EPIDERMAL PATTERNING FACTOR 2 (EPF2) is a small secreted cysteine-rich peptide hormone of the EPF/EPFL family (120-aa precursor with a signal peptide; the mature C-terminal peptide MEPF2 is held in a scaffold by three/four disulfide bonds and a functional loop). It is produced by meristemoid mother cells (MMCs) and their early stomatal-lineage descendants and acts non-cell-autonomously on neighbouring protodermal cells to limit how many cells enter, and remain in, the stomatal lineage. MEPF2 binds directly to the leucine-rich-repeat ectodomain of the receptor kinase ERECTA (its principal receptor; ERL1 is also bound) in a complex with the receptor-like co-receptor TOO MANY MOUTHS (TMM), and triggers rapid activation of the YODA-MKK4/5-MPK3/6 MAP kinase cascade that phosphorylates and destabilizes the stomatal-entry bHLH SPEECHLESS. EPF2 is antagonized by the positive peptide stomagen/EPFL9, which competes for the same ERECTA-TMM receptor, and is inactivated by cleavage by the CO2-induced secreted protease CRSP. Loss of EPF2 produces excess small stomatal-lineage cells and higher stomatal density, whereas overexpression nearly abolishes stomata; EPF2 is required for the reduction of stomatal development at elevated CO2.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005576 extracellular region | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt subcellular-location mapping (Secreted) to extracellular region. Reason: EPF2 has an N-terminal signal peptide and acts as a mobile extracellular peptide perceived at the cell surface by ERECTA-family receptors; extracellular localization is correct. Supporting Evidence: PMID:19435754 We demonstrate that a small secretory peptide, EPIDERMAL PATTERING FACTOR 2 (EPF2), is produced by the MMC and its early descendants |
| GO:0005576 extracellular region | ISM GO_REF:0000122 | ACCEPT | Summary: Sequence-model (signal peptide) prediction of extracellular localization. Reason: Consistent with the secreted-peptide nature of EPF2 and its direct binding to the ectodomains of ERECTA-family receptors. Supporting Evidence: PMID:22241782 Here we report the direct, robust, and saturable binding of bioactive EPF peptides to the ERECTA family. |
| GO:0007165 signal transduction | IEA GO_REF:0000108 | ACCEPT | Summary: Inter-ontology inference from receptor ligand activity to signal transduction. Reason: EPF2 initiates ERECTA/TMM receptor signalling that activates MPK3/MPK6; as the ligand it is part of this cell-cell signalling pathway. The term is broad but correct; the specific biological output is captured by negative regulation of stomatal complex development. Supporting Evidence: PMID:26083750 Application of MEPF2 peptides to Arabidopsis wild-type seedlings rapidly elicited phosphorylation of MPK3 and MPK6 in 10 min, a characteristic signature of MAPK activation |
| GO:0010052 guard cell differentiation | IBA GO_REF:0000033 | MODIFY | Summary: PAINT inference of involvement in guard cell differentiation from the EPF/EPFL family node. Reason: EPF2 does not drive guard cell differentiation; it acts as an inhibitory cue that limits entry of protodermal cells into the stomatal lineage (MMC fate) and asymmetric entry divisions. The family-level process is better and more accurately expressed for EPF2 by its regulatory role, negative regulation of stomatal complex development. The IBA is not wrong about the family being stomatal regulators, but 'guard cell differentiation' places EPF2 at the wrong step and in the wrong direction. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: REGULATORY SIGN INVERSION GRANULARITY MISMATCH Sources checked: PANTHER:PTN002104517 SUPPORTS SOURCE BUT NOT TARGET The node covers EPF-family peptides; for EPF2 the only experimental evidence is inhibitory and acts at stomatal lineage entry, so a positive guard cell differentiation process overstates the role. AGI_LocusCode:AT1G34245 · EPF2 SUPPORTS SOURCE BUT NOT TARGET The seed is EPF2's own acts-upstream IMP annotation; that genetic evidence supports an inhibitory regulator, better captured by GO:2000122. Proposed replacements: negative regulation of stomatal complex development Supporting Evidence: PMID:19435754 Our results suggest that EPF2 inhibits cells from adopting the MMC fate in a non-cell-autonomous manner, thus limiting the number of MMCs. |
| GO:0010052 guard cell differentiation | IMP PMID:19398336 The signaling peptide EPF2 controls asymmetric cell division... | MODIFY | Summary: epf2 mutants and EPF2 overexpressors alter stomatal production (Hunt and Gray 2009). Reason: The phenotype (excess stomatal-lineage cells in the mutant; virtually no stomata on overexpression) shows that EPF2 is an inhibitory regulator acting at stomatal lineage entry rather than at the GMC-to-guard-cell transition. The evidence is better captured by negative regulation of stomatal complex development. Proposed replacements: negative regulation of stomatal complex development Supporting Evidence: PMID:19398336 In the absence of EPF2, excessive numbers of cells enter the stomatal lineage and produce numerous small epidermal cells that express stomatal lineage reporter genes, whereas plants overexpressing EPF2 produce virtually no stomata. |
| GO:0010374 stomatal complex development | IBA GO_REF:0000033 | ACCEPT | Summary: PAINT inference of stomatal complex development for the EPF/EPFL clade. Reason: The EPF/EPFL peptides (EPF1, EPF2, EPFL9/stomagen) are a phylogenetically coherent clade of stomatal-development regulators; EPF2 is inside that clade and has direct experimental support. The more specific regulatory term (GO:2000122) is also annotated. Supporting Evidence: PMID:19398336 EPF2 expression during leaf development affects stomatal density on the mature leaf. |
