NIPI-3 is a conserved Tribbles family pseudokinase that serves as an adapter protein regulating innate immune signaling and larval development in C. elegans. Despite containing a protein kinase domain, NIPI-3 lacks key catalytic residues and is predicted to be catalytically inactive. It functions primarily in epidermal immunity against fungal pathogens (D. coniospora) by acting upstream of the PMK-1/p38 MAPK cascade to induce antimicrobial peptide genes (nlp-29, nlp-31). In the intestine, NIPI-3 mediates defense against Gram-negative (P. aeruginosa) and Gram-positive bacteria by negatively regulating the C/EBP transcription factor CEBP-1, which in turn controls VHP-1 phosphatase levels and PMK-1 phosphorylation status. NIPI-3 directly interacts with CEBP-1 and represses its transcription. Loss of nipi-3 causes larval arrest at L2-L3, demonstrating its essential role in development beyond immunity. The protein localizes to nuclei in epidermis, intestine, and neurons.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005634 nucleus | IBA GO_REF:0000033 | ACCEPT | Summary: NIPI-3 localizes predominantly to nuclei in epidermis, intestine, and neurons, as demonstrated by GFP knock-in studies (PMID:27927209). This nuclear localization is consistent with its role in transcriptional regulation of cebp-1. Reason: This annotation is well-supported by experimental evidence. PMID:27927209 demonstrates nuclear localization of NIPI-3 using a GFP knock-in strain. This localization is also consistent with the functional role of NIPI-3 in transcriptional repression of cebp-1. The IBA annotation based on phylogenetic inference is supported by direct experimental evidence in C. elegans. Supporting Evidence: PMID:27927209 GFP::NIPI-3 expression was observed predominantly in the nuclei (Fig. 1i), with an overall intensity peaking at the L2-L3 stages. This nuclear localization suggests a role for NIPI-3 in regulation of gene expression. file:worm/nipi-3/nipi-3-deep-research-falcon.md model: Edison Scientific Literature |
| GO:0031434 mitogen-activated protein kinase kinase binding | IBA GO_REF:0000033 | ACCEPT | Summary: NIPI-3 functions upstream of the p38 MAPK cascade in epidermal immunity. While genetic interactions between NIPI-3 and SEK-1 (MAPKK) are well-documented (PMID:18394898, PMID:27927209), direct physical binding to SEK-1 has not been demonstrated in C. elegans. The IBA annotation is based on mammalian Tribbles proteins which do interact with MAPK pathway components. Reason: The IBA annotation is phylogenetically reasonable. Mammalian Tribbles proteins interact with MAPK pathway components. In C. elegans, NIPI-3 genetically acts upstream of the SEK-1/PMK-1 cascade, though the molecular mechanism may differ from direct binding. PMID:27927209 shows that loss of sek-1 suppresses nipi-3 null lethality, indicating functional interaction. The annotation appropriately captures the conserved relationship between Tribbles and MAPK signaling. Supporting Evidence: PMID:27927209 Among the other suppressors of nipi-3(0), we found missense alterations in TIR-1/the sterile alpha (SAM) and Toll-interleukin receptor (TIR) motif-containing protein SARM), NSY-1/MAPKKK, SEK-1/MAPKK and MAK-2/MAPK-activated protein kinase (MAPKAPK) PMID:18394898 We identify a kinase, NIPI-3, related to human Tribbles homolog 1, that is likely to act upstream of the MAPKK SEK-1. |
| GO:0032436 positive regulation of proteasomal ubiquitin-dependent protein catabolic process | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: Mammalian Tribbles proteins (TRIB1, TRIB2) promote C/EBP protein degradation via the ubiquitin-proteasome system. In C. elegans, NIPI-3 regulates CEBP-1 at the transcriptional level rather than through proteasomal degradation (PMID:27927209). Reason: The IBA annotation is phylogenetically inferred from mammalian Tribbles which do promote C/EBP degradation. However, in C. elegans, NIPI-3 appears to regulate CEBP-1 primarily at the transcriptional level. PMID:27927209 states that a translational reporter showed no detectable differences in GFP expression in a nipi-3(0) background, indicating post-translational regulation is not the primary mechanism. The annotation may still apply to some extent given the conserved Tribbles-C/EBP interaction but is not a core function in C. elegans. Supporting Evidence: PMID:27927209 Consistent with the observed transcriptional regulation, we found that a translational CEBP-1::GFP reporter driven by a heterologous epidermal promoter showed no detectable differences in GFP expression in a nipi-3(0) background |
