.\",\n      \"method\": \"Single and double gene knockouts of eba genes combined with antibody invasion inhibition assays (growth inhibition assay)\",\n      \"journal\": \"Infection and immunity\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — epistasis via gene knockouts with functional antibody inhibition readout, single lab\",\n      \"pmids\": [\"21149582\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Native PfRH2a/2b is proteolytically processed near the N-terminus yielding a 220 kDa fragment (sialic acid-independent, trypsin-resistant, chymotrypsin-sensitive erythrocyte binding) and an 80 kDa fragment (sialic acid-dependent, trypsin-resistant, chymotrypsin-sensitive erythrocyte binding); the erythrocyte binding domain was mapped to a conserved 40 kDa N-terminal region common to both PfRH2a and PfRH2b, and recombinant protein of this domain blocks sialic acid-independent invasion.\",\n      \"method\": \"Western blot of native protein processing, erythrocyte binding assays with enzyme-treated RBCs, recombinant protein binding, invasion inhibition with antibodies\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods (binding assay, inhibition assay, recombinant domain mapping), single lab\",\n      \"pmids\": [\"21386888\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"PfRh2a undergoes proteolytic cleavage events in the transmembrane region during invasion consistent with PfROM4 activity, releasing the ectodomain; PfRh2a and PfRh2b bind red blood cells and the erythrocyte-binding domain was defined to a 15 kDa N-terminal region of each protein; antibodies to this region block merozoite invasion.\",\n      \"method\": \"Proteolytic processing analysis (western blot), erythrocyte binding assay, antibody invasion inhibition assay\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding domain mapping, proteolysis characterization, and functional antibody blocking in single study, single lab\",\n      \"pmids\": [\"21698217\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Using monoclonal antibodies against a unique region of PfRH2a, PfRH2a was shown to translocate from the rhoptry neck to the moving junction during merozoite invasion, independent of the invasion pathway used, suggesting a function beyond receptor binding; PfRH2a is processed both in the schizont and during invasion, producing fragments with different erythrocyte binding properties.\",\n      \"method\": \"Live and fixed immunofluorescence with anti-PfRH2a monoclonal antibodies, erythrocyte binding assays with processed fragments\",\n      \"journal\": \"Infection and immunity\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization during live invasion with monoclonal antibodies, pathway-independent movement established, single lab\",\n      \"pmids\": [\"21628513\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Aldolase does NOT bind the cytoplasmic tail of RH2a (only RH2b binds aldolase among the two paralogs), indicating RH2a lacks the aromatic amino acid context required for connection to the actin-myosin motor via aldolase; this is consistent with the functional distinction that RH2b but not RH2a is required for erythrocyte invasion.\",\n      \"method\": \"Peptide binding assay, surface plasmon resonance (SPR), X-ray crystal structure of aldolase with RH2b cytoplasmic peptide\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — negative result for RH2a confirmed by SPR and structural data with RH2b as comparator; mechanistically informative negative finding with rigorous biochemical methods\",\n      \"pmids\": [\"22991428\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"RH2a (as a member of the reticulocyte binding-like family) binds heparin, localizing to the apical tip of the merozoite surface; heparin affinity chromatography followed by immunoblot confirmed heparin binding, suggesting heparin masks the apical surface and blocks erythrocyte interaction.\",\n      \"method\": \"Heparin affinity chromatography, immunoblot with ligand-specific antibodies, immunofluorescence localization\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single pulldown/affinity chromatography experiment, RH2a identified among multiple proteins without specific functional follow-up for this protein alone\",\n      \"pmids\": [\"24212193\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Forward genetic linkage analysis of experimental genetic crosses identified the PfRh2a/PfRh2b locus as the single major determinant of phenotypic variation in two erythrocyte invasion pathways; CRISPR-Cas9 editing confirmed that variation in PfRh2a and PfRh2b expression specifies alternative invasion pathway use.