GA1 (ent-copalyl diphosphate synthase, CPS; At4g02780) is a plastid-targeted class II diterpene cyclase of the terpene synthase family that catalyses the protonation-initiated cyclization of (E,E,E)-geranylgeranyl diphosphate to ent-copalyl diphosphate, the first committed step of gibberellin biosynthesis. Catalysis uses a DXDD motif as general acid in an active site at the interface of the beta and gamma domains; activity is inhibited synergistically by Mg2+ and GGPP, a property suggested to limit flux into the GA pathway when plastid Mg2+ rises during de-etiolation. The enzyme is imported into chloroplasts/plastids, and its promoter is active mainly in provascular tissue of germinating seeds and in growing organs. ga1 null mutants (e.g. ga1-3) are severely GA-deficient, non-germinating, male-sterile dwarfs rescued by exogenous GA.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0000287 magnesium ion binding | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: Mg2+ binding by CPS. Class II diterpene cyclases use a DXDD general acid rather than a Mg2+-dependent diphosphate ionization, but AtCPS activity depends on and is regulated by Mg2+ (synergistic substrate inhibition with GGPP). Reason: Mg2+ binding is real and has a regulatory role (feed-forward inhibition at high plastid Mg2+), but it is a cofactor/regulatory property rather than the core catalytic function, which is ent-CPP synthase activity. Supporting Evidence: PMID:17384166 demonstrating that Mg(2+) and GGPP exert synergistic substrate inhibition effects on CPS activity PMID:20430888 Notably, this residue is conserved as a histidine in enzymes involved in gibberellin biosynthesis |
| GO:0000287 magnesium ion binding | IDA PMID:17384166 Synergistic substrate inhibition of ent-copalyl diphosphate ... | KEEP AS NON CORE | Summary: Kinetic analysis of recombinant pseudomature AtCPS showed Mg2+ and GGPP act synergistically to inhibit activity, implying direct Mg2+ binding (intrasteric binding to the DXDD motif). Reason: Experimentally supported cofactor/regulatory binding; retained as non-core because the core activity is ent-copalyl diphosphate synthase. Supporting Evidence: PMID:17384166 demonstrating that Mg(2+) and GGPP exert synergistic substrate inhibition effects on CPS activity |
| GO:0003824 catalytic activity | IEA GO_REF:0000117 | MODIFY | Summary: Generic catalytic activity from ARBA. Reason: Correct but uninformative; the specific activity (ent-copalyl diphosphate synthase, EC 5.5.1.13, RHEA:14841) is experimentally established. Proposed replacements: ent-copalyl diphosphate synthase activity Supporting Evidence: PMID:7994182 In Escherichia coli cells that express both the Arabidopsis GA1 gene and the Erwinia uredovora gene encoding GGPP synthase, CPP was accumulated. |
| GO:0009507 chloroplast | IBA GO_REF:0000033 | ACCEPT | Summary: Plastid (chloroplast) localization of CPS; the enzyme carries an N-terminal transit peptide and is imported and processed by chloroplasts. Reason: Consistent with direct import and GFP-targeting evidence. Supporting Evidence: PMID:7994182 The results showed that the GA1 protein is imported into and processed in pea chloroplasts in vitro. PMID:11722763 We show that the N-terminal pre-sequences of the Arabidopsis thaliana terpene cyclases copalyl diphosphate synthase (AtCPS1) and ent-kaurene synthase (AtKS1) direct GFP to chloroplasts in transient assays following microprojectile bombardment of tobacco leaves. |
| GO:0009507 chloroplast | IDA PMID:7994182 The Arabidopsis GA1 locus encodes the cyclase ent-kaurene sy... | ACCEPT | Summary: In vitro import into and processing in isolated pea chloroplasts. Reason: Direct experimental support for chloroplast localization; GA biosynthesis from GGPP to ent-kaurene occurs in plastids. Supporting Evidence: PMID:7994182 The results showed that the GA1 protein is imported into and processed in pea chloroplasts in vitro. |
