id: Q8AY16 gene_symbol: smad3b product_type: PROTEIN status: COMPLETE taxon: id: NCBITaxon:7955 label: Danio rerio description: 'smad3b (SMAD family member 3b) is one of two zebrafish co-orthologs of vertebrate SMAD3, retained from the teleost-specific genome duplication; its paralog is smad3a. It encodes a receptor-regulated SMAD with an MH1 DNA-binding domain, a linker and an MH2 domain ending in the receptor-phosphorylated SSVS motif. It mediates TGF-beta/activin/nodal signals by forming complexes with SMAD4 that regulate transcription. smad3b is the faster-evolving copy: it differs from human SMAD3 mainly in the linker and carries asparagine at the position of the CK1 phosphosite S418, while keeping the Zn-binding, DNA-binding and receptor-phosphorylation residues. It is maternally supplied and zygotically expressed at the gastrula margin, tail bud, lateral mesoderm, brain regions, retina, spinal cord, somites and heart; unlike smad3a it is not expressed with a circadian rhythm. Overexpressed smad3b induces mesoderm and organizer genes such as goosecoid, and constitutively active Smad3b increases cardiomyocyte proliferation and activates Smad3 target genes. Knockdown of smad3b, like smad3a, alters neural differentiation. Combined loss of smad3a and smad3b enlarges the ventral aorta but is compatible with adult survival.' existing_annotations: - term: id: GO:0000976 label: transcription cis-regulatory region binding evidence_type: IEA original_reference_id: GO_REF:0000117 qualifier: enables supporting_entities: - ARBA:ARBA00028970 review: summary: ARBA electronic transfer of cis-regulatory region binding, a parent of the SMAD-binding element binding captured by the IBA to GO:0000978. action: ACCEPT reason: The MH1 domain of smad3b keeps the Zn-coordinating residues and K40/K41 of human SMAD3 (94% MH1 identity), and a Smad3 reporter built on SMAD-binding elements responds to Smad3b. Correct though general. supported_by: - reference_id: PMID:25286120 supporting_text: Reporter fluorescence is activated in phospho-Smad3 positive cells and is responsive to both Smad3 isoforms, Smad3a and 3b. - term: id: GO:0000978 label: RNA polymerase II cis-regulatory region sequence-specific DNA binding evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: enables supporting_entities: - FB:FBgn0011648 - FB:FBgn0288966 - MGI:MGI:109452 - MGI:MGI:1328787 - MGI:MGI:894293 - PANTHER:PTN000344419 - UniProtKB:P84022 - UniProtKB:Q13485 - UniProtKB:Q15796 - UniProtKB:Q15797 - UniProtKB:Q99717 - ZFIN:ZDB-GENE-990603-7 review: summary: PAINT inference from the SMAD family node that R-SMADs bind RNA polymerase II cis-regulatory DNA. action: ACCEPT reason: smad3b is inside the node and keeps every annotated MH1 functional residue of human SMAD3 (smad3a-bioinformatics). No smad3b-specific loss is known. supported_by: - reference_id: file:DANRE/smad3a/smad3a-bioinformatics/RESULTS.md supporting_text: Conserved in both copies and in gar - term: id: GO:0000981 label: DNA-binding transcription factor activity, RNA polymerase II-specific evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: enables supporting_entities: - FB:FBgn0011648 - FB:FBgn0288966 - MGI:MGI:108051 - MGI:MGI:109452 - MGI:MGI:1201674 - MGI:MGI:1328787 - MGI:MGI:894293 - PANTHER:PTN000344419 - UniProtKB:P84022 - UniProtKB:Q13485 - UniProtKB:Q15796 - UniProtKB:Q15797 - ZFIN:ZDB-GENE-990603-7 review: summary: PAINT inference that SMAD proteins are sequence-specific RNA polymerase II transcription factors. action: ACCEPT reason: Core function. Smad3b activates a Smad3 reporter and target genes (constitutively active Smad3b raises Smad3 target-gene