\",\n      \"method\": \"Forward genetic screen, whole genome sequencing linkage analysis of genetic cross progeny, CRISPR-Cas9 gene editing, invasion pathway phenotyping\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — forward genetic screen confirmed by CRISPR-Cas9 editing with quantitative invasion phenotyping, two orthogonal genetic approaches\",\n      \"pmids\": [\"30496294\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Disruption of pfrh2a in P. falciparum had a less prominent effect on susceptibility to human inhibitory antibodies compared to loss of PfRH1 or PfRH2b, suggesting PfRH2a is a minor target of acquired growth-inhibitory antibodies and plays a lesser role in immune evasion through phenotypic variation relative to PfRH1 and PfRH2b.\",\n      \"method\": \"Gene disruption (pfrh2a knockout) parasites challenged with human immune sera in invasion inhibition assays\",\n      \"journal\": \"Frontiers in immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct gene disruption with functional invasion inhibition readout using human sera, single study\",\n      \"pmids\": [\"40201183\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"current_model\": \"RH2a (PF3D7_1335400) is a >300 kDa apical merozoite protein that is proteolytically processed (including by PfROM4 at its transmembrane region) to yield fragments with differential erythrocyte binding specificities; its conserved 15–40 kDa N-terminal ectodomain mediates sialic acid-independent, chymotrypsin-sensitive erythrocyte binding, while its divergent cytoplasmic domain (which does not bind aldolase) is responsible for its functional distinction from the paralogous PfRh2b; RH2a translocates from the rhoptry neck to the moving junction during invasion in a pathway-independent manner, cooperates with EBL proteins and PfRh2b for efficient sialic acid-independent invasion, and its variant expression — controlled at the PfRh2a/PfRh2b locus as the primary determinant of invasion pathway phenotype — contributes to evasion of host immunity.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"RH2a (PF3D7_1335400) is a >300 kDa apical merozoite protein of the reticulocyte binding-like family that participates in erythrocyte invasion by Plasmodium falciparum, contributing to the selection of alternative invasion pathways [#0, #12]. It is expressed in late schizonts and merozoites and localizes to apical organelles and the apical merozoite surface, with antibodies against it inhibiting invasion [#1, #2]. Unlike its paralog PfRh2b, disruption of RH2a does not by itself specify the sialic acid-independent, trypsin-resistant, chymotrypsin-sensitive invasion pathway, but both paralogs contribute to efficient sialic acid-independent invasion, and their variant expression at the PfRh2a/PfRh2b locus is the single major genetic determinant of invasion-pathway phenotype [#0, #4, #12]. The two paralogs share a conserved N-terminal ectodomain — mapped to a 15–40 kDa region — that mediates erythrocyte binding, while their functional divergence is conferred by their divergent cytoplasmic domains rather than the ectodomain or transmembrane region [#5, #7, #8]. RH2a is proteolytically processed both in the schizont and during invasion, including transmembrane cleavage consistent with the rhomboid protease PfROM4, yielding fragments with distinct erythrocyte binding specificities, and it translocates from the rhoptry neck to the moving junction during invasion independently of the pathway used [#3, #8, #9]. The RH2a cytoplasmic tail does not bind aldolase — in contrast to RH2b — explaining why RH2a is not directly coupled to the actin-myosin motor and is functionally distinct from RH2b [#10]. Relative to PfRH1 and PfRh2b, RH2a is a minor target of acquired human growth-inhibitory antibodies [#13].\",\n  \"teleology\": [\n    {\n      \"year\": 2001,\n      \"claim\": \"Establishing where and when RH2a acts was the first requirement; its identification as a large apical schizont/merozoite protein whose antibodies block invasion placed it at the invasion interface.\",\n      \"evidence\": \"Protein expression analysis, immunofluorescence, erythrocyte binding and antibody invasion-inhibition assays in P. falciparum\",\n      \"pmids\": [\"11160005\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No direct erythrocyte binding detected in this assay\", \"Receptor and molecular function undefined\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Whether RH2a expression is uniform across parasites was addressed by showing variable transcription/translation across lines, establishing RH2a as subject to phenotypic variation at the protein level.