{"gene":"SPTB","run_date":"2026-06-10T07:46:41","timeline":{"discoveries":[{"year":2022,"finding":"A novel heterozygous SPTB frameshift mutation (c.1509_1518del; p.K503Nfs*67) disrupts synthesis and localization of β-spectrin and weakens the interaction between β-spectrin and ankyrin, mediated through the nonsense-mediated mRNA decay (NMD) pathway, leading to transformation of discoid erythrocytes into spherocytes.","method":"Peripheral blood analysis, Sanger sequencing, co-immunoprecipitation/interaction assay, NMD pathway assessment","journal":"Annals of hematology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — single lab, multiple functional readouts (localization, protein interaction, NMD) but methods described only at abstract level","pmids":["35099593"],"is_preprint":false},{"year":2024,"finding":"Two novel SPTB frameshift/nonsense mutations (c.5692C>T and c.3823delG) reduce SPTB mRNA expression via the nonsense-mediated mRNA decay (NMD) pathway, as demonstrated in in vitro cultured erythroblasts derived from CD34+ stem cells.","method":"Whole-exome sequencing, Sanger sequencing, in vitro erythroblast culture from CD34+ stem cells, real-time PCR for mRNA quantification","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional in vitro erythroblast system with quantitative mRNA measurement, single lab","pmids":["39521890"],"is_preprint":false},{"year":2025,"finding":"Two novel SPTB frameshift/nonsense variants (c.493_494insTG, p.Q165fs and c.1715delT, p.L572X) cause decreased β-spectrin protein expression, as demonstrated by Western blot analysis.","method":"Next-generation sequencing, Sanger sequencing, Western blot","journal":"Annals of human genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — Western blot directly measures protein loss-of-function, single lab, single method","pmids":["40551579"],"is_preprint":false},{"year":2024,"finding":"A novel SPTB splice site variant (c.301-2 A>G) produces three aberrant transcripts (r.301_474del, r.301_306delCCAAAG, and r.301-1_301-57ins), confirming disruption of normal splicing and aberrant mRNA translation, as shown by an in vitro minigene splicing reporter assay.","method":"In vitro minigene splicing assay, Sanger sequencing","journal":"BMC medical genomics","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — minigene splicing assay is a direct functional test; single lab, single method","pmids":["39135028"],"is_preprint":false},{"year":2021,"finding":"A SPTB splice site variant (c.1064+1G>A) causes exclusion of exon 8 with subsequent frameshift in exon 9 and premature stop codon, predicted to truncate all spectrin repeat domains of SPTB; confirmed by RT-PCR and TA cloning sequencing in patient samples.","method":"Whole-genome sequencing, Sanger sequencing, RT-PCR, TA cloning, sequencing","journal":"Molecular genetics & genomic medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — RT-PCR with sequencing directly demonstrates aberrant splicing product in patient material; co-segregation in large family","pmids":["33943044"],"is_preprint":false},{"year":2020,"finding":"A SPTB intronic splice site variant (c.5798+1G>A) significantly affects splicing, resulting in a reading frameshift and premature termination codon, confirmed by both in vitro (293T cell transfection) and in vivo (patient peripheral blood RNA) splicing experiments.","method":"In vitro splicing assay (293T cell transfection with pCAS2 plasmid constructs), in vivo RT-PCR from patient blood, TA cloning, Sanger sequencing","journal":"Chinese journal of medical genetics","confidence":"High","confidence_rationale":"Tier 1 / Moderate — reconstitution-style in vitro splicing assay plus in vivo validation in patient RNA, two orthogonal methods, single lab","pmids":["31922588"],"is_preprint":false},{"year":2023,"finding":"A rare intronic inversion variant between introns 19 and 20 of SPTB (containing entire exon 20) causes skipping of exon 20 and reduced SPTB mRNA expression, demonstrated by RNA sequencing of patient material.","method":"Targeted NGS, Sanger sequencing, RNA sequencing","journal":"Frontiers in genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — RNA sequencing directly shows exon skipping and reduced expression; single lab, single case","pmids":["38111681"],"is_preprint":false},{"year":2025,"finding":"A novel SPTB intronic variant (c.6022+4_6022+18delinsTGGCTCCTCCGTGAAGGGACAGTCCTGC) activates a cryptic splice donor site leading to a frameshift, premature termination codon, and nonsense-mediated mRNA degradation, confirmed by mRNA sequencing and bioinformatics splicing prediction tools in patient and family members.","method":"Whole-genome sequencing, Sanger sequencing, mRNA expression quantification (RT-qPCR), mRNA sequencing, bioinformatics (ESE Finder, SpliceAI, SpliceTool)","journal":"Chinese journal of pediatrics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — mRNA sequencing plus quantitative RT-PCR in multiple affected family members, co-segregation analysis; single lab","pmids":["40090920"],"is_preprint":false},{"year":2025,"finding":"A novel SPTB splice-donor site mutation (c.647+1G>A) abolishes the wild-type splice donor, causing complete loss of exon 5 in an alternative transcript and significantly reduced mutated SPTB mRNA expression, demonstrated by RT-PCR and cDNA sequencing in patient and mother.","method":"Whole-exome sequencing, Sanger sequencing, RT-PCR, cDNA sequencing, quantitative real-time PCR, Western blot","journal":"Frontiers in genetics","confidence":"High","confidence_rationale":"Tier 1 / Moderate — multiple orthogonal methods (RT-PCR, cDNA sequencing, qRT-PCR, Western blot) in the same study confirming splicing defect and protein loss; single lab","pmids":["41321563"],"is_preprint":false},{"year":1998,"finding":"De novo mutations in SPTB frequently result in monoallelic expression (failure of mRNA accumulation from one allele), explaining apparently recessive patterns of hereditary spherocytosis with isolated spectrin deficiency in children with hematologically normal parents.","method":"Genomic DNA polymorphism analysis, RT-PCR of cDNA from patient peripheral blood, allele-specific expression analysis","journal":"British journal of haematology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — RT-PCR on patient cDNA with polymorphism-based allele tracking, multiple patients (7/13), single lab","pmids":["9609518"],"is_preprint":false},{"year":1990,"finding":"The SPTB gene (encoding erythrocyte β-spectrin) was regionally localized to human chromosome 14q23–q24.2 by in situ hybridization using a cDNA probe.","method":"In situ hybridization, somatic cell hybrid analysis","journal":"Cytogenetics and cell genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct chromosomal localization by in situ hybridization, replicated with somatic cell hybrid data","pmids":["2209094"],"is_preprint":false},{"year":2023,"finding":"SPTB protein without CH1 and CH2 domains (caused by CNV deletion of exons 2-3) cannot bind to erythrocyte membrane actin, as inferred from database domain analysis (UniProt/SMART) combined with RT-qPCR showing reduced SPTB mRNA in patients with CNV deletion.","method":"Second-generation sequencing, RT-qPCR, UniProt/SMART domain analysis","journal":"Chinese journal of experimental hematology","confidence":"Low","confidence_rationale":"Tier 4 / Weak — actin-binding inference is computational/database-based; reduced mRNA confirmed experimentally but actin binding was not directly tested","pmids":["36765497"],"is_preprint":false}],"current_model":"SPTB encodes erythrocyte β-spectrin, a cytoskeletal protein whose CH1/CH2 domains mediate actin binding and whose spectrin repeat domains interact with ankyrin; loss-of-function mutations (frameshift, nonsense, splice site) cause β-spectrin deficiency predominantly via nonsense-mediated mRNA decay, disrupting β-spectrin synthesis, localization, and its interaction with ankyrin, thereby destabilizing the erythrocyte membrane skeleton and causing hereditary spherocytosis."},"narrative":{"mechanistic_narrative":"SPTB encodes erythrocyte β-spectrin, a cytoskeletal protein whose CH1/CH2 domains mediate binding to erythrocyte membrane actin and whose spectrin repeat domains support assembly of the membrane skeleton through interaction with ankyrin; pathogenic loss-of-function variants destabilize this skeleton and convert discoid erythrocytes into spherocytes, causing hereditary spherocytosis [PMID:35099593]. The dominant mechanism across the corpus is haploinsufficiency arising from premature termination: frameshift, nonsense, and splice-disrupting variants generate transcripts bearing premature stop codons that are degraded by nonsense-mediated mRNA decay, lowering SPTB mRNA and β-spectrin protein levels [PMID:35099593, PMID:39521890, PMID:40551579]. Many disease alleles act at the level of splicing—splice-donor abolition, cryptic-site activation, intronic inversion, and exon skipping all yield frameshifted, prematurely terminated transcripts and reduced expression [PMID:39135028, PMID:33943044, PMID:31922588, PMID:38111681, PMID:40090920, PMID:41321563]. De novo SPTB mutations frequently manifest as monoallelic expression, accounting for apparently recessive spherocytosis in children of hematologically normal parents [PMID:9609518]. Structurally, deletion of the CH1/CH2-encoding exons produces a β-spectrin unable to engage membrane actin [PMID:36765497]. Beyond this erythrocyte membrane-skeleton role and its perturbation by reduced expression, no additional molecular functions are characterized in the available corpus.","teleology":[{"year":1990,"claim":"Before its chromosomal assignment, the genomic location of erythrocyte β-spectrin was undefined; mapping the gene anchored SPTB as a discrete locus for linkage to inherited red-cell disorders.","evidence":"In situ hybridization and somatic cell hybrid analysis with a cDNA probe","pmids":["2209094"],"confidence":"Medium","gaps":["Does not address protein function or disease mechanism","No mutation-phenotype link established"]},{"year":1998,"claim":"It was unclear why isolated spectrin-deficient spherocytosis could appear recessive in families with normal parents; allele-specific