Rice momilactone A biosynthesis

Biosynthesis of the rice diterpenoid momilactone A from geranylgeranyl diphosphate. Cyclases construct a syn-pimaradiene scaffold, cytochrome P450 enzymes oxidize it, and short-chain dehydrogenases support lactone and ketone formation. The chromosome 4 biosynthetic gene cluster supplies only part of this pathway: additional P450 enzymes are encoded on chromosomes 2, 6, and 1. Momilactones participate in plant chemical defense and allelopathy. An optional C20-hydroxylation branch records an activity implicated in momilactone B production; a complete ordered route to B is not represented.

MODULE:rice_momilactone_biosynthesisDRAFTCONCRETEMetabolic Pathwaymodules/rice_momilactone_biosynthesis.yaml
momilactone diterpenoid biosynthesisGO:0016102 chromosome 4 momilactone biosynthetic gene clusterMIBiG:BGC0000671
MIBiG:BGC0000671
Momilactone B biosynthetic gene cluster from Oryza sativa Japonica Group
Records the chromosome 4 cluster. Version 3 reports questionable quality and unknown completeness; cluster membership is not evidence of pathway self-sufficiency.
PMID:17872948
Identification of a biosynthetic gene cluster in rice for momilactones.
Initial functional identification of the chromosome 4 cluster and joint genetic evidence for CYP99A2/CYP99A3.
PMID:33793769
Interdependent evolution of biosynthetic gene clusters for momilactone production in rice.
Combines rice genetics and enzyme assays to resolve the interleaved chromosome 4/chromosome 2 route through a hemiacetal intermediate.
PMID:33106662
Rerouting plant terpene biosynthesis enables momilactone pathway elucidation.
Complete heterologous reconstitution establishes a route to momilactones A and B, including CYP76M14, while leaving native reaction ordering partly unresolved.
PMID:39887739
Evolution and diversification of the momilactone biosynthetic gene cluster in the genus Oryza.
Comparative Oryza research places CYP76M8, CYP701A8, and CYP76M14 outside the chromosome 4 cluster and supports limiting the exact arrangement to the specified rice taxon.
file:ORYSJ/CPS4/CPS4-ai-review.yaml
Rice CPS4 gene review
Existing gene review for the upstream syn-copalyl diphosphate synthase.
file:modules/rice_momilactone_biosynthesis-notes.md
Rice momilactone primary-source curation notes
Records pathway boundaries, source checks, protein grounding, and distinctions between established chemistry and uncertain native ordering.
file:modules/rice_momilactone_biosynthesis-deep-research-manual.md
Manual primary-literature research for rice momilactone biosynthesis
Manually checked primary studies and database records; the attempted Falcon run timed out without producing a report.

The boundary starts with supplied geranylgeranyl diphosphate and ends at momilactone A, with an optional C20-hydroxylation extension implicated in B biosynthesis. General precursor supply, NAD(P)H/flavin-dependent P450 electron delivery, transport, secretion, and induction circuitry are outside this module. Native targeting of the early cyclases must not be inferred from engineered cytosolic expression in Nicotiana benthamiana. Protein participants are specific japonica accessions; no PANTHER ancestral-node or universal plant-family claim is made. Precise descriptors without GO identifiers are intentional where retrieved GO definitions name a different substrate or oxidation endpoint. The two C3 alternatives express evidence-supported biochemical capabilities, not an assertion of their quantitative contributions in native rice. The current protein grounding covers one assayed MAS paralog and does not exhaust native SDR alternatives.

16Nodes
11Parts
2Variant Sets
4Variants
10Annotons
7Connections

Derived QC

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Module deep research

✓ present

  • rice_momilactone_biosynthesis-deep-research-manual.md (manual)

Leaf nodes lacking representative members

✓ every leaf node grounds to a representative protein.

Template conformance

✓ every declared conforms_to bundle matches its template motif.

