PI15 is a secreted, glycosylated CAP/CRISP-family serine-protease inhibitor synthesized as a 258-amino-acid precursor and processed to a mature extracellular protein. It was originally purified from human glioblastoma conditioned medium as p25TI and displays weak trypsin-inhibitor activity. During Chlamydia trachomatis infection, PI15 is recruited into the inclusion lumen, binds the CPAF protease zymogen, and concentration-dependently promotes or inhibits CPAF activation; altering PI15 abundance perturbs production of infectious chlamydial progeny. Circulating PI15 is elevated in a subset of cholangiocarcinoma patients and falls after tumor resection, supporting biomarker utility without establishing a causal tumor function. PI15's normal physiological protease targets and organismal role remain unknown.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005576 extracellular region | IBA GO_REF:0000033 | ACCEPT | Summary: PAINT infers extracellular activity across the CAP superfamily, including PI15 itself among the experimental sources for the ancestral localization. Reason: Human PI15 was purified from serum-free conditioned medium, has a cleaved secretory precursor, and is detectable in patient plasma. These direct data make extracellular activity a well-supported core context. GO has since obsoleted GO:0005615 extracellular space, the replacement previously proposed here, and merged it into GO:0005576 (replaced_by), so this term is now the correct location term. Propagation Review Root cause: NO FAILURE CORE Sources checked: UniProtKB:O43692 · human PI15 SUPPORTS TRANSFER Direct human purification and N-terminal sequencing support secretion and anchor the ancestral extracellular localization in the target itself. PANTHER:PTN000036124 · PTHR10334 ancestral node SUPPORTS TRANSFER Broad CAP-family extracellular transfer agrees with direct target evidence. Supporting Evidence: PMID:8882727 secreted by human glioblastoma cell line T98G were analyzed by gelatin reverse PMID:30638862 Plasma PI15 levels in CCA patients were obviously reduced (p = .0014) after |
| GO:0060090 molecular adaptor activity | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: PAINT transfers a molecular-adaptor term from mouse Glipr1l1. Separately, direct PI15 experiments show CPAF-zymogen binding and support a model in which oligomeric PI15 juxtaposes CPAF zymogens during chlamydial infection. Reason: The broad term is biologically defensible for the specialized infection context because PI15 binds CPAF zymogen and promotes concentration-dependent autoprocessing, plausibly by bringing zymogens together. However, simultaneous bridging is a proposed model, not a resolved constitutive activity. The PAINT path itself is unsafe because its sole seed is the divergent sperm paralog mouse Glipr1l1 rather than direct PI15 evidence. Propagation Review Root cause: PROPAGATION BAD Failure modes: WRONG ORTHOLOG OR PARALOG CONTEXT OR TISSUE MISMATCH ROLE CONFLATION Sources checked: MGI:MGI:1916536 · mouse Glipr1l1 SUPPORTS SOURCE BUT NOT TARGET Glipr1l1's IZUMO1-containing sperm complex does not establish the PI15 CPAF-zymogen mechanism; direct PMID:29900129 evidence independently rescues the broad term for an infection-specific context. PANTHER:PTN000036124 · broad PTHR10334 ancestral node UNRESOLVED The node spans divergent CAP subfamilies and cannot establish a shared adaptor mechanism from a single sperm-paralog seed. Supporting Evidence: PMID:29900129 We show that PI15 binds to the CPAF zymogen PMID:29900129 the oligomerizing properties of PI15 may position CPAF zymogens in close proximity |
| GO:0005576 extracellular region | IEA GO_REF:0000120 | ACCEPT | Summary: Combined automated mapping assigns extracellular localization from PI15's CAP/secretory features. Reason: A signal peptide, direct purification from conditioned medium, and plasma detection all confirm that the predicted location is correct. GO has since obsoleted GO:0005615 extracellular space, the replacement previously proposed here, and merged it into GO:0005576 (replaced_by), so this term is now the correct location term. Supporting Evidence: file:human/PI15/PI15-uniprot.txt FT SIGNAL 1..19 PMID:8882727 secreted by human glioblastoma cell line T98G were analyzed by gelatin reverse |