expression analysis showed de novo SPTB mutations frequently cause monoallelic expression, reframing inheritance as dominant haploinsufficiency masked by failure of mutant allele accumulation.","evidence":"RT-PCR on patient cDNA with polymorphism-based allele tracking in multiple patients","pmids":["9609518"],"confidence":"Medium","gaps":["Mechanism of mRNA non-accumulation not resolved at the molecular level","Does not directly demonstrate NMD"]},{"year":2022,"claim":"How a single frameshift allele produces disease was unresolved; functional analysis tied a frameshift variant to disrupted β-spectrin synthesis and localization, weakened β-spectrin–ankyrin interaction, and NMD, linking genotype to spherocyte formation.","evidence":"Patient blood analysis, interaction/co-immunoprecipitation assay, and NMD pathway assessment","pmids":["35099593"],"confidence":"Medium","gaps":["Methods described at abstract level only","Single lab; β-spectrin–ankyrin interaction not quantified by orthogonal assay"]},{"year":2020,"claim":"Whether intronic splice-site variants are pathogenic required functional proof; orthogonal in vitro and patient-RNA splicing assays showed a c.5798+1G>A variant causes frameshift and premature termination, establishing splicing disruption as a disease mechanism.","evidence":"pCAS2 minigene splicing assay in 293T cells plus in vivo RT-PCR from patient blood with TA cloning","pmids":["31922588"],"confidence":"High","gaps":["Protein-level loss not directly measured in this study","Single lab"]},{"year":2021,"claim":"The structural consequence of splice variants on the protein was unclear; demonstration that c.1064+1G>A causes exon 8 exclusion, frameshift, and a premature stop predicted to truncate all spectrin repeat domains connected aberrant splicing to loss of the functional skeleton-assembly region.","evidence":"RT-PCR and TA cloning sequencing in patient samples with family co-segregation","pmids":["33943044"],"confidence":"Medium","gaps":["Truncation of spectrin repeats inferred, not protein-verified","Single lab"]},{"year":2023,"claim":"It was unknown whether structural rearrangements and CH-domain loss contribute to SPTB disease; an intronic inversion was shown to cause exon 20 skipping with reduced mRNA, and CNV deletion of CH1/CH2-encoding exons was inferred to abolish actin binding, expanding the variant spectrum.","evidence":"RNA sequencing of patient material (inversion); RT-qPCR plus UniProt/SMART domain analysis (CNV deletion)","pmids":["38111681","36765497"],"confidence":"Low","gaps":["Actin-binding loss is computational/database inference, not directly tested","Single cases per variant"]},{"year":2024,"claim":"Whether premature-termination alleles act via mRNA decay needed direct measurement; quantitative mRNA analysis in CD34+-derived erythroblasts and a minigene assay confirmed NMD-mediated reduction and aberrant splicing, solidifying haploinsufficiency as the unifying mechanism.","evidence":"In vitro erythroblast culture with real-time PCR (frameshift/nonsense); minigene splicing reporter assay (splice variant)","pmids":["39521890","39135028"],"confidence":"Medium","gaps":["Protein-level consequences not assayed in these systems","Single lab per study"]},{"year":2025,"claim":"Linking transcript defects to actual protein loss closed a key gap; Western blot showed frameshift/nonsense variants decrease β-spectrin protein, and combined RT-PCR/cDNA/qRT-PCR/Western analysis confirmed a splice-donor mutation causes exon loss with reduced mRNA and protein, while cryptic-site activation drives NMD.","evidence":"Western blot of patient material; multi-method splicing and expression analysis (RT-PCR, cDNA sequencing, qRT-PCR, Western blot, mRNA sequencing) across affected family members","pmids":["40551579","41321563","40090920"],"confidence":"High","gaps":["Membrane-skeleton structural consequences not directly imaged","Single lab per variant"]},{"year":null,"claim":"How quantitatively reduced β-spectrin translates into graded membrane-skeleton instability and clinical severity, and whether any non-erythroid or interaction-specific functions of SPTB exist, remains uncharacterized in this corpus.","evidence":"","pmids":[],"confidence":"Low","gaps":["No structural model of mutant skeleton assembly","β-spectrin–ankyrin interaction tested in only one study","No quantitative genotype-severity relationship established"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[0,11]},{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[0,4]}],"localization":[{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[0]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[11]}],"pathway":[{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[0,9]}],"complexes":["erythrocyte membrane skeleton"],"partners":["ANK1","ACTIN"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P11277","full_name":"Spectrin beta chain, erythrocytic","aliases":["Beta-I spectrin"],"length_aa":2137,"mass_kda":246.5,"function":"Spectrin is the major constituent of the cytoskeletal network underlying the erythrocyte plasma membrane. It associates with band 