| GO:0009507 chloroplast | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt subcellular location mapping (chloroplast). Reason: Agrees with experimental chloroplast import and GFP data. Supporting Evidence: PMID:11722763 We show that the N-terminal pre-sequences of the Arabidopsis thaliana terpene cyclases copalyl diphosphate synthase (AtCPS1) and ent-kaurene synthase (AtKS1) direct GFP to chloroplasts in transient assays following microprojectile bombardment of tobacco leaves. |
| GO:0009507 chloroplast | ISM GO_REF:0000122 | ACCEPT | Summary: AtSubP prediction of chloroplast localization. Reason: Prediction agrees with experimental import and GFP targeting data. Supporting Evidence: PMID:11722763 We show that the N-terminal pre-sequences of the Arabidopsis thaliana terpene cyclases copalyl diphosphate synthase (AtCPS1) and ent-kaurene synthase (AtKS1) direct GFP to chloroplasts in transient assays following microprojectile bombardment of tobacco leaves. |
| GO:0009686 gibberellin biosynthetic process | IBA GO_REF:0000033 | ACCEPT | Summary: GA1/CPS catalyses the first committed step of gibberellin biosynthesis (GGPP to ent-CPP); ga1 null mutants are severely GA-deficient dwarfs rescued by GA1 cDNA. Reason: CPS performs the committed step of the pathway; phylogenetically conserved role. Supporting Evidence: PMID:17384166 The committed step in GA biosynthesis is catalyzed in plastids by ent-copalyl diphosphate synthase (CPS) PMID:7994182 In Escherichia coli cells that express both the Arabidopsis GA1 gene and the Erwinia uredovora gene encoding GGPP synthase, CPP was accumulated. |
| GO:0009686 gibberellin biosynthetic process | IDA PMID:7994182 The Arabidopsis GA1 locus encodes the cyclase ent-kaurene sy... | ACCEPT | Summary: GA1/CPS catalyses the first committed step of gibberellin biosynthesis (GGPP to ent-CPP); ga1 null mutants are severely GA-deficient dwarfs rescued by GA1 cDNA. Reason: Enzyme activity of recombinant GA1 directly demonstrates participation in GA biosynthesis. Supporting Evidence: PMID:7994182 In Escherichia coli cells that express both the Arabidopsis GA1 gene and the Erwinia uredovora gene encoding GGPP synthase, CPP was accumulated. |
| GO:0009686 gibberellin biosynthetic process | IEA GO_REF:0000041 | ACCEPT | Summary: GA1/CPS catalyses the first committed step of gibberellin biosynthesis (GGPP to ent-CPP); ga1 null mutants are severely GA-deficient dwarfs rescued by GA1 cDNA. Reason: UniPathway mapping consistent with experimental evidence. Supporting Evidence: PMID:17384166 The committed step in GA biosynthesis is catalyzed in plastids by ent-copalyl diphosphate synthase (CPS) |
| GO:0009686 gibberellin biosynthetic process | IMP PMID:7994182 The Arabidopsis GA1 locus encodes the cyclase ent-kaurene sy... | ACCEPT | Summary: GA1/CPS catalyses the first committed step of gibberellin biosynthesis (GGPP to ent-CPP); ga1 null mutants are severely GA-deficient dwarfs rescued by GA1 cDNA. Reason: ga1 mutants are GA-deficient and complemented by GA1. Supporting Evidence: PMID:7994182 The ga1 mutants of Arabidopsis are gibberellin-responsive male-sterile dwarfs. PMID:7994182 This cDNA clone encodes an active protein and is able to complement the dwarf phenotype in ga1-3 mutants by Agrobacterium-mediated transformation. |
| GO:0009740 gibberellic acid mediated signaling pathway | TAS PMID:7994182 The Arabidopsis GA1 locus encodes the cyclase ent-kaurene sy... | MARK AS OVER ANNOTATED | Summary: GA1/CPS synthesizes the hormone precursor; it is not a component of the GA perception/DELLA-degradation signaling pathway. Reason: Making the ligand is upstream of signaling but is not participation in it; the gene's process role is fully captured by gibberellin biosynthetic process. The TAS row adds no information about signal transduction. Supporting Evidence: PMID:17384166 The committed step in GA biosynthesis is catalyzed in plastids by ent-copalyl diphosphate synthase (CPS) |