| GO:0000166 nucleotide binding | IEA GO_REF:0000043 | ACCEPT | Summary: NIPI-3 contains a protein kinase domain with predicted ATP binding sites. However, the kinase domain lacks key catalytic residues and is predicted to be catalytically inactive (pseudokinase). Reason: The UniProt annotation indicates ATP binding sites at positions 206-214 and 235. While NIPI-3 is a pseudokinase, pseudokinases often retain nucleotide binding capability even though they lack catalytic activity. The IEA annotation based on keyword mapping is appropriate as it does not imply catalytic function. UniProt explicitly states the kinase domain is predicted to be catalytically inactive but still annotates ATP binding. Supporting Evidence: PMID:27927209 The Tribbles genes encode a family of highly conserved pseudokinases which lack key catalytic amino acids in the kinase domain |
| GO:0004672 protein kinase activity | IEA GO_REF:0000002 | REMOVE | Summary: NIPI-3 contains an InterPro-recognized protein kinase domain (IPR000719). However, multiple sources confirm that NIPI-3 is a pseudokinase lacking catalytic activity. Reason: This annotation is incorrect. NIPI-3 is a well-characterized pseudokinase that lacks key catalytic residues and is not expected to have protein kinase activity. UniProt explicitly states the protein kinase domain is predicted to be catalytically inactive (PROSITE-ProRule:PRU00159). PMID:27927209 confirms this. The IEA annotation based solely on domain presence is misleading for this pseudokinase. Supporting Evidence: PMID:27927209 The Tribbles genes encode a family of highly conserved pseudokinases which lack key catalytic amino acids in the kinase domain PMID:22470487 nipi-4 encodes a nematode-specific protein with a kinase-like domain that is predicted to be a pseudokinase |
| GO:0005524 ATP binding | IEA GO_REF:0000120 | ACCEPT | Summary: Based on InterPro kinase domain and UniProt keyword mapping. NIPI-3 contains ATP binding motifs in its kinase domain despite being a pseudokinase. Reason: Pseudokinases can retain ATP binding capability even without catalytic activity. UniProt annotates specific ATP binding sites at positions 206-214 and 235, suggesting the structure accommodates ATP. This is consistent with many characterized pseudokinases that bind ATP for regulatory purposes. The annotation is appropriate. Supporting Evidence: PMID:27927209 nipi-3 encodes a pseudokinase of the Tribbles family |
| GO:0005634 nucleus | IEA GO_REF:0000044 | ACCEPT | Summary: IEA annotation based on UniProt subcellular location mapping. This is redundant with the IBA annotation and is supported by direct experimental evidence. Reason: Nuclear localization is well-documented experimentally (PMID:27927209). This IEA annotation is consistent with the IBA annotation and experimental data. Duplicate annotations with different evidence codes are acceptable in GO. Supporting Evidence: PMID:27927209 GFP::NIPI-3 expression was observed predominantly in the nuclei |
| GO:0010468 regulation of gene expression | IMP PMID:27927200 Tribbles ortholog NIPI-3 and bZIP transcription factor CEBP-... | ACCEPT | Summary: NIPI-3 is required for proper expression of immune genes in both unstressed and infected animals. nipi-3 mutants show widespread misregulation of pathogen-responsive genes (PMID:27927200). NIPI-3 also represses transcription of cebp-1 (PMID:27927209). Reason: The annotation is well-supported. PMID:27927200 demonstrates in the abstract that NIPI-3 protects against intestinal infection by negatively regulating CEBP-1. PMID:27927209 shows that nipi-3 is necessary for the proper expression of immune genes. Supporting Evidence: PMID:27927200 NIPI-3 is a negative regulator of CEBP-1, which in turn negatively regulates protective immune mechanisms PMID:27927209 NIPI-3 negatively regulates expression of cebp-1 at the transcriptional level |