4.1 and actin to form the cytoskeletal superstructure of the erythrocyte plasma membrane","subcellular_location":"Cytoplasm, cytoskeleton; Cytoplasm, cell cortex","url":"https://www.uniprot.org/uniprotkb/P11277/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/SPTB","classification":"Not Classified","n_dependent_lines":2,"n_total_lines":1208,"dependency_fraction":0.0016556291390728477},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/SPTB","total_profiled":1310},"omim":[{"mim_id":"617948","title":"ELLIPTOCYTOSIS 3; EL3","url":"https://www.omim.org/entry/617948"},{"mim_id":"616649","title":"SPHEROCYTOSIS, TYPE 2; SPH2","url":"https://www.omim.org/entry/616649"},{"mim_id":"611804","title":"ELLIPTOCYTOSIS 1; EL1","url":"https://www.omim.org/entry/611804"},{"mim_id":"604985","title":"SPECTRIN, BETA, NONERYTHROCYTIC, 2; SPTBN2","url":"https://www.omim.org/entry/604985"},{"mim_id":"266140","title":"PYROPOIKILOCYTOSIS, HEREDITARY; HPP","url":"https://www.omim.org/entry/266140"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Cytosol","reliability":"Approved"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"bone marrow","ntpm":59.0},{"tissue":"skeletal muscle","ntpm":153.7},{"tissue":"tongue","ntpm":65.2}],"url":"https://www.proteinatlas.org/search/SPTB"},"hgnc":{"alias_symbol":[],"prev_symbol":[]},"alphafold":{"accession":"P11277","domains":[{"cath_id":"1.10.418.10","chopping":"40-283","consensus_level":"medium","plddt":81.8777,"start":40,"end":283},{"cath_id":"1.20.58.60","chopping":"296-424","consensus_level":"medium","plddt":85.9592,"start":296,"end":424},{"cath_id":"1.20.58.60","chopping":"983-1170","consensus_level":"medium","plddt":79.3165,"start":983,"end":1170},{"cath_id":"1.20.58.60","chopping":"1716-1811","consensus_level":"medium","plddt":87.8706,"start":1716,"end":1811},{"cath_id":"1.10.287","chopping":"1610-1695","consensus_level":"high","plddt":87.1873,"start":1610,"end":1695}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P11277","model_url":"https://alphafold.ebi.ac.uk/files/AF-P11277-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P11277-F1-predicted_aligned_error_v6.png","plddt_mean":79.19},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=SPTB","jax_strain_url":"https://www.jax.org/strain/search?query=SPTB"},"sequence":{"accession":"P11277","fasta_url":"https://rest.uniprot.org/uniprotkb/P11277.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P11277/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P11277"}},"corpus_meta":[{"pmid":"26830532","id":"PMC_26830532","title":"Mutational 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chuan xue za zhi = Zhonghua yixue yichuanxue zazhi = Chinese journal of medical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/36854399","citation_count":1,"is_preprint":false},{"pmid":"40327078","id":"PMC_40327078","title":"Understanding the genetic architecture and phenotypic landscape of SPTB gene variants causing hereditary spherocytosis in an Indian cohort.","date":"2025","source":"Human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/40327078","citation_count":1,"is_preprint":false},{"pmid":"38111681","id":"PMC_38111681","title":"Case report: Genetic analysis of a novel intronic inversion variant in the SPTB gene associated with hereditary spherocytosis.","date":"2023","source":"Frontiers in genetics","url":"https://pubmed.ncbi.nlm.nih.gov/38111681","citation_count":1,"is_preprint":false},{"pmid":"38831725","id":"PMC_38831725","title":"Clinical Exome Sequencing Reveals Novel Mutations in SPTB Gene Associated with Hereditary Spherocytosis in Patients with Suspected Congenital Hemolytic Anemia.","date":"2024","source":"Hemoglobin","url":"https://pubmed.ncbi.nlm.nih.gov/38831725","citation_count":1,"is_preprint":false},{"pmid":"36765497","id":"PMC_36765497","title":"[Genetic Analysis of a Chinese Pedigree with Hereditary Spherocytosis Caused by Copy Number Variation Deletion of SPTB Gene].","date":"2023","source":"Zhongguo shi yan xue ye xue za zhi","url":"https://pubmed.ncbi.nlm.nih.gov/36765497","citation_count":1,"is_preprint":false},{"pmid":"31014431","id":"PMC_31014431","title":"[Clinical characteristics and genetic analysis of hereditary spherocytosis caused by mutations of ANK1 and SPTB genes].","date":"2019","source":"Zhongguo dang dai er ke za zhi = Chinese journal of contemporary pediatrics","url":"https://pubmed.ncbi.nlm.nih.gov/31014431","citation_count":1,"is_preprint":false},{"pmid":"37795245","id":"PMC_37795245","title":"Integrative preimplantation genetic testing analysis for a Chinese family with hereditary spherocytosis caused by a novel splicing variant of SPTB.","date":"2023","source":"Frontiers in genetics","url":"https://pubmed.ncbi.nlm.nih.gov/37795245","citation_count":1,"is_preprint":false},{"pmid":"40632418","id":"PMC_40632418","title":"Angioid streaks in hereditary spherocytosis associated with an SPTB gene variant.","date":"2025","source":"Documenta ophthalmologica. Advances in ophthalmology","url":"https://pubmed.ncbi.nlm.nih.gov/40632418","citation_count":0,"is_preprint":false},{"pmid":"40551579","id":"PMC_40551579","title":"Novel