GO:0052725 inositol-1,3,4-trisphosphate 6-kinase activity: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0052725 describes the ATP-dependent phosphorylation of 1D-myo-inositol 1,3,4-trisphosphate to 1D-myo-inositol 1,3,4,6-tetrakisphosphate, a reaction catalyzed by inositol 1,3,4-trisphosphate 5/6-kinase (ITPK1).
• The enzyme is bifunctional, producing both 6-kinase and 5-kinase products from Ins(1,3,4)P3, and it also acts as a protein kinase on transcription factors such as c-Jun and ATF-2.
• ITPK1 activity is regulated by reversible lysine acetylation, linking inositol phosphate metabolism to cellular acetyl-CoA status.
• The enzyme associates with the COP9 signalosome via CSN1, connecting inositol polyphosphate synthesis to protein stability and signal transduction.
• Loss of ITPK1 function sensitizes cells to TNF-induced apoptosis, indicating a role in cell survival signaling.
• The crystal structure of ITPK1 revealed specificity determinants for inositol polyphosphate synthesis, enabling structure-guided studies.
Description
Inositol-1,3,4-trisphosphate 6-kinase activity (GO:0052725) is a molecular function that catalyzes the phosphorylation of 1D-myo-inositol 1,3,4-trisphosphate (Ins(1,3,4)P3) at the 6-position to yield 1D-myo-inositol 1,3,4,6-tetrakisphosphate (Ins(1,3,4,6)P4). This reaction is part of the inositol polyphosphate pathway, which generates second messengers and cofactors involved in diverse cellular processes, including calcium signaling, vesicle trafficking, and gene expression. The enzyme responsible, inositol 1,3,4-trisphosphate 5/6-kinase (ITPK1), was first purified from rat liver and subsequently cloned, revealing a bifunctional enzyme that can also phosphorylate at the 5-position. Beyond its inositol kinase activity, ITPK1 functions as a protein kinase that phosphorylates the transcription factors c-Jun and ATF-2, thereby linking inositol phosphate metabolism to transcriptional regulation. ITPK1 also associates with the COP9 signalosome through CSN1, suggesting a role in modulating signal transduction complexes. Regulation of ITPK1 by reversible lysine acetylation further connects its activity to cellular metabolic status. In plants, the Arabidopsis ortholog AtItpk-1 is involved in photomorphogenesis under red light, possibly via interaction with the COP9 signalosome, highlighting evolutionary conservation. Given its central role in inositol polyphosphate synthesis and its emerging functions in apoptosis, transcription, and signalosome biology, GO:0052725 is a critical node for researchers studying cell signaling, cancer, and metabolic regulation.
inositol-1,3,4-trisphosphate 6-kinase activity At A Glance
| GO ID | GO:0052725 |
|---|---|
| GO term | inositol-1,3,4-trisphosphate 6-kinase activity |
| Ontology | molecular_function |
| Synonym | 1D-myo-inositol-trisphosphate 6-kinase activity; inositol 1,3,4-trisphosphate 6-kinase activity; inositol-trisphosphate 6-kinase activity; ins(1,3,4)P(3) 6-kinase activity; Ins(1,3,4)P3 6-kinase activity; IP3 6-kinase activity |
| Major function | Phosphorylation of 1D-myo-inositol 1,3,4-trisphosphate at the 6-position to produce 1D-myo-inositol 1,3,4,6-tetrakisphosphate |
| Enzyme | Inositol 1,3,4-trisphosphate 5/6-kinase (ITPK1) |
| Substrate | 1D-myo-inositol 1,3,4-trisphosphate and ATP |
| Product | 1D-myo-inositol 1,3,4,6-tetrakisphosphate, ADP, and H+ |
| Cofactor | Mg2+ (implied by kinase mechanism) |
| Regulation | Reversible lysine acetylation |
What Is GO:0052725?
GO:0052725, inositol-1,3,4-trisphosphate 6-kinase activity, is defined as the catalysis of the reaction: 1D-myo-inositol 1,3,4-trisphosphate + ATP = 1D-myo-inositol 1,3,4,6-tetrakisphosphate + ADP + H+. In other words, it is the enzyme activity that transfers a phosphate group from ATP to the 6-hydroxyl position of Ins(1,3,4)P3, producing Ins(1,3,4,6)P4 and ADP. This activity is one of the two kinase activities of the bifunctional enzyme ITPK1, which can also phosphorylate at the 5-position.