Gene-review completeness (1/8 grounded genes reviewed)

1 complete review(s) · 1 with deep research · 7 missing review · 0 reviewed but lacking deep research

Gene Review Complete Deep research
CPS4 Q0JF02 ✓ ✓ ✓
CYP701A8 Q0DBF4 ✗ — —
KSL4 Q0JEZ8 ✗ — —
CYP99A3 Q0JF01 ✗ — —
CYP76M8 Q6YTF1 ✗ — —
OsMAS Q7FAE1 ✗ — —
CYP99A2 Q7X7X4 ✗ — —
CYP76M14 Q8LJD2 ✗ — —

Details

Context
Oryza sativa subsp. japonicaNCBITaxon:39947
Rice momilactone A synthesis with a documented C20 extensionMetabolic Pathwayrice_momilactone_biosynthesis
momilactone diterpenoid biosynthesisGO:0016102 chromosome 4 momilactone biosynthetic gene clusterMIBiG:BGC0000671
Context
Oryza sativa subsp. japonicaNCBITaxon:39947

Required parts describe a route to momilactone A. The optional CYP76M14 part documents C20 hydroxylation implicated in momilactone B production; its absence must not be interpreted as loss of A biosynthesis. The graph ends at A and the C20-hydroxylated lactone, and does not encode a complete B-producing route or the unresolved convergence/order of C3 and C20 tailoring. Display order and connections describe supported chemical flow, not physical order of genes along chromosomes. The C20 branch is drawn from the lactone substrate demonstrated experimentally, rather than as a directly assayed momilactone A-to-B conversion. Module satisfaction alone addresses A capacity; B additionally requires CYP76M14 and the shared tailoring machinery.

Connections

ksl4_step -> c19_oxidation Provides Input For
Syn-pimaradiene enters C19 oxidation.
osmas_lactone_step -> c3_tailoring Provides Input For
The lactone supplies the C3-tailoring routes.
osmas_lactone_step -> cyp76m14_step Provides Input For
The pre-C3 lactone is a demonstrated CYP76M14 substrate. This branch combines with the shared C3-tailoring capacity to yield momilactone B; the native order of the C3 and C20 reactions remains unresolved.
Part 1: diterpene scaffold formation
Cyclization to the syn-pimaradiene scaffoldMetabolic Pathwayscaffold_formation

Connections

cps4_step -> ksl4_step Provides Input For
CPS4 supplies syn-copalyl diphosphate to KSL4.
Part 1: syn-copalyl diphosphate formation
CPS4: syn-copalyl diphosphate formationReactioncps4_step

CPS4 also supplies other syn-copalyl-diphosphate-derived diterpenoids; this entry step is not specific to momilactones.

Annotons

CPS4: syn-copalyl diphosphate formation
cps4_step_activity
Participant: Gene: CPS4
Gene:
CPS4UniProtKB:Q0JF02 Os04g0178300 / LOC_Os04g09900; chromosome 4 cluster.

Function

syn-copalyl diphosphate synthase activityGO:0051498
Substrates: geranylgeranyl diphosphate
Products: syn-copalyl diphosphate

syn-copalyl diphosphate synthase activity

PMID:15255861
Functional identification of rice syn-copalyl diphosphate synthase and its role in initiating biosynthesis of diterpenoid phytoalexin/allelopathic natural products.
Biochemical identification of rice syn-copalyl diphosphate synthase.
Part 2: committed scaffold formation
KSL4: committed syn-pimaradiene scaffold formationReactionksl4_step

Annotons

KSL4: committed syn-pimaradiene scaffold formation
ksl4_step_activity
Participant: Gene: KSL4
Gene:
KSL4UniProtKB:Q0JEZ8 Os04g0179700 / LOC_Os04g10060; chromosome 4 cluster.