| GO:0010374 stomatal complex development | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO mapping from the EPF domain (IPR039455) to stomatal complex development. Reason: Consistent with experimental data on EPF2; broad but correct. Supporting Evidence: PMID:19398336 EPF2 expression during leaf development affects stomatal density on the mature leaf. |
| GO:0010374 stomatal complex development | IMP PMID:19398336 The signaling peptide EPF2 controls asymmetric cell division... | ACCEPT | Summary: Loss- and gain-of-function of EPF2 alter stomatal density and lineage entry. Reason: Direct genetic evidence; EPF2 is a negative regulator of stomatal development. Accepted at this general level, with the specific negative-regulation term carrying the core meaning. Supporting Evidence: PMID:19398336 In the absence of EPF2, excessive numbers of cells enter the stomatal lineage and produce numerous small epidermal cells that express stomatal lineage reporter genes, whereas plants overexpressing EPF2 produce virtually no stomata. |
| GO:0019901 protein kinase binding | IPI PMID:26083750 Competitive binding of antagonistic peptides fine-tunes stom... | MODIFY | Summary: Binding of EPF2 to TMM (AT1G80080) reported in Lee et al. 2015. Reason: TMM is a receptor-like protein that lacks a cytoplasmic kinase domain, so 'protein kinase binding' is an incorrect description of this interaction. The interaction reflects EPF2's activity as a peptide ligand of the ERECTA-TMM receptor complex, which is captured by receptor ligand activity. Proposed replacements: receptor ligand activity Supporting Evidence: PMID:26083750 Both EPF2 and Stomagen directly bind to ER and its co-receptor TOO MANY MOUTHS. PMID:22241782 Unlike ERECTA family RKs, TMM lacks a cytoplasmic effector domain |
| GO:0019901 protein kinase binding | IPI PMID:26083750 Competitive binding of antagonistic peptides fine-tunes stom... | MODIFY | Summary: Binding of EPF2 to the ERECTA receptor kinase (AT2G26330). Reason: The interaction is real (direct, saturable binding of MEPF2 to the ERECTA ectodomain), but it is a ligand-receptor interaction with the extracellular LRR domain, not binding to the kinase as an enzyme. Receptor ligand activity is the informative molecular function. Proposed replacements: receptor ligand activity Supporting Evidence: PMID:26083750 Both EPF2 and Stomagen directly bind to ER and its co-receptor TOO MANY MOUTHS. PMID:22241782 Our results place the ERECTA family as the primary receptors for EPFs with TMM as a signal modulator and establish EPF2-ERECTA and EPF1-ERL1 as ligand-receptor pairs specifying two steps of stomatal development |
| GO:0048018 receptor ligand activity | IBA GO_REF:0000033 | ACCEPT | Summary: PAINT inference of receptor ligand activity for EPF-family peptides. Reason: Core molecular function. MEPF2 binds the ERECTA (and ERL1) ectodomain and TMM, competes with stomagen for ERECTA, and elicits rapid MPK3/MPK6 activation, i.e. it is an agonist ligand of the ERECTA-TMM receptor complex. Supporting Evidence: PMID:22241782 MEPF2, unlike MEPF1, exhibited binding to TMM in addition to ERECTA and ERL1 PMID:26083750 Application of MEPF2 peptides to Arabidopsis wild-type seedlings rapidly elicited phosphorylation of MPK3 and MPK6 in 10 min, a characteristic signature of MAPK activation |
| GO:0090626 plant epidermis morphogenesis | IMP PMID:19435754 Epidermal cell density is autoregulated via a secretory pept... | KEEP AS NON CORE | Summary: epf2 mutants show increased density of guard and non-guard epidermal cells. Reason: EPF2 controls epidermal cell density because the stomatal lineage produces many non-guard (pavement) cells; this epidermal phenotype is a consequence of its core role in limiting stomatal lineage entry. Supporting Evidence: PMID:19435754 negatively regulates the density of guard and non-guard epidermal cells |
| GO:0160109 plant gross anatomical part developmental process | IEA GO_REF:0000117 | KEEP AS NON CORE | Summary: ARBA machine-learning annotation to a very general plant developmental process. Reason: Not wrong (EPF2 influences leaf epidermal development) but very general and uninformative relative to the experimentally supported negative regulation of stomatal complex development. |
| GO:2000122 negative regulation of stomatal complex development | IMP PMID:19435754 Epidermal cell density is autoregulated via a secretory pept... | ACCEPT | Summary: EPF2 overexpression suppresses and loss of EPF2 increases stomatal development. Reason: Core biological role. EPF2 is a non-cell-autonomous inhibitor of MMC fate and stomatal lineage entry, acting through ERECTA/TMM and the MAPK cascade; it also mediates repression of stomatal development at elevated CO2. Supporting Evidence: PMID:19435754 Our results suggest that EPF2 inhibits cells from adopting the MMC fate in a non-cell-autonomous manner, thus limiting the number of MMCs. PMID:25043023 Moreover, EPF2 is essential for CO2 control of stomatal development. |
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Download this section (compressed HTML)Q: What is the processed, bioactive form of EPF2 in planta, and which proteases (besides CRSP, which inactivates it) generate or degrade MEPF2 in the apoplast?
Q: How does EPF2 binding to the ERECTA-TMM complex recruit SERK co-receptors and activate the receptor kinase, given that stomagen binds the same receptor without activating MAPKs?
Experiment: Quantify in-planta MEPF2 levels and processing (e.g. targeted peptidomics of apoplastic fluid) in wild type, crsp and elevated-CO2 conditions to link CO2-dependent EPF2 abundance to ERECTA signalling output.
Type: peptidomics / mass spectrometry
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