Why Is inositol-1,3,4-trisphosphate 6-kinase activity Important in Cell Biology?
GO:0052725 is important because it represents a key step in the generation of higher inositol polyphosphates, which are essential for numerous cellular processes including signal transduction, apoptosis, and transcriptional regulation. The enzyme ITPK1, which carries this activity, is bifunctional and can also act as a protein kinase, thereby directly linking inositol phosphate metabolism to gene expression. Its association with the COP9 signalosome suggests a role in regulating protein stability and cellular responses to stress. Moreover, ITPK1 activity is modulated by acetylation, connecting it to metabolic state. Dysregulation of inositol polyphosphate signaling has been implicated in cancer and other diseases, making this activity a potential therapeutic target.
• Generates Ins(1,3,4,6)P4, a precursor for higher inositol polyphosphates involved in cellular signaling.
• ITPK1, the enzyme with this activity, inhibits TNF-induced apoptosis, linking it to cell survival.
• Acts as a protein kinase on c-Jun and ATF-2, connecting inositol metabolism to transcription.
• Regulated by lysine acetylation, integrating metabolic signals.
• Associates with the COP9 signalosome, impacting protein stability and signal transduction.
• Conserved in plants, where it affects photomorphogenesis.
• Crystal structure provides a basis for understanding specificity and inhibitor design.
• Potential role in cancer and other diseases through apoptosis regulation.
Molecular Mechanism of inositol-1,3,4-trisphosphate 6-kinase activity
Substrate recognition and binding
In simple terms: The enzyme grabs the inositol trisphosphate molecule and ATP.
ITPK1 specifically binds 1D-myo-inositol 1,3,4-trisphosphate (Ins(1,3,4)P3) and ATP. The crystal structure of ITPK1 revealed key residues that determine substrate specificity, including those that interact with the inositol ring and phosphate groups. The enzyme can also phosphorylate at the 5-position, but the 6-kinase activity is defined by the transfer to the 6-hydroxyl.
Catalytic mechanism
In simple terms: The enzyme transfers a phosphate from ATP to the inositol ring.
The reaction catalyzed is: 1D-myo-inositol 1,3,4-trisphosphate + ATP = 1D-myo-inositol 1,3,4,6-tetrakisphosphate + ADP + H+. This is a typical kinase reaction requiring a divalent metal ion, likely Mg2+, for ATP coordination. The enzyme is bifunctional, producing both 5- and 6-kinase products, but the 6-kinase activity specifically adds a phosphate to the 6-position.
Protein kinase activity
In simple terms: The same enzyme can also phosphorylate proteins.
Beyond its inositol kinase activity, ITPK1 acts as a protein kinase that phosphorylates the transcription factors c-Jun and ATF-2. This dual function suggests a direct role in transcriptional regulation.
Regulation by acetylation
In simple terms: The enzyme's activity can be turned on or off by adding acetyl groups.
ITPK1 is regulated by reversible lysine acetylation. This modification can affect its enzymatic activity and possibly its interactions, linking inositol phosphate metabolism to cellular acetyl-CoA levels.
Interaction with COP9 signalosome
In simple terms: The enzyme binds to a large protein complex that controls protein degradation.
ITPK1 associates with the COP9 signalosome by binding to the CSN1 subunit. This interaction may localize the enzyme to specific cellular compartments or modulate its activity in response to signals.
Key Genes Involved in GO:0052725 inositol-1,3,4-trisphosphate 6-kinase activity
The following genes and proteins are directly involved in or regulate inositol-1,3,4-trisphosphate 6-kinase activity (GO:0052725) and its associated pathways.