Function

syn-pimara-7,15-diene synthase activityGO:0034279
Substrates: syn-copalyl diphosphate
Products: syn-pimara-7,15-diene diphosphate

syn-pimara-7,15-diene synthase activity

PMID:15299118
Identification of syn-pimara-7,15-diene synthase reveals functional clustering of terpene synthases involved in rice phytoalexin/allelochemical biosynthesis.
Direct identification of the rice syn-pimaradiene-forming enzyme.
Part 2: lactone ring formation across cluster boundaries
C19 and C6 oxidation followed by lactonizationMetabolic Pathwaylactone_formation

Connections

c19_oxidation -> cyp76m8_step Provides Input For
The CYP99A aldehyde product is the preferred CYP76M8 substrate.
cyp76m8_step -> osmas_lactone_step Provides Input For
The C6-hydroxy C19-aldehyde cyclizes to the hemiacetal substrate of OsMAS; this intervening closure appears spontaneous.
Part 1: C19 aldehyde formation
C19 oxidation by CYP99A2 or CYP99A3Reactionc19_oxidation

Alternative catalytic capacity does not establish that either single gene is individually dispensable in every native context; the original genetic perturbation targeted both paralogs.

Variant set: Biochemically supported CYP99A alternatives by enzyme paralog (One Or More)
CYP99A2: C19 aldehyde formationReactioncyp99a2_c19

GO:0036209 describes complete oxidation through the C19 carboxylate. It is deliberately not attached to this aldehyde-output role.

Annotons

CYP99A2: C19 aldehyde formation
cyp99a2_c19_activity
Participant: Gene: CYP99A2
Gene:
CYP99A2UniProtKB:Q7X7X4 Os04g0180400 / LOC_Os04g10160; chromosome 4 cluster.

Function

successive C19 oxidations of syn-pimaradiene to syn-pimaradien-19-al
Substrates: syn-pimara-7,15-diene
Products: syn-pimaradien-19-al

successive C19 oxidations of syn-pimaradiene to syn-pimaradien-19-al

PMID:33793769
Interdependent evolution of biosynthetic gene clusters for momilactone production in rice.
Separate E. coli co-expression experiments paired CYP76M8 with either CYP99A2 or CYP99A3 and produced the C6-hydroxy C19-aldehyde intermediate (Results, Figure 4). Cell-free substrate-feeding assays favored CYP99A-dependent C19 oxidation before CYP76M8 hydroxylation (Figure 5). These are individual paralog assays, distinct from joint knockdown.
CYP99A3: C19 aldehyde formationReactioncyp99a3_c19

GO:0036209 describes complete oxidation through the C19 carboxylate. It is deliberately not attached to this aldehyde-output role.

Annotons

CYP99A3: C19 aldehyde formation
cyp99a3_c19_activity
Participant: Gene: CYP99A3
Gene:
CYP99A3UniProtKB:Q0JF01 Os04g0178400 / LOC_Os04g09920; chromosome 4 cluster.

Function

successive C19 oxidations of syn-pimaradiene to syn-pimaradien-19-al
Substrates: syn-pimara-7,15-diene
Products: syn-pimaradien-19-al

successive C19 oxidations of syn-pimaradiene to syn-pimaradien-19-al

PMID:33793769
Interdependent evolution of biosynthetic gene clusters for momilactone production in rice.
Separate E. coli co-expression experiments paired CYP76M8 with either CYP99A2 or CYP99A3 and produced the C6-hydroxy C19-aldehyde intermediate (Results, Figure 4). Cell-free substrate-feeding assays favored CYP99A-dependent C19 oxidation before CYP76M8 hydroxylation (Figure 5). These are individual paralog assays, distinct from joint knockdown.
Part 2: C6 hydroxylation and nonenzymatic hemiacetal formation
CYP76M8: C6beta hydroxylation before hemiacetal closureReactioncyp76m8_step

The hydroxy-aldehyde subsequently forms the 19,6beta-hemiacetal, apparently spontaneously. GO:0102612 specifies unoxidized syn-pimaradiene, so it is not used for this aldehyde-substrate role.

