id: A4QNX6 gene_symbol: ctdspl2b product_type: PROTEIN status: COMPLETE taxon: id: NCBITaxon:7955 label: Danio rerio description: ctdspl2b (CTD small phosphatase-like 2b) is one of two zebrafish co-orthologs of vertebrate CTDSPL2 (SCP4), a nuclear protein phosphatase of the haloacid dehalogenase (HAD) superfamily; its paralog is ctdspl2a. The protein has a disordered N-terminal regulatory region, which in the human protein carries nuclear localization sequences, and a C-terminal FCP1-homology phosphatase domain with the magnesium-dependent DxDx(T/V) catalytic motif. Human and mouse CTDSPL2 dephosphorylate serine and threonine residues on several nuclear substrates, including the RNA polymerase II C-terminal domain (Ser5-preferential), FoxO1/FoxO3a (promoting their nuclear retention and target-gene transcription, including gluconeogenic genes), Smad1/5/8 (attenuating BMP signalling) and mitotic histone H3 Thr3. The zebrafish protein keeps the catalytic motif (DLDET) and a phosphatase domain 96.2% identical to human CTDSPL2, and is 70.9% identical to ctdspl2a overall. ctdspl2b is maternally supplied and expressed throughout development and in all adult tissues sampled, in the same pattern as ctdspl2a but at two- to three-fold lower levels. No functional study of the zebrafish gene has been published. existing_annotations: - term: id: GO:0004721 label: phosphoprotein phosphatase activity evidence_type: IBA original_reference_id: GO_REF:0000033 qualifier: enables supporting_entities: - AGI_LocusCode:AT5G46410 - FB:FBgn0029067 - FB:FBgn0036556 - MGI:MGI:1196405 - PANTHER:PTN000258719 - SGD:S000001046 - SGD:S000003933 - UniProtKB:O14595 - UniProtKB:O95476 - UniProtKB:Q05D32 - UniProtKB:Q3ZCQ8 - UniProtKB:Q57ZA0 - UniProtKB:Q9GZU7 review: summary: Phosphoprotein phosphatase activity, propagated from the Dullard/SCP family node PTN000258719, whose experimental sources include human CTDSPL2 itself (Q05D32). action: ACCEPT reason: The function is ancestral to the family (FCP/SCP, Dullard/CTDNEP1, TIMM50 and plant and yeast homologs are all protein phosphatases), and ctdspl2b keeps every feature needed for it. The catalytic DxDx(T/V) motif is present as DLDET, aligned exactly to human D293-T297, and the FCP1-homology domain is 96.2% identical to human CTDSPL2. Mutating either aspartate of that motif abolishes human SCP4 function. This is the core molecular function of the gene; a more specific child (protein serine/threonine phosphatase activity) is recorded in core_functions. supported_by: - reference_id: PMID:35021089 supporting_text: SCP4 belongs to the haloacid dehalogenase (HAD) family of proteins, which employ conserved active site aspartates within a DxDx(V/T) motif to remove phosphates from protein or non-protein substrates - reference_id: PMID:35021089 supporting_text: Despite similar expression levels to the wild-type protein, SCP4D293A, SCP4D295A, and SCP4D293A/D295A mutants were unable to support MOLM-13 growth (Figures 3G and 3H). - reference_id: file:DANRE/ctdspl2a/ctdspl2a-bioinformatics/RESULTS.md supporting_text: Both copies keep the DxDx(T/V) catalytic motif (DLDET) and a phosphatase domain at least 96% identical to human CTDSPL2. - term: id: GO:0004721 label: phosphoprotein phosphatase activity evidence_type: IEA original_reference_id: GO_REF:0000117 qualifier: enables supporting_entities: - ARBA:ARBA00027651 review: summary: Phosphoprotein phosphatase activity from an ARBA machine-learning rule. action: ACCEPT reason: Same assertion as the IBA row, consistent with the conserved catalytic motif and phosphatase domain. Duplicate rows with different evidence are acceptable. supported_by: - reference_id: file:DANRE/ctdspl2a/ctdspl2a-bioinformatics/RESULTS.md supporting_text: Both copies keep the DxDx(T/V) catalytic motif (DLDET) and a phosphatase domain at least 96% identical to human CTDSPL2. - term: id: GO:0005634 label: nucleus evidence_type: IEA original_reference_id: GO_REF:0000117 qualifier: located_in