| Gene | Major Role | Research Relevance |
|---|---|---|
| ITPK1 | Bifunctional inositol 1,3,4-trisphosphate 5/6-kinase and protein kinase; catalyzes the 6-kinase reaction | Central enzyme for GO:0052725; target for knockout, point mutation, and overexpression studies |
| CSN1 | Subunit of the COP9 signalosome; binds ITPK1 | Links ITPK1 to signalosome functions; potential target for interaction studies |
| JUN | Transcription factor c-Jun; phosphorylated by ITPK1 | Connects ITPK1 to AP-1 signaling; relevant in cancer and stress responses |
| ATF2 | Activating transcription factor 2; phosphorylated by ITPK1 | Links ITPK1 to transcriptional regulation; potential disease relevance |
| TNF | Tumor necrosis factor; its induced apoptosis is inhibited by ITPK1 | Used to study ITPK1's role in cell survival |
| IPK1 | Inositol pentakisphosphate 2-kinase; downstream enzyme in inositol polyphosphate pathway | May act downstream of ITPK1; relevant for pathway analysis |
| IPK2 | Inositol polyphosphate multikinase; produces precursors for ITPK1 | Upstream enzyme; potential target for combinatorial studies |
| PLC | Phospholipase C; generates Ins(1,4,5)P3, which can be converted to Ins(1,3,4)P3 | Upstream of ITPK1 substrate production |
| ITPKA | Inositol-trisphosphate 3-kinase A; phosphorylates Ins(1,4,5)P3 to Ins(1,3,4,5)P4 | Related enzyme in inositol phosphate metabolism |
| ITPKB | Inositol-trisphosphate 3-kinase B; similar to ITPKA | Related enzyme; potential redundancy |
| ITPKC | Inositol-trisphosphate 3-kinase C; involved in calcium signaling | Related enzyme; may affect substrate availability |
| MINPP1 | Multiple inositol polyphosphate phosphatase 1; degrades inositol polyphosphates | Counteracts ITPK1 by dephosphorylation |
| PTEN | Phosphatase and tensin homolog; dephosphorylates PIP3, affecting inositol phosphate pools | Indirectly influences substrate levels |
| INPP4A | Inositol polyphosphate-4-phosphatase type I; degrades Ins(1,3,4)P3 | Regulates substrate availability for ITPK1 |
| INPP4B | Inositol polyphosphate-4-phosphatase type II; similar to INPP4A | Potential regulator of ITPK1 substrate |
| ATItpk-1 | Arabidopsis inositol 1,3,4-trisphosphate 5/6 kinase; involved in photomorphogenesis | Plant ortholog for comparative studies |
| CSN5 | COP9 signalosome subunit 5; may interact with ITPK1 complex | Potential component of ITPK1-associated complex |
| CSN7 | COP9 signalosome subunit 7; part of the complex | Potential interaction partner |
How Is inositol-1,3,4-trisphosphate 6-kinase activity Regulated?
ITPK1, the enzyme responsible for GO:0052725, is regulated by reversible lysine acetylation. This post-translational modification can alter its enzymatic activity and potentially its interactions with other proteins, thereby linking inositol phosphate metabolism to cellular metabolic status. Additionally, ITPK1 associates with the COP9 signalosome via CSN1, which may regulate its stability or localization. The enzyme's dual function as a protein kinase also suggests that its activity could be modulated by phosphorylation or other signaling events, though specific upstream regulators remain to be fully elucidated.
inositol-1,3,4-trisphosphate 6-kinase activity and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| ITPK1 | Cancer (apoptosis resistance) | Knockout in cancer cell lines to assess TNF-induced apoptosis |
| ITPK1 | Metabolic disorders (acetylation-linked) | Point mutation of acetylation sites to study activity changes |
| ITPK1 | Signalosome-related diseases | Knock-in of tagged ITPK1 to study interaction with CSN1 |
| ITPK1 | Transcriptional dysregulation | Overexpression to study c-Jun/ATF-2 phosphorylation |
| AtItpk-1 | Plant photomorphogenesis | Arabidopsis knockout for red light response |
Cancer and apoptosis
ITPK1, the enzyme with inositol-1,3,4-trisphosphate 6-kinase activity, inhibits tumor necrosis factor (TNF)-induced apoptosis. This suggests that loss of ITPK1 function could sensitize cancer cells to apoptosis, making it a potential target for cancer therapy. The protein kinase activity of ITPK1 on c-Jun and ATF-2 further implicates it in pathways that control cell proliferation and survival.
Metabolic and signaling disorders
The regulation of ITPK1 by lysine acetylation connects it to cellular metabolism. Dysregulation of acetylation is observed in metabolic disorders and cancer, suggesting that ITPK1 activity could be altered in these conditions. Additionally, its interaction with the COP9 signalosome, a key regulator of protein stability, implies a role in diseases characterized by defective signalosome function.
Plant development and photomorphogenesis
In Arabidopsis, the ITPK1 ortholog AtItpk-1 is involved in photomorphogenesis under red light, possibly through interaction with the COP9 signalosome. This highlights a conserved role for this enzyme activity in developmental processes, though direct human disease links are not established.