expression in heart). It was weaker than Smad3a on the myf5 promoter, a quantitative difference that does not remove the activity. supported_by: - reference_id: PMID:25286120 supporting_text: Reporter fluorescence is activated in phospho-Smad3 positive cells and is responsive to both Smad3 isoforms, Smad3a and 3b. - reference_id: PMID:29196619 supporting_text: an upregulation in Smad3 target gene expression in caSmad3a and caSmad3b hearts - term: id: GO:0005515 label: protein binding evidence_type: IPI original_reference_id: PMID:16890162 qualifier: enables supporting_entities: - ZFIN:ZDB-GENE-031030-4 review: summary: Interaction with Tob1a, which the paper reports inhibits Smad3 transcriptional activity. action: REMOVE reason: Generic protein binding carries no functional information. The Tob1a interaction is not disputed, but the cached abstract gives no evidence for a more informative Smad3b activity term. Treated identically on smad3a. supported_by: - reference_id: PMID:16890162 supporting_text: Although Tob1a can also inhibit the transcriptional activity of the Nodal effector Smad3, its role in limiting dorsal development is executed primarily by antagonizing the beta-catenin signal. - term: id: GO:0005634 label: nucleus evidence_type: IEA original_reference_id: GO_REF:0000044 qualifier: located_in supporting_entities: - UniProtKB-SubCell:SL-0191 review: summary: Nuclear location mapped from the UniProt subcellular-location rule for SMADs. action: ACCEPT reason: Activated R-SMADs act in the nucleus; Smad3b drives nuclear reporter and target-gene transcription. - term: id: GO:0005667 label: transcription regulator complex evidence_type: IEA original_reference_id: GO_REF:0000002 qualifier: part_of supporting_entities: - InterPro:IPR013019 review: summary: InterPro2GO mapping from the MH1 domain. action: ACCEPT reason: R-SMADs act in transcription regulator complexes with SMAD4; the IBA to heteromeric SMAD protein complex is more specific. - term: id: GO:0005737 label: cytoplasm evidence_type: IEA original_reference_id: GO_REF:0000044 qualifier: located_in supporting_entities: - UniProtKB-SubCell:SL-0086 review: summary: Cytoplasmic location mapped from the UniProt subcellular-location rule. action: ACCEPT reason: Unstimulated R-SMADs are cytoplasmic and shuttle to the nucleus after receptor phosphorylation. - term: id: GO:0006355 label: regulation of DNA-templated transcription evidence_type: IEA original_reference_id: GO_REF:0000002 qualifier: involved_in supporting_entities: - InterPro:IPR001132 - InterPro:IPR003619 - InterPro:IPR013019 - InterPro:IPR013790 review: summary: InterPro2GO mapping from the SMAD/MH1/MH2 domains. action: ACCEPT reason: Correct but general; the IBA to positive regulation of transcription by RNA polymerase II is more specific. - term: id: GO:0007179 label: transforming growth factor beta receptor signaling pathway evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: involved_in supporting_entities: - MGI:MGI:108051 - MGI:MGI:1201674 - PANTHER:PTN002672225 - RGD:3031 - RGD:3032 - UniProtKB:P84022 - UniProtKB:Q15796 - ZFIN:ZDB-GENE-000509-3 - ZFIN:ZDB-GENE-990603-7 review: summary: PAINT inference at the SMAD2/3 node; smad3a (ZFIN:ZDB-GENE-000509-3) is among the descendant evidence. action: ACCEPT reason: Core process. The receptor-phosphorylated C-terminal SSVS motif is conserved in smad3b, and the Smad3 reporter that responds to Smad3b is blocked by ALK4/ALK5 inhibitors. supported_by: - reference_id: PMID:25286120 supporting_text: Moreover, Alk4 and Alk5 inhibitors strongly repress the reporter activity. - term: id: GO:0009653 label: anatomical structure