| GO:0009905 ent-copalyl diphosphate synthase activity | EXP PMID:17457817 Probing the role of the DXDD motif in Class II diterpene cyc... | ACCEPT | Summary: Mutational analysis of the DXDD motif of class II diterpene cyclases including AtCPS (abstract not available in cache). Reason: The ent-CPP synthase activity of AtCPS is independently established (PMID:7994182, PMID:17384166); UniProt curators read the full text. Supporting Evidence: PMID:20430888 Class II diterpene cyclases mediate the acid-initiated cycloisomerization reaction that serves as the committed step in biosynthesis of the large class of labdane-related diterpenoid natural products, which includes the important gibberellin plant hormones. |
| GO:0009905 ent-copalyl diphosphate synthase activity | EXP PMID:20430888 A single residue switch for Mg(2+)-dependent inhibition char... | ACCEPT | Summary: Mann et al. characterised class II diterpene cyclases including AtCPS, identifying the His/Arg switch residue governing Mg2+ inhibition. Reason: Direct biochemical characterization of AtCPS activity. Supporting Evidence: PMID:20430888 Class II diterpene cyclases mediate the acid-initiated cycloisomerization reaction that serves as the committed step in biosynthesis of the large class of labdane-related diterpenoid natural products, which includes the important gibberellin plant hormones. PMID:20430888 Notably, this residue is conserved as a histidine in enzymes involved in gibberellin biosynthesis |
| GO:0009905 ent-copalyl diphosphate synthase activity | IDA PMID:17384166 Synergistic substrate inhibition of ent-copalyl diphosphate ... | ACCEPT | Summary: Kinetic analysis of recombinant pseudomature AtCPS converting GGPP to ent-CPP. Reason: Direct enzyme assay. Supporting Evidence: PMID:17384166 demonstrating that Mg(2+) and GGPP exert synergistic substrate inhibition effects on CPS activity |
| GO:0009905 ent-copalyl diphosphate synthase activity | IDA PMID:7994182 The Arabidopsis GA1 locus encodes the cyclase ent-kaurene sy... | ACCEPT | Summary: GA1 expressed in E. coli with a GGPP synthase accumulates CPP. Reason: Core activity; the first committed step of GA biosynthesis (RHEA:14841). Supporting Evidence: PMID:7994182 In Escherichia coli cells that express both the Arabidopsis GA1 gene and the Erwinia uredovora gene encoding GGPP synthase, CPP was accumulated. |
| GO:0009905 ent-copalyl diphosphate synthase activity | IEA GO_REF:0000120 | ACCEPT | Summary: EC/Rhea-based mapping to ent-copalyl diphosphate synthase activity. Reason: Matches experimental evidence. Supporting Evidence: PMID:7994182 In Escherichia coli cells that express both the Arabidopsis GA1 gene and the Erwinia uredovora gene encoding GGPP synthase, CPP was accumulated. |
| GO:0009905 ent-copalyl diphosphate synthase activity | IMP PMID:7994182 The Arabidopsis GA1 locus encodes the cyclase ent-kaurene sy... | ACCEPT | Summary: ga1 loss-of-function and complementation support the ent-CPP synthase function. Reason: Genetic support consistent with biochemical activity. Supporting Evidence: PMID:7994182 This cDNA clone encodes an active protein and is able to complement the dwarf phenotype in ga1-3 mutants by Agrobacterium-mediated transformation. |
| GO:0010333 terpene synthase activity | IBA GO_REF:0000033 | MODIFY | Summary: Terpene synthase activity propagated at a PANTHER node shared by CPS and KS. In GO, terpene synthase activity is classified under lyase activity (diphosphate-releasing class I cyclases), whereas ent-CPP synthase is a class II cyclase classified as an intramolecular isomerase (EC 5.5.1.13) and is not a descendant of GO:0010333. Reason: The GO definition of terpene synthase activity (cyclization of linear prenyl diphosphates) loosely covers CPS, but its placement under lyase activity misrepresents the class II isomerase chemistry; the precise term GO:0009905 should be used. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: PANTHER:PTN001272394 SUPPORTS SOURCE BUT NOT TARGET Node spanning CPS and KS clades; terpene synthase activity (a lyase-branch term) fits the class I KS descendants, while the CPS descendants are class II isomerases better described by GO:0009905. AGI_LocusCode:AT1G79460 SUPPORTS SOURCE BUT NOT TARGET GA2/KS, a genuine class I ent-kaurene synthase (terpene synthase activity), is the Arabidopsis seed; its chemistry differs from CPS. Proposed replacements: ent-copalyl diphosphate synthase activity Supporting Evidence: PMID:20430888 Class II diterpene cyclases mediate the acid-initiated cycloisomerization reaction that serves as the committed step in biosynthesis of the large class of labdane-related diterpenoid natural products, which includes the important gibberellin plant hormones. |
| GO:0010333 terpene synthase activity | IEA GO_REF:0000002 | MODIFY | Summary: InterPro2GO terpene synthase activity from the terpene synthase N-terminal domain. Reason: As for the IBA row: CPS is a class II cyclase/isomerase; replace with the specific activity. Proposed replacements: ent-copalyl diphosphate synthase activity Supporting Evidence: PMID:20430888 Class II diterpene cyclases mediate the acid-initiated cycloisomerization reaction that serves as the committed step in biosynthesis of the large class of labdane-related diterpenoid natural products, which includes the important gibberellin plant hormones. |
| GO:0016102 diterpenoid biosynthetic process | IEA GO_REF:0000002 | MODIFY | Summary: Diterpenoid biosynthetic process (InterPro2GO). Reason: Correct but general; gibberellins are the diterpenoids made via this enzyme and GO:0009686 is the informative process. Proposed replacements: gibberellin biosynthetic process Supporting Evidence: PMID:20430888 Class II diterpene cyclases mediate the acid-initiated cycloisomerization reaction that serves as the committed step in biosynthesis of the large class of labdane-related diterpenoid natural products, which includes the important gibberellin plant hormones. |
| GO:0016114 terpenoid biosynthetic process | IEA GO_REF:0000002 | MODIFY | Summary: Terpenoid biosynthetic process (InterPro2GO). Reason: Correct but general; use gibberellin biosynthetic process. Proposed replacements: gibberellin biosynthetic process Supporting Evidence: PMID:17384166 The committed step in GA biosynthesis is catalyzed in plastids by ent-copalyl diphosphate synthase (CPS) |
| GO:0016829 lyase activity | IEA GO_REF:0000002 | REMOVE | Summary: Lyase activity from InterPro terpene synthase domain mappings. Reason: ent-copalyl diphosphate synthase (EC 5.5.1.13) is a protonation-initiated class II cyclase that retains the diphosphate group; in GO and EC it is an isomerase (intramolecular lyase branch of isomerase activity), not a lyase. The InterPro mapping is driven by the shared terpene synthase fold of class I enzymes such as KS. Supporting Evidence: PMID:20430888 Class II diterpene cyclases mediate the acid-initiated cycloisomerization reaction that serves as the committed step in biosynthesis of the large class of labdane-related diterpenoid natural products, which includes the important gibberellin plant hormones. |
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Download this section (compressed HTML)Q: Is CPS the main flux-controlling step of GA biosynthesis in vivo, and how important is Mg2+/GGPP substrate inhibition relative to transcriptional control of GA1?
Q: How is ent-CPP or a downstream early intermediate moved from provascular cells (where GA1 is expressed) to the cortex/endodermis cells that express late GA biosynthetic genes in germinating seeds?
Experiment: Express GA1 variants with altered Mg2+ sensitivity (His/Arg switch residue) in the ga1-3 background and measure GA levels and de-etiolation phenotypes to test the physiological relevance of Mg2+ inhibition.
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