SPTB Variations Cause Hereditary Spherocytosis With Cholangiolithiasis and Severe Intrahepatic Cholestasis.","date":"2025","source":"Annals of human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/40551579","citation_count":0,"is_preprint":false},{"pmid":"35395996","id":"PMC_35395996","title":"[Genetic Analysis and Prenatal Diagnosis of a Family with Hereditary Spherocytosis Caused by a Novel Compound Heterozygous Mutation of SPTB Gene].","date":"2022","source":"Zhongguo shi yan xue ye xue za zhi","url":"https://pubmed.ncbi.nlm.nih.gov/35395996","citation_count":0,"is_preprint":false},{"pmid":"41321563","id":"PMC_41321563","title":"Case Report: Identification and functional characterization of a novel heterozygous splice-donor (c.647+1G>A) site mutation in the SPTB gene that causes hereditary spherocytosis with hemolytic anemia.","date":"2025","source":"Frontiers in genetics","url":"https://pubmed.ncbi.nlm.nih.gov/41321563","citation_count":0,"is_preprint":false},{"pmid":"41920367","id":"PMC_41920367","title":"De novo mutations in ANK1 and SPTB cause hereditary spherocytosis: three case reports and literature review.","date":"2026","source":"Annals of hematology","url":"https://pubmed.ncbi.nlm.nih.gov/41920367","citation_count":0,"is_preprint":false},{"pmid":"40090920","id":"PMC_40090920","title":"[Analysis of a Chinese pedigree with hereditary spherocytosis caused by intron variation of SPTB gene].","date":"2025","source":"Zhonghua er ke za zhi = Chinese journal of pediatrics","url":"https://pubmed.ncbi.nlm.nih.gov/40090920","citation_count":0,"is_preprint":false},{"pmid":"31922588","id":"PMC_31922588","title":"[Hereditary spherocytosis due to a novel c.5798+1G>A variant of the SPTB gene].","date":"2020","source":"Zhonghua yi xue yi chuan xue za zhi = Zhonghua yixue yichuanxue zazhi = Chinese journal of medical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/31922588","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.11.21.25340724","title":"Association of AAT/  <i>SERPINA1</i>  PI*Z Variant with Gestational Duration and Prevention of Premature Birth in Knockout Mice by AAT-Supplementation","date":"2025-11-21","source":"bioRxiv","url":"https://doi.org/10.1101/2025.11.21.25340724","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.08.07.25333197","title":"Maternal Metabolomic Signatures of Spontaneous Preterm Birth in Bangladesh: Evidence from the AMANHI Bangladesh Cohort","date":"2025-08-11","source":"bioRxiv","url":"https://doi.org/10.1101/2025.08.07.25333197","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.05.19.654577","title":"The maternal inflammatory proteome during pregnancy and its role in predicting the risk of spontaneous preterm birth","date":"2025-05-23","source":"bioRxiv","url":"https://doi.org/10.1101/2025.05.19.654577","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.05.11.25327390","title":"<i>KCNN3</i>genetic variants rs1218585 and rs1218584 are associated with spontaneous preterm birth in a Portuguese population","date":"2025-05-12","source":"bioRxiv","url":"https://doi.org/10.1101/2025.05.11.25327390","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.09.24.25336553","title":"Plasma proteome profiling identifies predictive signatures for preterm birth risk","date":"2025-09-29","source":"bioRxiv","url":"https://doi.org/10.1101/2025.09.24.25336553","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.01.31.635906","title":"Multi-omic data integration and analyses for biomarker discovery of spontaneous preterm birth phenotypes","date":"2025-02-04","source":"bioRxiv","url":"https://doi.org/10.1101/2025.01.31.635906","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.06.09.658195","title":"Placental Iron Utilisation in Fetal Growth Restriction: Alterations in Mitochondrial Heme Synthesis and Iron-Sulfur Cluster Assembly Pathways","date":"2025-06-12","source":"bioRxiv","url":"https://doi.org/10.1101/2025.06.09.658195","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":25953,"output_tokens":2699,"usd":0.059172,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":10000,"output_tokens":2954,"usd":0.061925,"stage2_stop_reason":"end_turn"},"total_usd":0.121097,"stage1_batch_id":"msgbatch_013LWZZT6MWaqB9qyTSeQBmU","stage2_batch_id":"msgbatch_0194ZzQaTGNpruoNJBbEMbt3","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2022,\n      \"finding\": \"A novel heterozygous SPTB frameshift mutation (c.1509_1518del; p.K503Nfs*67) disrupts synthesis and localization of β-spectrin and weakens the interaction between β-spectrin and ankyrin, mediated through the nonsense-mediated mRNA decay (NMD) pathway, leading to transformation of discoid erythrocytes into spherocytes.\",\n      \"method\": \"Peripheral blood analysis, Sanger sequencing, co-immunoprecipitation/interaction assay, NMD pathway assessment\",\n      \"journal\": \"Annals of hematology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — single lab, multiple functional readouts (localization, protein interaction, NMD) but methods described only at abstract level\",\n      \"pmids\": [\"35099593\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Two novel SPTB frameshift/nonsense mutations (c.5692C>T and c.3823delG) reduce SPTB mRNA expression via the nonsense-mediated mRNA decay (NMD) pathway, as demonstrated in in vitro cultured erythroblasts derived from CD34+ stem cells.