\",\n      \"evidence\": \"Western blot, immunofluorescence and RT-PCR across multiple parasite lines (3D7, T996, FCB1) in P. falciparum\",\n      \"pmids\": [\"12228308\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism controlling variant expression not defined\", \"Consequence of expression variation for invasion untested here\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7, T996, FCB1\"}]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"The question of whether RH2a and RH2b are functionally interchangeable was answered by disruption showing RH2a, unlike RH2b, does not mediate the sialic acid-independent invasion pathway, framing invasion-pathway variation as receptor exploitation.\",\n      \"evidence\": \"Targeted gene disruption with enzyme-treated RBC invasion assays in P. falciparum\",\n      \"pmids\": [\"12606570\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Positive function of RH2a itself not defined\", \"Molecular basis of paralog divergence unknown\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"How RH2a is released from the merozoite surface was probed by identifying it as a substrate of the rhomboid protease PfROM4, establishing a candidate mechanism for transmembrane processing during invasion.\",\n      \"evidence\": \"Heterologous rhomboid cleavage assay with transmembrane-domain swaps in P. falciparum\",\n      \"pmids\": [\"17040128\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Cleavage shown in heterologous system, not in native parasite\", \"Functional consequence of cleavage untested here\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Reciprocal disruptions clarified that RH2a is not merely redundant, showing both paralogs contribute to efficient sialic acid-independent invasion and that selection favors RH2b over RH2a expression.\",\n      \"evidence\": \"Individual PfRh2a and PfRh2b knockouts with sialic acid-depleted erythrocyte invasion assays in W2mef P. falciparum\",\n      \"pmids\": [\"19400777\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Quantitative contribution of RH2a alone not fully separable\", \"Basis of selective pressure toward RH2b unexplained\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"W2mef\"}]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"The domain responsible for functional divergence between the paralogs was mapped by chimeras to the cytoplasmic domain, localizing pathway specification to intracellular signaling rather than receptor binding.\",\n      \"evidence\": \"PfRh2a/2b domain-swap chimeric parasites with invasion pathway assays in P. falciparum\",\n      \"pmids\": [\"20487292\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Cytoplasmic-domain binding partners not identified in this study\", \"How the tail transduces pathway choice unknown\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Whether RH2 acts alone was tested by showing EBA-181 loss abolishes RH2a/b antibody inhibition, establishing functional cooperation between PfRh and EBL protein families.\",\n      \"evidence\": \"eba gene knockouts combined with antibody growth-inhibition assays in P. falciparum\",\n      \"pmids\": [\"21149582\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct physical interaction not demonstrated\", \"RH2a-specific vs RH2b-specific cooperation not separated\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Multiple studies defined how processing generates functionally distinct fragments and mapped the erythrocyte-binding activity to a conserved N-terminal ectodomain (15–40 kDa) shared with RH2b.\",\n      \"evidence\": \"Native protein processing western blots, enzyme-treated RBC binding assays, recombinant domain mapping and antibody inhibition in P. falciparum\",\n      \"pmids\": [\"21386888\", \"21698217\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Erythrocyte receptor for the binding domain not identified\", \"Assays largely single-lab\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Tracking RH2a during live invasion revealed pathway-independent translocation from rhoptry neck to moving junction, implying a role beyond receptor binding.