morphogenesis evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: involved_in supporting_entities: - FB:FBgn0011648 - FB:FBgn0025800 - FB:FBgn0288966 - MGI:MGI:108051 - MGI:MGI:109452 - MGI:MGI:1100518 - MGI:MGI:1201674 - MGI:MGI:1336883 - MGI:MGI:894293 - PANTHER:PTN000344419 - RGD:620158 - RGD:71004 - UniProtKB:A0A8J0R1N9 - UniProtKB:A0A8J0T288 - UniProtKB:A0A8J1IQ47 - UniProtKB:O43541 - UniProtKB:Q6IRR4 - WB:WBGene00004856 - WB:WBGene00004857 - WB:WBGene00004858 - Xenbase:XB-GENE-1018193 - Xenbase:XB-GENE-482930 - Xenbase:XB-GENE-484686 - Xenbase:XB-GENE-485691 - Xenbase:XB-GENE-486004 - ZFIN:ZDB-GENE-000509-3 - ZFIN:ZDB-GENE-990603-7 - ZFIN:ZDB-GENE-990603-8 - ZFIN:ZDB-GENE-990603-9 review: summary: Very broad family-level PAINT process. action: KEEP_AS_NON_CORE reason: Not wrong for a SMAD, but uninformative and describes downstream outcomes rather than what smad3b does. - term: id: GO:0009880 label: embryonic pattern specification evidence_type: IEA original_reference_id: GO_REF:0000117 qualifier: involved_in supporting_entities: - ARBA:ARBA00084933 review: summary: ARBA transfer of embryonic patterning. action: KEEP_AS_NON_CORE reason: Smad3b overexpression induces mesoderm and organizer genes, and dominant-negative Smad3b blocks mesendoderm induction and neural induction; these reagents act on Smad2/3 signalling generally. Plausible but non-core. supported_by: - reference_id: PMID:12112463 supporting_text: We show that zebrafish Smad3b, in contrast to the related zebrafish Smad2, can induce mesoderm independently of TGF-beta signaling. - reference_id: PMID:19580801 supporting_text: Collectively, these data indicate that Smad2/3 activities are required for neural induction and neuroectodermal posteriorization in zebrafish. - term: id: GO:0009953 label: dorsal/ventral pattern formation evidence_type: IEA original_reference_id: GO_REF:0000117 qualifier: involved_in supporting_entities: - ARBA:ARBA00027261 review: summary: ARBA transfer of dorsal/ventral patterning, a SMAD family-wide rule dominated by BMP-pathway SMADs. action: MARK_AS_OVER_ANNOTATED reason: Dorsal/ventral patterning in zebrafish is driven mainly by BMP-SMAD1/5. Smad3b activates goosecoid when overexpressed, but no smad3b loss-of-function evidence shows a dorsal/ventral patterning defect. Treated identically on smad3a. supported_by: - reference_id: PMID:12112463 supporting_text: Although mammalian Smad3 was shown to inhibit expression of the organizer-specific genes goosecoid, zebrafish smad3b activates organizer genes such as goosecoid. - term: id: GO:0030154 label: cell differentiation evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: involved_in supporting_entities: - FB:FBgn0011648 - FB:FBgn0025800 - FB:FBgn0288966 - MGI:MGI:108051 - MGI:MGI:109452 - MGI:MGI:1201674 - MGI:MGI:1328787 - MGI:MGI:1859993 - MGI:MGI:894293 - PANTHER:PTN000344419 - RGD:3032 - RGD:3033 - UniProtKB:O15198 - UniProtKB:O43541 - UniProtKB:Q13485 - UniProtKB:Q15797 - UniProtKB:Q99717 - ZFIN:ZDB-GENE-000509-3 - ZFIN:ZDB-GENE-031014-1 - ZFIN:ZDB-GENE-990603-7 - ZFIN:ZDB-GENE-990603-8 - ZFIN:ZDB-GENE-991119-8 review: summary: Broad family-level PAINT process. action: KEEP_AS_NON_CORE reason: Correct in general (smad3b knockdown alters neural differentiation) but uninformative. supported_by: - reference_id: PMID:25286120 supporting_text: Similarly, smad3a and 3b knock-down alter neural differentiation showing that both paralogues play a positive role in neural differentiation. - term: id: GO:0030509 label: BMP signaling pathway evidence_type: IEA original_reference_id: GO_REF:0000117 qualifier: involved_in supporting_entities: - ARBA:ARBA00086900 