| GO:0010629 negative regulation of gene expression | IMP PMID:34407394 Tribbles pseudokinase NIPI-3 regulates intestinal immunity i... | ACCEPT | Summary: NIPI-3 negatively regulates CEBP-1 expression, which in turn affects downstream gene expression including vhp-1. Loss of nipi-3 leads to increased CEBP-1 levels and subsequent upregulation of VHP-1 (PMID:34407394). Reason: The annotation is supported by the finding that NIPI-3 negatively regulates CEBP-1 transcription. PMID:34407394 states in the abstract that NIPI-3 protects against intestinal infection by negatively regulating CEBP-1. Supporting Evidence: PMID:34407394 NIPI-3 has been reported to protect against intestinal infection by negatively regulating the CCAT enhancer binding protein (C/EBP) bZIP transcription factor CEBP-1 PMID:27927209 NIPI-3 negatively regulates expression of cebp-1 at the transcriptional level |
| GO:0050829 defense response to Gram-negative bacterium | IMP PMID:27927200 Tribbles ortholog NIPI-3 and bZIP transcription factor CEBP-... | ACCEPT | Summary: nipi-3(fr4) mutants are hypersusceptible to Pseudomonas aeruginosa. Intestinal NIPI-3 is both necessary and sufficient to mediate defense against this Gram-negative pathogen (PMID:27927200). Reason: This is a well-characterized core function of NIPI-3. PMID:27927200 abstract indicates that NIPI-3 protects against intestinal infection. Defense against P. aeruginosa is a key immune function. Supporting Evidence: PMID:27927200 the Tribbles pseudokinase ortholog nipi-3, is essential for host survival following exposure to P. aeruginosa or ToxA |
| GO:0050830 defense response to Gram-positive bacterium | IMP PMID:34407394 Tribbles pseudokinase NIPI-3 regulates intestinal immunity i... | ACCEPT | Summary: nipi-3 knockdown in intestinal cells causes hypersusceptibility to the Gram-positive bacterium Enterococcus faecalis (PMID:34407394, PMID:27927200). Reason: PMID:34407394 demonstrates that NIPI-3 protects against intestinal infection. This is consistent with NIPI-3's role in intestinal immunity and represents a core immune function. Supporting Evidence: PMID:34407394 NIPI-3 has been reported to protect against intestinal infection by negatively regulating the CCAT enhancer binding protein (C/EBP) bZIP transcription factor CEBP-1 |
| GO:0010629 negative regulation of gene expression | IMP PMID:27927209 Coordinated inhibition of C/EBP by Tribbles in multiple tiss... | ACCEPT | Summary: NIPI-3 represses transcription of cebp-1 in multiple tissues. Loss of nipi-3 results in increased cebp-1 mRNA levels (PMID:27927209). Reason: This is a core function. PMID:27927209 shows that the expression of the cebp-1 transcriptional reporter was highly and significantly increased in nipi-3 mutants and Quantitative RT-PCR analysis also showed significantly increased expression of cebp-1 mRNAs. This negative transcriptional regulation is central to NIPI-3 function. Supporting Evidence: PMID:27927209 Strikingly, the expression of the cebp-1 transcriptional reporter was highly and significantly increased in both nipi-3(fr4) and nipi-3(0) mutants, compared with wild-type animals PMID:27927209 Quantitative RT-PCR analysis also showed significantly increased expression of cebp-1 mRNAs in nipi-3 mutant animals |
| GO:0140297 DNA-binding transcription factor binding | IPI PMID:27927209 Coordinated inhibition of C/EBP by Tribbles in multiple tiss... | ACCEPT | Summary: NIPI-3 physically interacts with CEBP-1, a bZIP transcription factor. The N-terminal region of CEBP-1 is sufficient for binding NIPI-3 (PMID:27927209). Reason: Yeast two-hybrid studies demonstrated direct physical interaction between NIPI-3 and CEBP-1. PMID:27927209 identified CEBP-1 as a prominent candidate interacting partner and showed that the N-terminal region of CEBP-1 was sufficient for binding NIPI-3. This is a core molecular function. Supporting Evidence: PMID:27927209 One prominent candidate interacting partner was CEBP-1, a member of the C/EBP family of transcription factors PMID:27927209 In further analyses using the yeast two-hybrid assay, we found that the N-terminal region (amino acids 1-115) of CEBP-1 was sufficient for binding NIPI-3 |