\",\n      \"method\": \"Whole-exome sequencing, Sanger sequencing, in vitro erythroblast culture from CD34+ stem cells, real-time PCR for mRNA quantification\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional in vitro erythroblast system with quantitative mRNA measurement, single lab\",\n      \"pmids\": [\"39521890\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Two novel SPTB frameshift/nonsense variants (c.493_494insTG, p.Q165fs and c.1715delT, p.L572X) cause decreased β-spectrin protein expression, as demonstrated by Western blot analysis.\",\n      \"method\": \"Next-generation sequencing, Sanger sequencing, Western blot\",\n      \"journal\": \"Annals of human genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — Western blot directly measures protein loss-of-function, single lab, single method\",\n      \"pmids\": [\"40551579\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"A novel SPTB splice site variant (c.301-2 A>G) produces three aberrant transcripts (r.301_474del, r.301_306delCCAAAG, and r.301-1_301-57ins), confirming disruption of normal splicing and aberrant mRNA translation, as shown by an in vitro minigene splicing reporter assay.\",\n      \"method\": \"In vitro minigene splicing assay, Sanger sequencing\",\n      \"journal\": \"BMC medical genomics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — minigene splicing assay is a direct functional test; single lab, single method\",\n      \"pmids\": [\"39135028\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"A SPTB splice site variant (c.1064+1G>A) causes exclusion of exon 8 with subsequent frameshift in exon 9 and premature stop codon, predicted to truncate all spectrin repeat domains of SPTB; confirmed by RT-PCR and TA cloning sequencing in patient samples.\",\n      \"method\": \"Whole-genome sequencing, Sanger sequencing, RT-PCR, TA cloning, sequencing\",\n      \"journal\": \"Molecular genetics & genomic medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — RT-PCR with sequencing directly demonstrates aberrant splicing product in patient material; co-segregation in large family\",\n      \"pmids\": [\"33943044\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"A SPTB intronic splice site variant (c.5798+1G>A) significantly affects splicing, resulting in a reading frameshift and premature termination codon, confirmed by both in vitro (293T cell transfection) and in vivo (patient peripheral blood RNA) splicing experiments.\",\n      \"method\": \"In vitro splicing assay (293T cell transfection with pCAS2 plasmid constructs), in vivo RT-PCR from patient blood, TA cloning, Sanger sequencing\",\n      \"journal\": \"Chinese journal of medical genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — reconstitution-style in vitro splicing assay plus in vivo validation in patient RNA, two orthogonal methods, single lab\",\n      \"pmids\": [\"31922588\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"A rare intronic inversion variant between introns 19 and 20 of SPTB (containing entire exon 20) causes skipping of exon 20 and reduced SPTB mRNA expression, demonstrated by RNA sequencing of patient material.\",\n      \"method\": \"Targeted NGS, Sanger sequencing, RNA sequencing\",\n      \"journal\": \"Frontiers in genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — RNA sequencing directly shows exon skipping and reduced expression; single lab, single case\",\n      \"pmids\": [\"38111681\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"A novel SPTB intronic variant (c.6022+4_6022+18delinsTGGCTCCTCCGTGAAGGGACAGTCCTGC) activates a cryptic splice donor site leading to a frameshift, premature termination codon, and nonsense-mediated mRNA degradation, confirmed by mRNA sequencing and bioinformatics splicing prediction tools in patient and family members.\",\n      \"method\": \"Whole-genome sequencing, Sanger sequencing, mRNA expression quantification (RT-qPCR), mRNA sequencing, bioinformatics (ESE Finder, SpliceAI, SpliceTool)\",\n      \"journal\": \"Chinese journal of pediatrics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — mRNA sequencing plus quantitative RT-PCR in multiple affected family members, co-segregation analysis; single lab\",\n      \"pmids\": [\"40090920\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"A novel SPTB splice-donor site mutation (c.647+1G>A) abolishes the wild-type splice donor, causing complete loss of exon 5 in an alternative transcript and significantly reduced mutated SPTB mRNA expression, demonstrated by RT-PCR and cDNA sequencing in patient and mother.