CG5565 encodes a 240-residue haloacid dehalogenase (HAD) subfamily IA phosphatase of the Gpp1/Gpp2-like (PTHR18901) family, with an intact N-terminal DxD catalytic motif (Asp16/Asp18). It is one of five Drosophila paralogs (with Gs1l, CG5561, CG5556 and CG5564) produced by a dipteran-lineage duplication within the eukaryotic clade that includes human and mouse PUDP and yeast YKL033W-A. Human PUDP is a highly selective pseudouridine 5'-phosphatase, which dephosphorylates pseudouridine 5'-phosphate released by turnover of pseudouridine-containing RNA so that free pseudouridine can be excreted. CG5565 is therefore predicted to have the same activity, but the fly protein has not been assayed, and whether all five paralogs retain this substrate preference is unknown. CG5565 transcripts are strongly testis-enriched and the protein has been detected in the sperm proteome; no loss-of-function phenotype linked to its enzymatic activity has been reported.
Summary: PAINT places generic phosphatase activity at the eukaryotic node PTN000431251, from which CG5565 descends. The term is correct, but CG5565 also descends from the Unikont node PTN000431252, which carries the more specific pseudouridine 5'-phosphatase IBD seeded by human PUDP and yeast YKL033W-A.
Reason: The broad phosphatase IBD is phylogenetically sound and CG5565 retains the HAD catalytic Asp pair, so the claim holds. It is less specific than the evidence allows. A PANTHER 19 tree walk places CG5565 under PTN000431252, the node bearing the GO:1990738 IBD (dated 20260729 and not yet propagated to GOA, which is why this snapshot carries only the PTN000431251 term). GO:1990738 is_a phosphatase activity and the gene already carries it by FlyBase ISS, so refine to it. The plant glycerol-3-phosphatase IBD placed in the bikont lineage (PTN008360158) is not inherited by CG5565 and is deliberately not proposed.
Propagation Review
Root cause:TERM SCOPING PROBLEM
Failure modes:GRANULARITY MISMATCH
Sources checked:
PANTHER:PTN000431251SUPPORTS TRANSFER
The eukaryote-level phosphatase IBD is compatible with CG5565's intact HAD-IA scaffold. The only issue is granularity; the more specific descendant-node IBD (PTN000431252, GO:1990738) also covers CG5565 but has not yet reached the GOA snapshot.
CG5565 (FlyBase:FBgn0031335) descends from PTN000431252 (Unikonts), the node carrying the PAINT IBD for GO:1990738 pseudouridine 5'-phosphatase activity seeded by human PUDP and yeast YKL033W-A.
Purified human recombinant HDHD1 dephosphorylated pseudouridine 5'-phosphate with a kcat of 1.6 s-1, a Km of 0.3 ฮผM and a catalytic efficiency at least 1000-fold higher than that on which it acted on other phosphate esters, including 5'-UMP.
Summary: InterPro2GO mapping from the Gpp1/Gpp2-like family signature (IPR045228). The HAD phosphatase class is correct for CG5565, but the term is less specific than the pseudouridine 5'-phosphatase activity supported by phylogenetic placement.
Reason: Same claim as the IBA row and must be treated consistently. The family-level phosphatase assignment is correct; refine to GO:1990738, which CG5565 already carries (ISS) and which is supported by its descent from the PTN000431252 PUDP node.
CG5565 (FlyBase:FBgn0031335) descends from PTN000431252 (Unikonts), the node carrying the PAINT IBD for GO:1990738 pseudouridine 5'-phosphatase activity seeded by human PUDP and yeast YKL033W-A.
Summary: FlyBase transfer from human PUDP (Q08623), whose purified recombinant enzyme is kinetically selective for pseudouridine 5'-phosphate. The transfer is phylogenetically grounded. CG5565 and human PUDP both descend from PANTHER node PTN000431252, where PAINT curators independently placed the same function.