| GO:0005576 extracellular region | EXP PMID:8882727 Purification and identification of a novel and four known se... | ACCEPT | Summary: PI15/p25TI was directly purified and N-terminally sequenced from serum-free T98G glioblastoma conditioned medium. Reason: The experimental evidence for a soluble protein recovered from conditioned medium is strong. GO has since obsoleted GO:0005615 extracellular space, the replacement previously proposed here, and merged it into GO:0005576 (replaced_by), so this term is now the correct location term. Supporting Evidence: PMID:8882727 The serum-free conditioned medium of T98G cells showed PMID:30638862 High levels of plasma PI15 were evident in CCA patients |
| GO:0070062 extracellular exosome | HDA PMID:23533145 In-depth proteomic analyses of exosomes isolated from expres... | KEEP AS NON CORE | Summary: PI15 was detected in a pooled EPS-urine exosome proteome from low-grade prostate-cancer and non-cancer samples. Reason: The curator's high-throughput detection should be retained, but the source study notes that proteins in exosome-enriched clinical-fluid preparations may also be soluble or contaminating. Exosome association is contextual and is not required for PI15's defining secreted inhibitor activity. Supporting Evidence: PMID:23533145 In pooled EPS-urine exosome samples, ~900 proteins were detected. file:human/PI15/PI15-deep-research-manual.md Exosome localization should therefore remain non-core pending vesicle-protection or immunocapture evidence. |
| GO:0004867 serine-type endopeptidase inhibitor activity | IDA PMID:8882727 Purification and identification of a novel and four known se... | NEW | Summary: Proposed new direct annotation. Purified human p25TI/PI15 weakly inhibited trypsin in reverse zymography, and recombinant PI15 also inhibited CPAF at higher concentrations. Reason: Two independent biochemical settings support direct inhibition of serine-type endopeptidases. The specific MF term captures PI15's demonstrated activity better than a generic peptidase-inhibitor or binding term. Supporting Evidence: PMID:8882727 p25TI showed weak inhibitory activity against trypsin in reverse PMID:29900129 increasing concentrations of rPI15 inhibited CPAF protease activity |
| GO:0010951 negative regulation of endopeptidase activity | IDA PMID:29900129 Peptidase Inhibitor 15 (PI15) Regulates Chlamydial CPAF Acti... | NEW | Summary: Proposed new direct annotation. Higher concentrations of recombinant PI15 suppress CPAF protease activity and zymogen maturation. Reason: Direct concentration-response assays establish a negative regulatory effect on CPAF endopeptidase activity, while the broader physiological relevance remains infection-context-specific. Supporting Evidence: PMID:29900129 increasing concentrations of rPI15 inhibited CPAF protease activity |
| GO:0051851 host-mediated perturbation of symbiont process | IMP PMID:29900129 Peptidase Inhibitor 15 (PI15) Regulates Chlamydial CPAF Acti... | NEW | Summary: Proposed new direct annotation. PI15 depletion and overexpression perturb CPAF maturation and production of infectious Chlamydia progeny in human cells. Reason: Genetic perturbation and CPAF-mutant controls establish that host PI15 modulates a symbiont process. The neutral perturbation term avoids asserting a uniformly positive or negative effect because both too little and too much PI15 impair chlamydial development. Supporting Evidence: PMID:29900129 We observed decreased CPAF activation if PI15 levels were reduced PMID:29900129 both absence of PI15 as well as overexpression of PI15 have an impact on the activation of CPAF |
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Download this section (compressed HTML)Q: Which endogenous extracellular serine proteases bind and are inhibited by mature human PI15 at physiological concentrations?
Q: Does oligomeric PI15 directly bridge two CPAF zymogens, and what structural transition switches PI15 from activation to inhibition as concentration rises?
