PPP4R4

UniProt ID: A0A9L0S961
Organism: Equus caballus
Review Status: COMPLETE
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Gene Description

PPP4R4 (protein phosphatase 4 regulatory subunit 4) is a noncatalytic HEAT/ARM-repeat protein with weak similarity to the scaffolding A subunit of PP2A. In human cells it binds the catalytic subunit of protein phosphatase 4 (PP4c) specifically, and not PP2Ac or PP6c, to form a stable cytosolic PP4c-PP4R4 heterodimer. This complex is separate from the PP4c-PP4R1 and PP4c-PP4R2-PP4R3 complexes, and PP4R4 does not bridge PP4c to those subunits. PP4c bound to PP4R4 dephosphorylates model substrates less actively than free PP4c, so PPP4R4 modulates PP4 rather than providing phosphatase catalysis itself. Its physiological substrates and cellular role are not known. Horse PPP4R4 is the ortholog of human PPP4R4 in PANTHER subfamily PTHR21467:SF0. The UniProt entry A0A9L0S961 is a 865-residue alternative product of the horse gene. It matches the canonical horse product except for a short alternative C-terminus, and it keeps the HEAT repeats and the residues needed for PP4c binding.

Existing Annotations Review

GO Term Evidence Action Reason
GO:0005737 cytoplasm
IEA
GO_REF:0000044 		MODIFY
Summary: Correct, but the evidence supports the more specific cytosol. The human ortholog was isolated as a stable cytosolic PP4c partner, and PAINT places cytosol at PTN001087371, the node that contains horse PPP4R4.
Reason: This row comes from the UniProt SubCell mapping of an ARBA-predicted "Cytoplasm" line, so on its own it is a prediction. The claim is still sound for this protein, which sits in PTHR21467:SF0 with experimentally characterised human PPP4R4 (Q6NUP7). Human PPP4R4 was identified as a stable cytosolic PP4c partner (PMID:18715871). Later work describes PP4R1 and PP4R4 as the cytoplasmic PP4 regulatory subunits, unlike the nuclear PP4R2/PP4R3 (PMID:37605615). PAINT propagated GO:0005829 cytosol from node PTN001087371 to the canonical horse product A0A9L0S938. A0A9L0S961 differs from that product only in its last few residues, inside the disordered C-terminal tail. Cytosol (GO:0005829) is the specific location the evidence supports and is the location used in core_functions, so the general term is refined to it.
Proposed replacements: cytosol
Supporting Evidence:
PMID:18715871
we identify a novel, stable cytosolic PP4c interacting partner, KIAA1622, which we have renamed PP4R4.
PMID:37605615
PP4R2, PP4R3Ξ±, and PP4R3Ξ² are present in nuclei, while PP4R1 and PP4R4 are located in the cytoplasm
file:HORSE/PPP4R4/PPP4R4-bioinformatics/RESULTS-rereview.md
A0A9L0S961 and A0A9L0S938 are identical over residues 1-861. They differ only at the extreme C-terminus.
GO:0030289 protein phosphatase 4 complex
ISS
PMID:18715871
PP4R4/KIAA1622 forms a novel stable cytosolic complex with p... 		NEW
Summary: Proposed. PPP4R4 is a subunit of a PP4c-PP4R4 heterodimer, which by definition is a protein phosphatase 4 complex.
Reason: GO:0030289 is defined as "a protein serine/threonine phosphatase complex formed by the catalytic subunit of protein phosphatase 4 plus one or more regulatory subunits". Its only parent is GO:0008287, the term that PAINT (PTN001087371) and the human IDA use for PPP4R4. The comparators in the same role, the other PP4 regulatory subunits PPP4R1, PPP4R2 and PPP4R3A and the catalytic subunit PPP4C, all carry GO:0030289 in human GOA. So the specific term is the convention for PP4 subunits. PPP4R4 has the generic parent only because its annotations predate any choice of the specific child; no curator argued against it. Human PPP4R4 binds PP4c specifically and does not bind PP2Ac or PP6c (PMID:18715871). The proteomic study of PPP holoenzymes treats PP4c-PP4R4 as one of the PP4 holoenzymes and measures its formation directly (PMID:34845248). Horse PPP4R4 shares subfamily PTHR21467:SF0 and the conserved PP4c-binding residues. If GO:0030289 is not accepted, the PAINT term GO:0008287 is the fallback.
Supporting Evidence:
PMID:18715871
PP4R4 displays weak sequence homology with the A (scaffolding) subunit of the PP2A holoenzyme and specifically associates with PP4c (and not with the related PP2Ac or PP6c phosphatases).
PMID:34845248
PP4R2, PP4R3Ξ±, and PP4R4 were mostly unaffected by the reduction in methylation
file:HORSE/PPP4R4/PPP4R4-bioinformatics/RESULTS-rereview.md
Both residues are conserved at the same positions in A0A9L0S961, with identical local context