supporting_entities: - ARBA:ARBA00026330 review: summary: Nucleus from an ARBA rule. action: ACCEPT reason: Human and mouse CTDSPL2 are nuclear, chromatin-associated phosphatases, and their N-terminal segment carries importin-binding nuclear localization sequences. Its best-characterized substrates (Pol II CTD, FoxO1/3a, Smad1/5/8, histone H3) are nuclear proteins. The zebrafish N-terminal region is less conserved (47% identical) than the phosphatase domain, so nuclear targeting is an inference, but a well supported one for the family. supported_by: - reference_id: PMID:26920047 supporting_text: Therefore, SCP4 is a unique chromatin-associated, Ser5-preferential CTD phosphatase that preferentially distributes to transcriptionally silenced gene regions and may participate in gene regulation during erythroid differentiation. - reference_id: PMID:35021089 supporting_text: This result suggests that a function of the SCP4 N-terminal segment is to promote nuclear localization of the full-length protein. - term: id: GO:0016791 label: phosphatase activity evidence_type: IEA original_reference_id: GO_REF:0000002 qualifier: enables supporting_entities: - InterPro:IPR011948 review: summary: Phosphatase activity from the InterPro Dullard-like phosphatase domain (IPR011948) mapping. action: ACCEPT reason: Correct but general; the domain-level mapping cannot say more than phosphatase activity. The protein-level term (phosphoprotein phosphatase activity) is carried by the IBA row, so this IEA adds no error. supported_by: - reference_id: PMID:26674342 supporting_text: It is well known that CTDSPL2 is a nuclear protein containing FCPH domain and DXDX(T/V) motif. - term: id: GO:0045722 label: positive regulation of gluconeogenesis evidence_type: IEA original_reference_id: GO_REF:0000117 qualifier: involved_in supporting_entities: - ARBA:ARBA00084180 review: summary: Positive regulation of gluconeogenesis from an ARBA rule, reflecting the mouse and human finding that SCP4 dephosphorylates FoxO1/3a in hepatocytes. action: KEEP_AS_NON_CORE reason: In mammals SCP4 promotes hepatic gluconeogenesis by dephosphorylating FoxO1/3a and increasing transcription of PEPCK1 and G6PC, and SCP4 knockout mice are hypoglycemic as neonates. That is a direct role (the phosphatase acts on the transcription factors that drive the gluconeogenic genes), not just a requirement. ctdspl2b is expressed in zebrafish liver, but the role has not been examined in fish, and it is one of several physiological roles of the enzyme (FoxO in muscle and cartilage, BMP-Smad, Pol II, mitosis). Kept as a plausible, non-core, homology-based process; the same call is made for ctdspl2a, which is also expressed in liver. supported_by: - reference_id: PMID:28851713 supporting_text: In this study, we discovered a nuclear phosphatase, SCP4/CTDSPL2 (SCP4), that dephosphorylated FoxO1/3a and promoted FoxO1/3a transcription activity. - reference_id: PMID:28851713 supporting_text: Moreover, we demonstrated that gene ablation of SCP4 led to hypoglycemia in neonatal mice. core_functions: - description: Nuclear HAD-family serine/threonine protein phosphatase (CTDSPL2/SCP4 ortholog) with a conserved DxDx(T/V) catalytic motif; by homology it dephosphorylates nuclear substrates such as the RNA polymerase II CTD, FoxO transcription factors and Smad1/5/8. Enzyme activity has not been measured for the zebrafish protein; the catalytic motif and phosphatase domain are shared with ctdspl2a. molecular_function: id: GO:0004721 label: phosphoprotein phosphatase activity locations: - id: GO:0005634 label: nucleus supported_by: - reference_id: PMID:26920047 supporting_text: SCP4 exhibited Ser5-preferential CTD phosphatase activity in vitro, while small interfering RNA-mediated SCP4 knockdown in HeLa cells increased phosphorylation levels of Pol II at Ser5 and Ser7, but not at Ser2. - reference_id: PMID:28506762 supporting_text: 'Our interest in SCP4 (also called CTDSPL2) was stimulated by the finding that SCP4 specifically dephosphorylates transcription factor Smad1/5/8; these reactions attenuate BMP-induced transcriptional activities [26;27].' - reference_id: PMID:42315649 supporting_text: Here, we identify the nuclear phosphatase SCP4 as a novel H3T3 phosphatase through a systematic phosphatase library screening. - reference_id: file:DANRE/ctdspl2a/ctdspl2a-bioinformatics/RESULTS.md supporting_text: Both copies keep the DxDx(T/V) catalytic motif (DLDET) and a phosphatase domain at least 96% identical to human CTDSPL2. 