Q: What normal tissue or developmental process requires PI15, and is the chlamydial CPAF mechanism an exploitation of that physiological activity?
Q: Is elevated plasma PI15 in cholangiocarcinoma merely a tumor-derived biomarker, or does PI15 alter the tumor extracellular protease environment?
Experiment: Produce correctly processed and glycosylated human PI15, determine binding and inhibition constants across a focused extracellular-serine-protease panel, and map activity with CAP-domain mutants. Validate candidates by secretome degradomics in PI15-null and rescue cells.
Hypothesis: Mature PI15 selectively inhibits a restricted set of extracellular human serine proteases through its CAP domain.
Type: Quantitative protease profiling, structural mutagenesis, and degradomics
Experiment: Measure PI15 oligomer stoichiometry and CPAF-zymogen binding by SEC-MALS, native mass spectrometry, cross-linking, and cryo-EM or crystallography across the activating and inhibitory concentration ranges. Test oligomerization-defective mutants in cell-free processing assays and PI15-null infected cells.
Hypothesis: PI15 oligomerization produces the concentration-dependent switch between CPAF activation and inhibition.
Type: Structural biochemistry and separation-of-function rescue
Experiment: Create PI15 knockout and endogenous-rescue models in primary prostate, lymphoid, smooth-muscle, and cholangiocyte contexts selected by verified expression. Quantify secretome proteolysis, matrix remodeling, differentiation, and stress responses without and with Chlamydia infection.
Hypothesis: Endogenous PI15 is required for a normal extracellular protease-regulatory process independent of infection.
Type: Context-resolved CRISPR genetics and extracellular protease phenotyping
Experiment: Fractionate conditioned medium and EPS urine by orthogonal vesicle methods, then combine protease protection, detergent release, immunocapture, and immunoelectron microscopy with soluble-protein controls.
Hypothesis: PI15 is packaged within a specific extracellular-vesicle population rather than simply co-purifying as a soluble secreted protein.
Type: Extracellular-vesicle localization and topology analysis
What is not known — curated, literature-grounded statements of the open unknowns (the inverse of core functions).
Gap: PI15's physiological human protease substrates, target specificity, inhibitory constants, and molecular inhibition mechanism are unknown.
OPEN BIOLOGY MF_DARK
What is known: Direct assays show weak trypsin inhibition and high-concentration CPAF inhibition, but neither substrate represents an established normal human extracellular target.
Significance: Identifying endogenous targets is necessary to explain why PI15 evolved as a secreted CAP protein and to distinguish broad from highly selective inhibition.
What would resolve it: Quantitative inhibition and binding screens against extracellular serine proteases, followed by endogenous degradomics and target-specific rescue.
Provenance (the field's own admissions):
Gap: PI15's normal physiological biological process and organismal phenotype are unknown.
OPEN BIOLOGY BP_DARK
What is known: Human-cell experiments establish a CPAF-dependent role during chlamydial infection, while normal-tissue expression and chicken developmental expression do not establish a causal conserved process.
Significance: The infection phenotype may exploit a pre-existing host function rather than reveal PI15's evolved physiological role.
What would resolve it: Use gene-specific loss-of-function and rescue in normal PI15-expressing human tissues and conditional mouse models, with extracellular protease activity and tissue phenotypes measured in parallel.
Provenance (the field's own admissions):
Gap: It is unclear whether PI15 is a true exosome/prostasome cargo or a soluble protein that co-purifies with extracellular vesicles.
OPEN BIOLOGYCURATION CC_DARK
What is known: PI15 appears in an EPS-urine exosome proteome, but the source study highlights soluble-protein and clinical-fluid contamination as alternative explanations.
Significance: Resolving vesicle association would distinguish a trafficking mechanism from incidental recovery of a secreted protein.
What would resolve it: Apply density gradients, size-exclusion chromatography, protease protection, detergent sensitivity, immunogold microscopy, and PI15 immunocapture to purified vesicles from prostate and other PI15-positive sources.
Provenance (the field's own admissions):
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