Core Functions

Noncatalytic regulatory subunit of protein phosphatase 4. It forms a stable cytosolic PP4c-PP4R4 heterodimer, separate from the other PP4 holoenzymes, and changes the activity of the bound catalytic subunit. Its physiological substrates are unknown.

Molecular Function:
protein phosphatase regulator activity
Cellular Locations:
cytosol
In Complex:
protein phosphatase 4 complex
Supporting Evidence:
  • PMID:18715871
    we identify a novel, stable cytosolic PP4c interacting partner, KIAA1622, which we have renamed PP4R4.
  • PMID:18715871
    PP4c.PP4R4 complexes exhibit phosphatase activity toward a fluorogenic substrate and gammaH2AX, but this activity is lower than that associated with the PP4c.PP4R2.PP4R3 complex, which itself is less active than the free PP4c catalytic subunit.
  • file:HORSE/PPP4R4/PPP4R4-bioinformatics/RESULTS-rereview.md
    Both residues are conserved at the same positions in A0A9L0S961, with identical local context

References

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Suggested Questions for Experts

Q: Which physiological substrates does the cytosolic PP4c-PP4R4 complex dephosphorylate, and does PP4R4 act mainly to target substrates or to restrain PP4c activity?

Q: Should PPP4R4's GO:0008287 annotations (human IDA, PAINT PTN001087371) be refined to GO:0030289 protein phosphatase 4 complex, as is done for the other PP4 regulatory subunits?

Suggested Experiments

Experiment: Phosphoproteomics after PPP4R4 knockout, compared with PPP4C depletion, in a cell type that expresses PPP4R4, to identify substrates specific to the PP4c-PP4R4 complex.

Type: phosphoproteomics

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.

ProtNLM2 External predictions

View prediction review YAML Β· PPP4R4-protnlm-predictions-review.yaml Β· Review status: COMPLETE

ProtNLM2 emitted three GO terms for the horse PPP4R4 alternative product A0A9L0S961. The molecular function (protein phosphatase regulator activity) and complex (protein serine/threonine phosphatase complex) predictions are correct, supported by the human PP4c-PP4R4 study and the PAINT node shared by horse PPP4R4, but both terms are already carried by IBA on the canonical horse product A0A9L0S938 of the same gene, so they are not novel. Blastocyst hatching rests only on a pooled RNAi screen of the mouse ortholog that scored a combined hatching and outgrowth defect; that necessity evidence does not show that PPP4R4 does any of the work of hatching, so the prediction is uncertain.

Source documents: projects/PROTNLM_EVALUATION/mammal-benchmark/horse40-predictions.csv Β· projects/PROTNLM_EVALUATION/mammal-benchmark/predictions.jsonl.gz Β· projects/PROTNLM_EVALUATION/mammal-benchmark/paired-sequences/PPP4R4.json

Review score: 2 = concordant with evidence; 1 = uncertain; 0 = discordant with evidence. This is an assessment score, not a model probability.