From inositol-1,3,4-trisphosphate 6-kinase activity-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does ITPK1 knockout affect TNF-induced apoptosis? | ITPK1 knockout cell lines treated with TNF |
| Which residues are critical for 6-kinase activity? | Point mutations in ITPK1 catalytic domain |
| Does acetylation regulate ITPK1 activity? | Knock-in of acetylation-mimetic or -dead mutants |
| Where does ITPK1 localize in cells? | Tagged knock-in (e.g., GFP) for imaging |
| Does ITPK1 overexpression alter transcription? | Overexpression of ITPK1 followed by RNA-seq |
| Is ITPK1 required for COP9 signalosome function? | Knockout of ITPK1 and assessment of CSN1 interaction |
How to Study the inositol-1,3,4-trisphosphate 6-kinase activity Process
| Method | What It Measures | Typical Application |
|---|---|---|
| Radioactive kinase assay | 6-kinase activity converting Ins(1,3,4)P3 to Ins(1,3,4,6)P4 | Enzyme purification and kinetic studies |
| X-ray crystallography | Three-dimensional structure of ITPK1 | Understanding substrate specificity and catalysis |
| Co-immunoprecipitation | Protein-protein interactions (e.g., ITPK1-CSN1) | Identifying signaling complexes |
| Western blot with acetylation antibodies | Acetylation status of ITPK1 | Studying regulation by acetylation |
| Site-directed mutagenesis | Effect of specific residues on activity | Mapping catalytic and regulatory sites |
| Apoptosis assays (e.g., caspase-3) | Cell death after TNF treatment | Assessing ITPK1 role in survival |
| Luciferase reporter assays | Transcriptional activity of c-Jun/ATF-2 | Linking ITPK1 to transcription |
| Plant photomorphogenesis assays | Red light response in Arabidopsis | Studying conserved function |
Enzymatic assays for 6-kinase activity
The 6-kinase activity of ITPK1 can be measured using radioactive ATP and Ins(1,3,4)P3 as substrates, followed by HPLC or thin-layer chromatography to separate products. This method was used in the original purification and characterization of the enzyme.
Structural biology
X-ray crystallography of ITPK1 provided insights into substrate specificity and catalytic mechanism. The crystal structure revealed key residues involved in binding Ins(1,3,4)P3 and ATP, enabling structure-guided mutagenesis.
Protein interaction studies
Co-immunoprecipitation and pull-down assays can identify ITPK1 interaction partners such as CSN1. These methods help elucidate how ITPK1 integrates into larger signaling complexes.
Post-translational modification analysis
Mass spectrometry and acetylation-specific antibodies can detect lysine acetylation of ITPK1. Mutagenesis of acetylation sites followed by enzymatic assays can determine the functional impact.
How CRISPR Can Be Used to Study GO:0052725 inositol-1,3,4-trisphosphate 6-kinase activity
Knockout
CRISPR knockout of ITPK1 can be used to eliminate inositol-1,3,4-trisphosphate 6-kinase activity, enabling studies on its role in TNF-induced apoptosis and inositol polyphosphate synthesis. Knockout cell lines can also reveal compensatory pathways and the importance of ITPK1 in cellular signaling.
Point Mutation
Point mutations in the catalytic domain of ITPK1 can dissect the contribution of specific residues to 6-kinase activity versus 5-kinase or protein kinase activities. Such mutants help validate structural predictions and acetylation sites.
Knock-in
Knock-in of tagged ITPK1 (e.g., GFP or FLAG) allows for localization and interaction studies, such as assessing binding to the COP9 signalosome. Knock-in of acetylation-mimetic mutants can probe the regulatory role of acetylation.
Overexpression
Overexpression of ITPK1 can enhance 6-kinase activity, leading to increased Ins(1,3,4,6)P4 levels. This can be used to study downstream effects on transcription and apoptosis.
How EDITGENE Supports inositol-1,3,4-trisphosphate 6-kinase activity Research
Researchers studying inositol-1,3,4-trisphosphate 6-kinase activity-related genes often need to determine whether a candidate gene is causally involved in a specific signaling or disease pathway. EDITGENE provides comprehensive CRISPR-based services to generate precisely engineered cell models, enabling functional validation of genes like ITPK1 and its interaction partners.
Contact EDITGENE today to design your custom CRISPR model for inositol-1,3,4-trisphosphate 6-kinase activity research.
Frequently Asked Questions About inositol-1,3,4-trisphosphate 6-kinase activity
What is inositol-1,3,4-trisphosphate 6-kinase activity?