review: summary: ARBA transfer of BMP signalling, a SMAD family-wide rule. action: MARK_AS_OVER_ANNOTATED reason: SMAD3 is an R-SMAD of the TGF-beta/activin/nodal branch; in zebrafish the Smad3 reporter that responds to Smad3b is blocked by ALK4/ALK5 inhibitors. No evidence places smad3b downstream of BMP receptors. Treated identically on smad3a. supported_by: - reference_id: PMID:25286120 supporting_text: Moreover, Alk4 and Alk5 inhibitors strongly repress the reporter activity. - term: id: GO:0032924 label: activin receptor signaling pathway evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: involved_in supporting_entities: - FB:FBgn0025800 - PANTHER:PTN001008908 - UniProtKB:P84022 - UniProtKB:Q15796 review: summary: PAINT inference at the SMAD2/3 node for activin signalling. action: ACCEPT reason: SMAD2/3 are the R-SMADs of the activin/nodal branch; dominant-negative Smad3b blocks nodal-induced mesendoderm, and smad3b keeps the receptor-phosphorylation motif. supported_by: - reference_id: PMID:18025082 supporting_text: Overexpression of these mutants abolished mesendoderm induction by ectopic Nodal signaling in zebrafish embryos. - term: id: GO:0045944 label: positive regulation of transcription by RNA polymerase II evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: involved_in supporting_entities: - FB:FBgn0025800 - MGI:MGI:108051 - MGI:MGI:1201674 - PANTHER:PTN001008908 - RGD:3031 - RGD:3032 - UniProtKB:P84022 - UniProtKB:Q15796 - WB:WBGene00000904 review: summary: PAINT inference at the SMAD2/3 node. action: ACCEPT reason: Smad3b activates transcription of organizer and Smad3 target genes; it is inside the node with no loss of functional residues. supported_by: - reference_id: PMID:29196619 supporting_text: an upregulation in Smad3 target gene expression in caSmad3a and caSmad3b hearts - term: id: GO:0060395 label: SMAD protein signal transduction evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: involved_in supporting_entities: - FB:FBgn0011648 - FB:FBgn0025800 - FB:FBgn0288966 - MGI:MGI:108051 - MGI:MGI:109452 - MGI:MGI:1100518 - MGI:MGI:1201674 - MGI:MGI:1328787 - MGI:MGI:1859993 - MGI:MGI:894293 - PANTHER:PTN000344419 - RGD:3031 - RGD:3032 - UniProtKB:O15105 - UniProtKB:P84022 - UniProtKB:Q13485 - UniProtKB:Q15796 - UniProtKB:Q15797 - UniProtKB:Q9W734 review: summary: PAINT inference at the SMAD family node. action: ACCEPT reason: Core process for an R-SMAD. - term: id: GO:0070411 label: I-SMAD binding evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: enables supporting_entities: - PANTHER:PTN000344419 - UniProtKB:O15105 - UniProtKB:O43541 - UniProtKB:Q13485 - UniProtKB:Q15796 - UniProtKB:Q15797 review: summary: PAINT inference that SMADs bind inhibitory SMADs (SMAD6/7). action: ACCEPT reason: MH2 is 97.9% identical to human SMAD3; zebrafish smad7 is regulated by Smad3. No smad3b-specific loss is known. supported_by: - reference_id: PMID:12112463 supporting_text: Furthermore, we show that Smad3 and BMP signals activate smad7. - term: id: GO:0071144 label: heteromeric SMAD protein complex evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: part_of supporting_entities: - FB:FBgn0011648 - FB:FBgn0025800 - FB:FBgn0288966 - PANTHER:PTN000344419 - UniProtKB:P84022 - UniProtKB:Q13485 - UniProtKB:Q15796 - UniProtKB:Q15797 review: summary: PAINT inference that R-SMADs form heteromeric complexes with SMAD4. action: ACCEPT reason: Conserved mechanism; smad3b keeps the MH2 domain and C-terminal SSVS motif. No smad3b-specific data. references: - id: GO_REF:0000002 title: Gene Ontology annotation through association of InterPro records with GO terms findings: [] - id: GO_REF:0000033 title: Annotation inferences using phylogenetic trees findings: [] - id: GO_REF:0000044 title: Gene Ontology annotation based on UniProtKB/Swiss-Prot Subcellular Location vocabulary mapping, accompanied by conservative changes to GO terms applied by UniProt findings: [] - id: GO_REF:0000117 title: Electronic Gene Ontology annotations created by ARBA machine learning models findings: [] - id: PMID:16890162 title: Tob1 controls dorsal development of zebrafish embryos by antagonizing maternal beta-catenin transcriptional activity. findings: [] reference_review: relevance: LOW correctness: VERIFIED review_notes: Abstract only. Tob1a inhibits Smad3 transcriptional activity; source of the generic protein-binding IPI rows on both smad3 paralogs. - id: PMID:12112463 title: Zebrafish Smad7 is regulated by Smad3 and BMP signals. findings: - statement: Smad3b induces mesoderm and activates organizer genes such as goosecoid when overexpressed. supporting_text: We show that zebrafish Smad3b, in contrast to the related zebrafish Smad2, can induce mesoderm independently of TGF-beta signaling. reference_review: relevance: HIGH correctness: VERIFIED review_notes: Abstract only; first description of smad3b, its expression and overexpression activity. - id: PMID:18025082 title: smad2 and smad3 are required for mesendoderm induction by transforming growth factor-beta/nodal signals in zebrafish. findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: Abstract only; dominant-negative Smad2, Smad3a and Smad3b block nodal-induced mesendoderm. - id: PMID:19580801 title: Smad2/3 activities are required for induction and patterning of the neuroectoderm in zebrafish. findings: [] reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: Abstract only; dnsmad3b and smad3b overexpression used as general Smad2/3 tools in neural induction. - id: PMID:21159776 title: dickkopf-3-related gene regulates the expression of zebrafish myf5 gene through phosphorylated p38a-dependent Smad4 activity. findings: - statement: Smad3b plays only a minor role in myf5 promoter activation compared with Smad2 and Smad3a. supporting_text: Although the ability of Smad2 and Smad3a to activate the myf5 promoter requires Smad4 to form a complex and enter the nucleus, Smad3b only plays a minor role in myf5 promoter activation. reference_review: relevance: HIGH correctness: VERIFIED review_notes: Full text; side-by-side comparison where smad3b, unlike smad3a, did not activate the myf5 promoter. - id: PMID:25286120 title: A Smad3 transgenic reporter reveals TGF-beta control of zebrafish spinal cord development. findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: Abstract only; reporter responds to both paralogs; both knockdowns alter neural differentiation. - id: PMID:27703851 title: 'Evolution history of duplicated smad3 genes in teleost: insights from Japanese flounder, Paralichthys olivaceus.' findings: - statement: Teleost smad3b evolves faster than smad3a and has linker sites under positive selection. supporting_text: Five candidate positive selected sites were identified, two of which were significantly positively selected (219L**, 221L**, posterior probability > 0.99) in smad3b. reference_review: relevance: HIGH correctness: VERIFIED review_notes: Full text; synteny/phylogeny place the duplication at the TGD; faster smad3b evolution and linker sites under positive selection in teleost smad3b. - id: PMID:29196619 title: Opposite effects of Activin type 2 receptor ligands on cardiomyocyte proliferation during development and repair. findings: [] reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: Full