GO:0001835 blastocyst hatching GO_BP
UNC β€” Uncertain Review score: 1/2
Prediction method: ProtNLM2 Β· Version: UniProt API snapshot 2026-09-08
Review rationale: The only experimental link between PPP4R4 and this process is a pooled RNAi screen in mouse zygotes, in which knockdown of the ortholog Ppp4r4 (same PANTHER subfamily PTHR21467:SF0) was one of 35 knockdowns scored as defective in a combined hatching and outgrowth assay. That is a single unreplicated screen result, it measures necessity rather than participation, and the authors themselves flag that Ppp4r4 is not expressed at the stages where it appears to be required. No mechanism connects the cytosolic PP4c-PP4R4 complex, whose substrates are unknown, to zona escape. PubMed searches for PPP4R4 or KIAA1622 with embryo or blastocyst found no other study. Neither horse product carries the term in GOA, and the horse gene review does not address it. The prediction is neither supported nor refuted; a validated knockout or rescue phenotype with a traced mechanism would be needed.
Supporting Evidence:
  • PMID:27869233: "In this way, 35 genes (Supplementary Table S2) were identified as indispensable for normal hatching and outgrowth."
  • PMID:27869233: "Akap3, Ankrd7, Fbll1, Ppp4r4, St8sia6 and Ube2a are not expressed in later stages but are required for successful OG"
  • PMID:18715871: "we identify a novel, stable cytosolic PP4c interacting partner, KIAA1622, which we have renamed PP4R4."
GO:0019888 protein phosphatase regulator activity GO_MF
CNN β€” Correct but not novel Review score: 2/2
Prediction method: ProtNLM2 Β· Version: UniProt API snapshot 2026-09-08
Review rationale: Human PP4R4 is an experimentally isolated stable cytosolic partner of PP4c, and the PP4c-PP4R4 complex has lower phosphatase activity than free PP4c, which establishes a noncatalytic regulatory role. Horse PPP4R4 is in the same PANTHER subfamily (PTHR21467:SF0) and PAINT node, and A0A9L0S961 keeps the HEAT repeats and the PP4c-binding residues, so the transfer is justified. The term is the core molecular function in the horse gene review. It is not novel for the gene: the canonical horse product A0A9L0S938, which is identical to A0A9L0S961 except at the extreme C-terminus, already carries GO:0019888 by IBA.
Supporting Evidence:
GO:0008287 protein serine/threonine phosphatase complex GO_CC
CNN β€” Correct but not novel Review score: 2/2
Prediction method: ProtNLM2 Β· Version: UniProt API snapshot 2026-09-08
Review rationale: Human PP4R4 binds PP4c specifically, and not PP2Ac or PP6c, in a stable cytosolic complex, so PPP4R4 is a subunit of a serine/threonine phosphatase complex; the transfer to horse rests on shared subfamily PTHR21467:SF0 and conserved PP4c-binding residues. The exact term is already carried by IBA on the canonical horse product A0A9L0S938 of the same gene, so the prediction is not novel. The horse gene review refines this complex to the more specific child GO:0030289 protein phosphatase 4 complex, but that refinement is a proposal rather than an existing annotation, so the prediction matches the existing term rather than being less precise than one.
Supporting Evidence:

Deep Research

Cyberian

(PPP4R4-deep-research-cyberian.md)

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πŸ“š Additional Documentation

Notes

(PPP4R4-notes.md)

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Bioinformatics Results

(RESULTS.md)

PPP4R4: human–horse sequence comparison

The downloaded human Q6NUP7 sequence (873 residues) and selected horse A0A9L0S961 sequence (865 residues) share 95.0% identity among 865 paired residues. Paired coverage is 99.1% of human and 100.0% of horse.

Full alignment, hashes, parameters and reproducible script: PPP4R4.json and compare.py.

Pairwise similarity supports homology but does not establish reciprocal orthology, intact gene models, conservation of targeting signals or every biological claim. Current UniProt sequences are compared; identity with the original ProtNLM input sequence is unverified.

πŸ“„ View Raw YAML

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