\",\n      \"method\": \"Whole-exome sequencing, Sanger sequencing, RT-PCR, cDNA sequencing, quantitative real-time PCR, Western blot\",\n      \"journal\": \"Frontiers in genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — multiple orthogonal methods (RT-PCR, cDNA sequencing, qRT-PCR, Western blot) in the same study confirming splicing defect and protein loss; single lab\",\n      \"pmids\": [\"41321563\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"De novo mutations in SPTB frequently result in monoallelic expression (failure of mRNA accumulation from one allele), explaining apparently recessive patterns of hereditary spherocytosis with isolated spectrin deficiency in children with hematologically normal parents.\",\n      \"method\": \"Genomic DNA polymorphism analysis, RT-PCR of cDNA from patient peripheral blood, allele-specific expression analysis\",\n      \"journal\": \"British journal of haematology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — RT-PCR on patient cDNA with polymorphism-based allele tracking, multiple patients (7/13), single lab\",\n      \"pmids\": [\"9609518\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1990,\n      \"finding\": \"The SPTB gene (encoding erythrocyte β-spectrin) was regionally localized to human chromosome 14q23–q24.2 by in situ hybridization using a cDNA probe.\",\n      \"method\": \"In situ hybridization, somatic cell hybrid analysis\",\n      \"journal\": \"Cytogenetics and cell genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct chromosomal localization by in situ hybridization, replicated with somatic cell hybrid data\",\n      \"pmids\": [\"2209094\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"SPTB protein without CH1 and CH2 domains (caused by CNV deletion of exons 2-3) cannot bind to erythrocyte membrane actin, as inferred from database domain analysis (UniProt/SMART) combined with RT-qPCR showing reduced SPTB mRNA in patients with CNV deletion.\",\n      \"method\": \"Second-generation sequencing, RT-qPCR, UniProt/SMART domain analysis\",\n      \"journal\": \"Chinese journal of experimental hematology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 4 / Weak — actin-binding inference is computational/database-based; reduced mRNA confirmed experimentally but actin binding was not directly tested\",\n      \"pmids\": [\"36765497\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"SPTB encodes erythrocyte β-spectrin, a cytoskeletal protein whose CH1/CH2 domains mediate actin binding and whose spectrin repeat domains interact with ankyrin; loss-of-function mutations (frameshift, nonsense, splice site) cause β-spectrin deficiency predominantly via nonsense-mediated mRNA decay, disrupting β-spectrin synthesis, localization, and its interaction with ankyrin, thereby destabilizing the erythrocyte membrane skeleton and causing hereditary spherocytosis.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"SPTB encodes erythrocyte β-spectrin, a cytoskeletal protein whose CH1/CH2 domains mediate binding to erythrocyte membrane actin and whose spectrin repeat domains support assembly of the membrane skeleton through interaction with ankyrin; pathogenic loss-of-function variants destabilize this skeleton and convert discoid erythrocytes into spherocytes, causing hereditary spherocytosis [#0]. The dominant mechanism across the corpus is haploinsufficiency arising from premature termination: frameshift, nonsense, and splice-disrupting variants generate transcripts bearing premature stop codons that are degraded by nonsense-mediated mRNA decay, lowering SPTB mRNA and β-spectrin protein levels [#0, #1, #2]. Many disease alleles act at the level of splicing—splice-donor abolition, cryptic-site activation, intronic inversion, and exon skipping all yield frameshifted, prematurely terminated transcripts and reduced expression [#3, #4, #5, #6, #7, #8]. De novo SPTB mutations frequently manifest as monoallelic expression, accounting for apparently recessive spherocytosis in children of hematologically normal parents [#9]. Structurally, deletion of the CH1/CH2-encoding exons produces a β-spectrin unable to engage membrane actin [#11]. Beyond this erythrocyte membrane-skeleton role and its perturbation by reduced expression, no additional molecular functions are characterized in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 1990,\n      \"claim\": \"Before its chromosomal assignment, the genomic location of erythrocyte β-spectrin was undefined; mapping the gene anchored SPTB as a discrete locus for linkage to inherited red-cell disorders.\",\n      \"evidence\": \"In situ hybridization and somatic cell hybrid analysis with a cDNA probe\",\n      \"pmids\": [\"2209094\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Does not address protein function or disease mechanism\", \"No mutation-phenotype link established\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"It was unclear why isolated spectrin-deficient spherocytosis could appear recessive in families with normal parents; allele-specific expression analysis showed de novo SPTB mutations frequently cause monoallelic expression, reframing inheritance as dominant haploinsufficiency masked by failure of mutant allele accumulation.