text; caSmad3a and caSmad3b both increase cardiomyocyte proliferation (70% and 31%). - id: PMID:29940038 title: Interactions between the circadian clock and TGF-β signaling pathway in zebrafish. findings: [] reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: Full text; smad3b is not rhythmically expressed, unlike smad3a. - id: PMID:40066353 title: 'Modeling thoracic aortic genetic variants in the zebrafish: useful for predicting clinical pathogenicity?' findings: [] reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: Review; both smad3 ohnologs had to be targeted because of redundancy (single-mutant data not shown). - id: PMID:42584512 title: Smad3 deficiency attenuates disease severity in a zebrafish model for Smad6-related aortic disease. findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: Abstract only; smad3a/b double knockout enlarges ventral aorta and survives to adulthood. - id: file:DANRE/smad3a/smad3a-bioinformatics/RESULTS.md title: smad3a/smad3b/human SMAD3/gar SMAD3 protein comparison and Bgee/ZFIN expression comparison reference_review: relevance: HIGH correctness: VERIFIED review_notes: Manual audit verified the sidecar supports both copies as SMAD3 orthologs retaining the MH1/MH2 domains and receptor-phosphorylated tail, with smad3b linker divergence that remains biochemically untested. findings: - statement: smad3b keeps all annotated functional residues of human SMAD3 except S418 (N); its differences concentrate in the linker. supporting_text: Conserved in both copies and in gar core_functions: - description: Receptor-regulated SMAD of the TGF-beta/activin/nodal branch, forming heteromeric complexes with SMAD4 that activate RNA polymerase II target genes. Molecular function shared with smad3a. molecular_function: id: GO:0000981 label: DNA-binding transcription factor activity, RNA polymerase II-specific directly_involved_in: - id: GO:0007179 label: transforming growth factor beta receptor signaling pathway - id: GO:0032924 label: activin receptor signaling pathway locations: - id: GO:0005634 label: nucleus - id: GO:0005737 label: cytoplasm in_complex: id: GO:0071144 label: heteromeric SMAD protein complex supported_by: - reference_id: PMID:25286120 supporting_text: Reporter fluorescence is activated in phospho-Smad3 positive cells and is responsive to both Smad3 isoforms, Smad3a and 3b. - reference_id: PMID:29196619 supporting_text: an upregulation in Smad3 target gene expression in caSmad3a and caSmad3b hearts - reference_id: PMID:18025082 supporting_text: We generated potent and specific dominant-negative forms of zebrafish Smad2, Smad3a, and Smad3b by mutating multiple amino acids. suggested_questions: - question: Do smad3b single mutants have any phenotype, and is smad3a upregulated in smad3b PTC mutants (transcriptional adaptation)? - question: Does the S418N substitution or the diverged linker change Smad3b activity or its regulation by CK1/MAPK/CDK phosphorylation? - question: Is the brain and spinal cord expression of smad3b (Pogoda and Meyer 2002) shared by smad3a, which has not been surveyed at the same resolution? suggested_experiments: - hypothesis: Smad3b is transcriptionally weaker than Smad3a because of linker divergence. description: Equal-dose SBE-reporter and target-gene assays of Smad3a, Smad3b, Smad3b N418S and linker-swapped chimeras in smad3a;smad3b double mutant embryos. experiment_type: cross-rescue - hypothesis: The two copies overlap in expression at cellular resolution. description: Double fluorescent in situ or single-cell RNA-seq comparison of smad3a and smad3b across embryonic and adult tissues. experiment_type: expression analysis