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:0000117 title: Electronic Gene Ontology annotations created by ARBA machine learning models findings: [] - id: PMID:26674342 title: A systematic study of nuclear interactome of C-terminal domain small phosphatase-like 2 using inducible expression system and shotgun proteomics. findings: [] reference_review: relevance: LOW correctness: VERIFIED review_notes: Human CTDSPL2 nuclear interactome; background statement on domain and motif. - id: PMID:26920047 title: Human SCP4 is a chromatin-associated CTD phosphatase and exhibits the dynamic translocation during erythroid differentiation. findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: Abstract only; in vitro Ser5-preferential CTD phosphatase activity; chromatin localization. - id: PMID:28506762 title: The nuclear phosphatase SCP4 regulates FoxO transcription factors during muscle wasting in chronic kidney disease. findings: [] reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: Mouse muscle; SCP4 domain organization (N-terminal NLS region, C-terminal phosphatase domain), FoxO and Smad1/5 dephosphorylation. - id: PMID:28851713 title: SCP4 Promotes Gluconeogenesis Through FoxO1/3a Dephosphorylation. findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: Abstract only in cache; source of the human IDA for positive regulation of gluconeogenesis and of the ARBA rule's biology. - id: PMID:35021089 title: SCP4-STK35/PDIK1L complex is a dual phospho-catalytic signaling dependency in acute myeloid leukemia. findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: Human SCP4 catalytic aspartates D293/D295 in the DxDx(V/T) motif; N-terminal importin-binding NLS region. - id: PMID:42315649 title: SCP4 dephosphorylates mitotic histone H3 to maintain chromosome stability. findings: [] reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: Human SCP4 as histone H3 Thr3 phosphatase; D295N phosphatase-dead mutant. - id: file:DANRE/ctdspl2a/ctdspl2a-bioinformatics/RESULTS.md title: ctdspl2a / ctdspl2b protein, expression and synteny comparison reference_review: relevance: HIGH correctness: VERIFIED review_notes: >- Local pair analysis directly supports CTDSPL2 orthology, retention of the DLDET catalytic motif and FCP1 homology domain in both paralogs, and broad co-expression without a resolved expression partition. findings: [] suggested_questions: - question: Do ctdspl2a and ctdspl2b act on the same nuclear substrates in zebrafish (Pol II CTD, FoxO, Smad1/5/9), and is the divergent N-terminal region of either copy still able to target it to the nucleus? - question: Is the combined ctdspl2a + ctdspl2b expression in zebrafish comparable to the single gar gene, as expected if the pair is retained for dosage? suggested_experiments: - hypothesis: ctdspl2a and ctdspl2b are functionally redundant phosphatases. description: Generate single and double mutants (RNA-less alleles where possible, to avoid transcriptional adaptation); test early BMP-dependent dorsoventral patterning, larval glucose levels under fasting, and cartilage development; test cross-rescue by mRNA injection of each copy and of catalytic-dead (D-to-N) forms. experiment_type: genetic interaction - hypothesis: Both zebrafish proteins keep CTD phosphatase activity. description: In vitro phosphatase assays of recombinant ctdspl2a and ctdspl2b on phospho-CTD and phospho-Smad1 peptides, with human CTDSPL2 as control. experiment_type: biochemical assay