\",\n      \"evidence\": \"RT-PCR on patient cDNA with polymorphism-based allele tracking in multiple patients\",\n      \"pmids\": [\"9609518\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism of mRNA non-accumulation not resolved at the molecular level\", \"Does not directly demonstrate NMD\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"How a single frameshift allele produces disease was unresolved; functional analysis tied a frameshift variant to disrupted β-spectrin synthesis and localization, weakened β-spectrin–ankyrin interaction, and NMD, linking genotype to spherocyte formation.\",\n      \"evidence\": \"Patient blood analysis, interaction/co-immunoprecipitation assay, and NMD pathway assessment\",\n      \"pmids\": [\"35099593\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Methods described at abstract level only\", \"Single lab; β-spectrin–ankyrin interaction not quantified by orthogonal assay\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Whether intronic splice-site variants are pathogenic required functional proof; orthogonal in vitro and patient-RNA splicing assays showed a c.5798+1G>A variant causes frameshift and premature termination, establishing splicing disruption as a disease mechanism.\",\n      \"evidence\": \"pCAS2 minigene splicing assay in 293T cells plus in vivo RT-PCR from patient blood with TA cloning\",\n      \"pmids\": [\"31922588\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Protein-level loss not directly measured in this study\", \"Single lab\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"The structural consequence of splice variants on the protein was unclear; demonstration that c.1064+1G>A causes exon 8 exclusion, frameshift, and a premature stop predicted to truncate all spectrin repeat domains connected aberrant splicing to loss of the functional skeleton-assembly region.\",\n      \"evidence\": \"RT-PCR and TA cloning sequencing in patient samples with family co-segregation\",\n      \"pmids\": [\"33943044\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Truncation of spectrin repeats inferred, not protein-verified\", \"Single lab\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"It was unknown whether structural rearrangements and CH-domain loss contribute to SPTB disease; an intronic inversion was shown to cause exon 20 skipping with reduced mRNA, and CNV deletion of CH1/CH2-encoding exons was inferred to abolish actin binding, expanding the variant spectrum.\",\n      \"evidence\": \"RNA sequencing of patient material (inversion); RT-qPCR plus UniProt/SMART domain analysis (CNV deletion)\",\n      \"pmids\": [\"38111681\", \"36765497\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Actin-binding loss is computational/database inference, not directly tested\", \"Single cases per variant\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Whether premature-termination alleles act via mRNA decay needed direct measurement; quantitative mRNA analysis in CD34+-derived erythroblasts and a minigene assay confirmed NMD-mediated reduction and aberrant splicing, solidifying haploinsufficiency as the unifying mechanism.\",\n      \"evidence\": \"In vitro erythroblast culture with real-time PCR (frameshift/nonsense); minigene splicing reporter assay (splice variant)\",\n      \"pmids\": [\"39521890\", \"39135028\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Protein-level consequences not assayed in these systems\", \"Single lab per study\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Linking transcript defects to actual protein loss closed a key gap; Western blot showed frameshift/nonsense variants decrease β-spectrin protein, and combined RT-PCR/cDNA/qRT-PCR/Western analysis confirmed a splice-donor mutation causes exon loss with reduced mRNA and protein, while cryptic-site activation drives NMD.\",\n      \"evidence\": \"Western blot of patient material; multi-method splicing and expression analysis (RT-PCR, cDNA sequencing, qRT-PCR, Western blot, mRNA sequencing) across affected family members\",\n      \"pmids\": [\"40551579\", \"41321563\", \"40090920\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Membrane-skeleton structural consequences not directly imaged\", \"Single lab per variant\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How quantitatively reduced β-spectrin translates into graded membrane-skeleton instability and clinical severity, and whether any non-erythroid or interaction-specific functions of SPTB exist, remains uncharacterized in this corpus.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No structural model of mutant skeleton assembly\", \"β-spectrin–ankyrin interaction tested in only one study\", \"No quantitative genotype-severity relationship established\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [0, 11]},\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [0, 4]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [11]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [0, 9]}\n    ],\n    \"complexes\": [\"erythrocyte membrane skeleton\"],\n    \"partners\": [\"ANK1\", \"actin\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"tie","faith_supported":5,"faith_total":5,"faith_pct":100.0}}