| GO:0010628 positive regulation of gene expression | IMP PMID:18394898 Distinct innate immune responses to infection and wounding i... | ACCEPT | Summary: NIPI-3 is required for induction of antimicrobial peptide genes nlp-29 and nlp-31 in the epidermis after fungal infection. Loss of nipi-3 results in failure to upregulate these genes (PMID:18394898). Reason: PMID:18394898 shows in the abstract that mutants were isolated that failed to induce nlp-29 expression after D. coniospora infection, including NIPI-3. This positive regulation of AMP gene expression is a core function of NIPI-3 in epidermal immunity. Supporting Evidence: PMID:18394898 Through a forward genetic screen, we isolated mutants that failed to induce nlp-29 expression after D. coniospora infection. We identify a kinase, NIPI-3, related to human Tribbles homolog 1, that is likely to act upstream of the MAPKK SEK-1. |
| GO:0050832 defense response to fungus | IMP PMID:22470487 The pseudokinase NIPI-4 is a novel regulator of antimicrobia... | ACCEPT | Summary: nipi-3 mutants fail to induce antimicrobial peptide genes after D. coniospora infection. PMID:22470487 describes nipi-3 in the context of fungal defense pathways. Reason: This is a core function of NIPI-3. Multiple studies (PMID:18394898, PMID:22470487) demonstrate that NIPI-3 is essential for the epidermal innate immune response to fungal infection. The name "nipi" stands for "no induction of peptide after Drechmeria infection." Supporting Evidence: PMID:22470487 infection specifically involves the Tribbles-like kinase NIPI-3 |
| GO:0061760 antifungal innate immune response | IMP PMID:18394898 Distinct innate immune responses to infection and wounding i... | ACCEPT | Summary: NIPI-3 is required for the epidermal innate immune response to fungal infection by D. coniospora. It acts upstream of the p38 MAPK cascade to induce antimicrobial peptides (PMID:18394898). Reason: This is the defining function for which nipi-3 was named. PMID:18394898 shows in the abstract that NIPI-3 is required specifically for nlp-29 induction after fungal infection and not after wounding. The annotation accurately captures this specific immune function. Supporting Evidence: PMID:18394898 We find NIPI-3 is required only for nlp-29 induction after infection and not after wounding. PMID:18394898 We identify a kinase, NIPI-3, related to human Tribbles homolog 1, that is likely to act upstream of the MAPKK SEK-1. |
| GO:0050832 defense response to fungus | IMP PMID:18394898 Distinct innate immune responses to infection and wounding i... | ACCEPT | Summary: Same as above entry from PMID:22470487. NIPI-3 is essential for antifungal defense in the epidermis. Reason: Duplicate annotation with different reference. Both PMIDs support this core function. PMID:18394898 originally characterized nipi-3's role in fungal defense. Supporting Evidence: PMID:18394898 We identify a kinase, NIPI-3, related to human Tribbles homolog 1, that is likely to act upstream of the MAPKK SEK-1. We find NIPI-3 is required only for nlp-29 induction after infection |
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Download this section (compressed HTML)Q: Does NIPI-3 retain any nucleotide binding or have structural roles independent of catalysis?
Q: What is the mechanism by which nuclear NIPI-3 represses cebp-1 transcription?
Q: Are there additional transcription factor targets of NIPI-3 beyond CEBP-1?
Experiment: Crystallographic or cryo-EM studies of NIPI-3 to determine if it binds ATP
Experiment: ChIP-seq for NIPI-3 to identify direct transcriptional targets
Experiment: Mass spectrometry to identify additional NIPI-3 interaction partners beyond CEBP-1
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