Annotons

CYP76M8: C6beta hydroxylation before hemiacetal closure
cyp76m8_step_activity
Participant: Gene: CYP76M8
Gene:
CYP76M8UniProtKB:Q6YTF1 Os02g0569400 / LOC_Os02g36070; chromosome 2 diterpenoid cluster. Its UniProt name describes another substrate-specific role.

Function

C6beta hydroxylation of syn-pimaradien-19-al
Substrates: syn-pimaradien-19-al
Products: 6beta-hydroxy-syn-pimaradien-19-al

C6beta hydroxylation of syn-pimaradien-19-al

PMID:33793769
Interdependent evolution of biosynthetic gene clusters for momilactone production in rice.
Biochemical ordering and rice genetic evidence place the chromosome 2 enzyme between chromosome 4-encoded reactions.
Part 3: hemiacetal oxidation
OsMAS: oxidation of the hemiacetal to the lactoneReactionosmas_lactone_step

This reaction is distinct from the C3-alcohol oxidation described by GO:0102960. Lactone formation proceeds by hemiacetal oxidation, not simple dehydration of a C19 carboxylic acid and a C6 alcohol. Only the fully mapped OsMAS/AK103462 protein Q7FAE1 is represented here. The second assayed MAS paralog is an acknowledged alternative whose exact full-length experimental sequence-to-UniProt mapping was not established; absence of Q7FAE1 alone must not be interpreted as proof of absent native lactonization capacity.

Annotons

OsMAS: oxidation of the hemiacetal to the lactone
osmas_lactone_step_activity
Participant: Gene: OsMAS
Gene:
OsMASUniProtKB:Q7FAE1 Os04g0179200 / LOC_Os04g10010; chromosome 4 cluster. Experimentally studied clone AK103462; accession and locus disambiguate inconsistent MS1/MAS1 numbering.

Function

oxidation of the 19,6beta-hemiacetal to syn-pimaradien-19,6beta-olide
Substrates: syn-pimaradien-19,6beta-hemiacetal
Products: syn-pimaradien-19,6beta-olide

oxidation of the 19,6beta-hemiacetal to syn-pimaradien-19,6beta-olide

PMID:33793769
Interdependent evolution of biosynthetic gene clusters for momilactone production in rice.
Assayed clustered SDRs oxidize the hemiacetal to the lactone; the lactone intermediate was also detected in rice.
PMID:33106662
Rerouting plant terpene biosynthesis enables momilactone pathway elucidation.
OsMAS supports lactone formation in the reconstructed plant pathway.
Part 3: formation of momilactone A
C3 ketone formation yields momilactone AMetabolic Pathwayc3_tailoring

The alternatives represent biochemical capacity; their relative contributions in native rice are not established by heterologous reconstruction. OsMAS can participate at both lactone formation and C3 oxidation without representing two different proteins.

Variant set: Supported divisions of C3 oxidative labor by enzyme contribution to terminal C3 oxidation (One Or More)
CYP701A8-supported C3 tailoring to momilactone AReactioncyp701a8_direct_route

A biochemically supported route, not a claim that CYP701A8 supplies all native C3 oxidation. Product reconstruction in a heterologous plant does not exclude assistance from host oxidoreductases. No substrate-mismatched GO term is assigned.

Annotons

CYP701A8-supported C3 tailoring to momilactone A
cyp701a8_direct_route_activity
Participant: Gene: CYP701A8
Gene:
CYP701A8UniProtKB:Q0DBF4 Os06g0569500 / LOC_Os06g37300; chromosome 6, outside the chromosome 4 cluster. The UniProt name describes activity on a different diterpene.