Reason: The donor activity is directly measured (PMID:20722631). The relationship is established by tree placement, not similarity alone. CG5565 is a co-ortholog of human PUDP through a dipteran duplication (PTN000431298) and lies below the PTN000431252 IBD for GO:1990738. The SF38 (fly) versus SF43 (human) subfamily labels do not argue against transfer, because SF43 arises only after a recent Catarrhini duplication and mouse Pudp is SF38. The catalytic Asp pair is conserved. Caveat - no fly paralog has been assayed, and post-duplication substrate divergence among the five fly paralogs cannot be excluded.
Purified human recombinant HDHD1 dephosphorylated pseudouridine 5'-phosphate with a kcat of 1.6 s-1, a Km of 0.3 ฮผM and a catalytic efficiency at least 1000-fold higher than that on which it acted on other phosphate esters, including 5'-UMP.
CG5565 (FlyBase:FBgn0031335) descends from PTN000431252 (Unikonts), the node carrying the PAINT IBD for GO:1990738 pseudouridine 5'-phosphatase activity seeded by human PUDP and yeast YKL033W-A.
The SF38 (CG5565) versus SF43 (human PUDP) label difference therefore reflects a recent primate-lineage split, not a deep divergence separating the fly gene from the characterised mammalian enzyme.
Human annotated catalytic Asp14 and Asp16 map to target Asp16 and Asp18.
Core Functions
Predicted hydrolysis of pseudouridine 5'-phosphate to pseudouridine and phosphate, the activity of the human and yeast PUDP enzymes. CG5565 inherits it through descent from the PANTHER Unikont node PTN000431252 and retains the HAD catalytic Asp pair. The fly protein has not been assayed.
Purified human recombinant HDHD1 dephosphorylated pseudouridine 5'-phosphate with a kcat of 1.6 s-1, a Km of 0.3 ฮผM and a catalytic efficiency at least 1000-fold higher than that on which it acted on other phosphate esters, including 5'-UMP.
CG5565 (FlyBase:FBgn0031335) descends from PTN000431252 (Unikonts), the node carrying the PAINT IBD for GO:1990738 pseudouridine 5'-phosphatase activity seeded by human PUDP and yeast YKL033W-A.
Q: Have all five dipteran PUDP-clade paralogs (CG5565, Gs1l, CG5561, CG5556, CG5564) retained pseudouridine 5'-phosphate selectivity after the PTN000431298 duplication, or has the strongly testis-enriched CG5565 diverged in substrate preference?
Suggested Experiments
Experiment: Express recombinant CG5565 and assay phosphatase activity against pseudouridine 5'-phosphate, UMP and other sugar or nucleotide phosphates, in parallel with Gs1l, to obtain kinetic constants comparable to those measured for human PUDP.
Type: in vitro enzyme kinetics
Experiment: Measure pseudouridine 5'-phosphate and free pseudouridine in testis and whole-fly extracts from CG5565 loss-of-function or RNAi animals, alone and combined with paralog knockdown, to test whether CG5565 contributes to pseudouridine 5'-phosphate clearance in vivo.
Type: metabolomics
External Prediction Reviews
These computational predictions are reviewed separately from the GOA annotation set used for this review. The assessments below are from this project and do not constitute official GO annotations or endorsement by GO/UniProt. They are not included in the existing annotation review above.
The ProtNLM2 API snapshot retrieved 2026-09-08 contains no GO-term predictions for Q9VQ04. The re-reviewed gene supports pseudouridine 5'-phosphatase activity (GO:1990738), already carried in GOA by FlyBase ISS transfer from experimentally characterized human PUDP and by the PAINT IBD at PANTHER node PTN000431252, under which CG5565 descends; the broader phosphatase activity (GO:0016791) is also already annotated. An emitted prediction of either term would therefore not be novel. The near-complete alignment to human PUDP with retained catalytic Asp16/Asp18 supports the HAD phosphatase activity, but the substrate assignment is an orthology-based inference, not a direct CG5565 assay, and no CG5565 publication was found in PubMed. The separate ProtNLM2 FUNCTION text claiming in-vitro D,L-glyceraldehyde 3-phosphate dephosphorylation is UNC and is not disproved by the better-supported physiological substrate. This completed record assesses the absence of GO output; predictions is empty because no GO or EC prediction was emitted.