It is the enzyme activity that catalyzes the phosphorylation of 1D-myo-inositol 1,3,4-trisphosphate to 1D-myo-inositol 1,3,4,6-tetrakisphosphate, using ATP.
What gene encodes inositol-1,3,4-trisphosphate 6-kinase activity?
The activity is primarily carried out by the ITPK1 gene, which encodes a bifunctional inositol 1,3,4-trisphosphate 5/6-kinase.
What is the function of ITPK1?
ITPK1 catalyzes the 5- and 6-kinase reactions in inositol polyphosphate synthesis and also acts as a protein kinase on c-Jun and ATF-2.
How is inositol-1,3,4-trisphosphate 6-kinase activity regulated?
It is regulated by reversible lysine acetylation and potentially by interaction with the COP9 signalosome.
What diseases are associated with ITPK1?
ITPK1 has been linked to cancer through its role in inhibiting TNF-induced apoptosis, and to metabolic disorders via acetylation.
What is the reaction catalyzed by GO:0052725?
1D-myo-inositol 1,3,4-trisphosphate + ATP = 1D-myo-inositol 1,3,4,6-tetrakisphosphate + ADP + H+.
What are the synonyms for inositol-1,3,4-trisphosphate 6-kinase activity?
Synonyms include IP3 6-kinase activity, Ins(1,3,4)P3 6-kinase activity, and inositol-trisphosphate 6-kinase activity.
How can I study inositol-1,3,4-trisphosphate 6-kinase activity?
Enzymatic assays with radioactive ATP, structural biology, and CRISPR knockout models are common approaches.
Is ITPK1 conserved in plants?
Yes, the Arabidopsis ortholog AtItpk-1 is involved in photomorphogenesis, indicating conservation.
What CRISPR models are available for ITPK1?
Knockout, point mutation, knock-in, and overexpression models can be generated to study ITPK1 function.
Conclusion
Inositol-1,3,4-trisphosphate 6-kinase activity (GO:0052725) is a key enzymatic function in inositol polyphosphate metabolism, carried out by the bifunctional enzyme ITPK1. Its role in generating Ins(1,3,4,6)P4, regulating apoptosis, and interacting with the COP9 signalosome underscores its importance in cell signaling and disease. Continued research using CRISPR models will further elucidate its mechanistic and therapeutic potential.
References
- 1. Sun Y et al.. 2003. Inositol 1,3,4-trisphosphate 5/6-kinase inhibits tumor necrosis factor-induced apoptosis.. J Biol Chem 278(44):43645-53 PMID: 12925536
- 2. Wilson MP et al.. 2001. Inositol 1,3,4-trisphosphate 5/6-kinase is a protein kinase that phosphorylates the transcription factors c-Jun and ATF-2.. J Biol Chem 276(44):40998-1004 PMID: 11533064
- 3. Zhang C et al.. 2012. Regulation of inositol 1,3,4-trisphosphate 5/6-kinase (ITPK1) by reversible lysine acetylation.. Proc Natl Acad Sci U S A 109(7):2290-5 PMID: 22308441
- 4. Wilson MP et al.. 1996. Isolation of inositol 1,3,4-trisphosphate 5/6-kinase, cDNA cloning and expression of the recombinant enzyme.. J Biol Chem 271(20):11904-10 PMID: 8662638
- 5. Sun Y et al.. 2002. Inositol 1,3,4-trisphosphate 5/6-kinase associates with the COP9 signalosome by binding to CSN1.. J Biol Chem 277(48):45759-64 PMID: 12324474
- 6. Abdullah M et al.. 1992. Purification and characterization of inositol-1,3,4-trisphosphate 5/6-kinase from rat liver using an inositol hexakisphosphate affinity column.. J Biol Chem 267(31):22340-5 PMID: 1331051
- 7. Miller GJ et al.. 2005. Specificity determinants in inositol polyphosphate synthesis: crystal structure of inositol 1,3,4-trisphosphate 5/6-kinase.. Mol Cell 18(2):201-12 PMID: 15837423
- 8. Qin ZX et al.. 2005. The Arabidopsis inositol 1,3,4-trisphosphate 5/6 kinase, AtItpk-1, is involved in plant photomorphogenesis under red light conditions, possibly via interaction with COP9 signalosome.. Plant Physiol Biochem 43(10-11):947-54 PMID: 16310364