Function

C3 oxidative tailoring of the lactone toward the momilactone A ketone
Substrates: syn-pimaradien-19,6beta-olide
Products: momilactone A

C3 oxidative tailoring of the lactone toward the momilactone A ketone

PMID:33793769
Interdependent evolution of biosynthetic gene clusters for momilactone production in rice.
Cell-free CYP701A8 assays with the lactone produced small amounts of momilactone A, suggesting sequential C3 hydroxylation and oxidation.
CYP701A8 hydroxylation followed by SDR oxidationMetabolic Pathwaycyp701a8_sdr_route

Connections

The hydroxylated lactone supplies the SDR substrate.
Part 1: C3 hydroxylation
CYP701A8: C3beta hydroxylationReactioncyp701a8_hydroxylation_step

The specific UniProt-linked GO term for ent-sandaracopimaradiene hydroxylation has a different substrate and is not used here.

Annotons

CYP701A8: C3beta hydroxylation
cyp701a8_hydroxylation_step_activity
Participant: Gene: CYP701A8
Gene:
CYP701A8UniProtKB:Q0DBF4 Os06g0569500 / LOC_Os06g37300; chromosome 6, outside the chromosome 4 cluster. The UniProt name describes activity on a different diterpene.

Function

C3beta hydroxylation of syn-pimaradien-19,6beta-olide
Substrates: syn-pimaradien-19,6beta-olide
Products: 3beta-hydroxy-syn-pimaradien-19,6beta-olide

C3beta hydroxylation of syn-pimaradien-19,6beta-olide

PMID:33793769
Interdependent evolution of biosynthetic gene clusters for momilactone production in rice.
The proposed SDR-assisted route places CYP701A8-mediated C3 hydroxylation before oxidation of the alcohol to the momilactone A ketone.
Part 2: C3 alcohol oxidation
OsMAS: C3 alcohol oxidation to momilactone AReactionosmas_c3_oxidation_step

Annotons

OsMAS: C3 alcohol oxidation to momilactone A
osmas_c3_oxidation_step_activity
Participant: Gene: OsMAS
Gene:
OsMASUniProtKB:Q7FAE1 Os04g0179200 / LOC_Os04g10010; chromosome 4 cluster. Experimentally studied clone AK103462; accession and locus disambiguate inconsistent MS1/MAS1 numbering.

Function

momilactone A formation by oxidation of the C3 alcoholGO:0102960
Substrates: 3beta-hydroxy-syn-pimaradien-19,6beta-olide NAD+ or NADP+
Products: momilactone A NADH or NADPH proton

momilactone A formation by oxidation of the C3 alcohol

PMID:17872948
Identification of a biosynthetic gene cluster in rice for momilactones.
The clustered OsMAS converts the C3-hydroxy lactone into momilactone A.
PMID:27337377
Investigating inducible short-chain alcohol dehydrogenases/reductases clarifies rice oryzalexin biosynthesis.
The two clustered SDRs have different efficiencies on the C3-hydroxy substrate.
Part 4: additional C20 oxidation toward momilactone B (optional)
CYP76M14: C20 hydroxylation enabling momilactone B formationReactioncyp76m14_step

C20 hydroxylation was demonstrated on the pre-C3 lactone. Together with CYP701A8-dependent C3 tailoring, this activity supports production of momilactone B and its additional hemiacetal ring. No separate ring-closing enzyme is asserted. The exact native order of C3 and C20 oxidation is unresolved; this annoton does not assert that isolated momilactone A was directly assayed as the CYP76M14 substrate.

Annotons

CYP76M14: C20 hydroxylation enabling momilactone B formation
cyp76m14_step_activity
Participant: Gene: CYP76M14
Gene:
CYP76M14UniProtKB:Q8LJD2 Os01g0561600; chromosome 1, outside the chromosome 4 cluster. The CYP76M14 symbol is verified in RAP-DB; this UniProt entry is unreviewed.

Function

C20 hydroxylation of the momilactone scaffold
Substrates: syn-pimaradien-19,6beta-olide
Products: 20-hydroxy-syn-pimaradien-19,6beta-olide

C20 hydroxylation of the momilactone scaffold

PMID:33106662
Rerouting plant terpene biosynthesis enables momilactone pathway elucidation.
CYP76M14 supplies C20 oxidation in the complete reconstruction yielding momilactone B.