\",\n      \"evidence\": \"Live and fixed immunofluorescence with anti-PfRH2a monoclonal antibodies and binding assays of processed fragments in P. falciparum\",\n      \"pmids\": [\"21628513\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"The moving-junction function of RH2a is not mechanistically defined\", \"Interactions at the junction not identified\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"The mechanistic basis of RH2a/RH2b functional distinction was resolved biochemically: the RH2a cytoplasmic tail does not bind aldolase, unlike RH2b, explaining why RH2a is not coupled to the actin-myosin motor.\",\n      \"evidence\": \"Peptide binding, surface plasmon resonance and X-ray crystallography of aldolase with the RH2b tail as comparator in P. falciparum\",\n      \"pmids\": [\"22991428\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Alternative binding partners of the RH2a tail not identified\", \"Whether RH2a couples to the motor by another route unknown\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"A screen for heparin-binding surface proteins included RH2a among apical merozoite ligands, offering a possible route by which heparin masks invasion ligands.\",\n      \"evidence\": \"Heparin affinity chromatography, immunoblot and immunofluorescence in P. falciparum\",\n      \"pmids\": [\"24212193\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single pulldown without RH2a-specific functional follow-up\", \"Physiological relevance of heparin binding for RH2a untested\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Genetics established the causal locus: forward linkage and CRISPR-Cas9 editing identified the PfRh2a/PfRh2b locus as the single major determinant of alternative invasion-pathway phenotype.\",\n      \"evidence\": \"Forward genetic cross linkage analysis and CRISPR-Cas9 editing with invasion phenotyping in P. falciparum\",\n      \"pmids\": [\"30496294\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Precise contribution of RH2a versus RH2b expression at the locus not fully partitioned\", \"Molecular switch controlling expression not defined\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"The relative immunological importance of RH2a was assessed by showing its disruption has a smaller effect on susceptibility to human inhibitory antibodies than PfRH1 or RH2b loss, ranking RH2a as a minor immune-evasion contributor.\",\n      \"evidence\": \"pfrh2a knockout parasites challenged with human immune sera in invasion inhibition assays in P. falciparum\",\n      \"pmids\": [\"40201183\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Epitopes and antibody targets on RH2a not mapped\", \"Single study\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The intracellular signaling role of the RH2a cytoplasmic tail and its binding partners at the moving junction remain unresolved, leaving open how RH2a functions during invasion beyond ectodomain receptor binding.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No RH2a cytoplasmic-tail interactor identified\", \"Erythrocyte receptor for the RH2a ectodomain unknown\", \"Function at the moving junction undefined\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0030246\", \"supporting_discovery_ids\": [7, 8, 11]},\n      {\"term_id\": \"GO:0008201\", \"supporting_discovery_ids\": [11]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0020009\", \"supporting_discovery_ids\": [1, 2, 9]},\n      {\"term_id\": \"GO:0020008\", \"supporting_discovery_ids\": [3, 9]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0044409\", \"supporting_discovery_ids\": [0, 4, 12]},\n      {\"term_id\": \"GO:0006508\", \"supporting_discovery_ids\": [3, 7, 8]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\n      \"PfROM4 (rhomboid protease that cleaves RH2a transmembrane domain) [#3, #8]\",\n      \"PfRh2b (paralog at the shared PfRh2a/PfRh2b locus) [#4, #5, #12]\",\n      \"EBA-181 (functional cooperation during invasion) [#6]\"\n    ],\n    \"other_free_text\": [\n      \"RH2a cytoplasmic tail does not bind aldolase, unlike RH2b, and is not directly coupled to the actin-myosin motor [#10]\",\n      \"Functional divergence from RH2b is conferred by the divergent cytoplasmic domain, not the ectodomain or transmembrane region [#5]\",\n      \"Erythrocyte binding maps to a conserved 15-40 kDa N-terminal ectodomain shared with RH2b [#7, #8]\",\n      \"Variant/phenotypic expression at the PfRh2a/PfRh2b locus determines invasion-pathway choice [#0, #2, #12]\",\n      \"Minor target of acquired human growth-inhibitory antibodies relative to PfRH1 and RH2b [#13]\"\n    ]\n  }\n}","audit_flag":null,"strains":[{"strain":"Pf3D7","gene_id":"PF3D7_1335400","name":null,"product":"reticulocyte binding protein 2 homologue a","uniprot_acc":null,"chromosome":null,"start_pos":null}],"gene_symbol":"RH2a","evaluation":null}