Abstract
• First functional analysis of antagonistic EPF-like peptides in cotton. • GhEPF2-2 and GhEPFL9-6 show opposite roles via the GhERECTA1 receptor. • Cross-species assays confirm conserved yet divergent stomatal regulation. • GhEPFL9-6 enhances drought tolerance despite increasing stomatal density. • EPF-ERECTA signaling module provides promising targets for cotton breeding. Precise regulation of stomatal development is essential for optimizing water use efficiency and drought adaptation in crops. EPIDERMAL PATTERNING FACTOR (EPF) and EPF-LIKE (EPFL) peptides are well-established regulators of stomatal patterning in model species, yet their functional roles in cotton ( Gossypium hirsutum ) remain insufficiently characterized. Here, we investigate two cotton homologs, GhEPF2-2 and GhEPFL9-6 , and show that they exert contrasting effects on stomatal development and drought-associated physiological responses. Transcript profiling revealed leaf-preferential expression of both genes and opposite transcriptional responses to dehydration. Virus-induced gene silencing (VIGS) of GhEPF2-2 increased stomatal density and aggravated oxidative stress, whereas silencing GhEPFL9-6 reduced stomatal density and improved antioxidant status under PEG-induced dehydration. Heterologous expression in Arabidopsis thaliana further supported their divergent roles: GhEPF2-2 overexpression decreased stomatal density and altered antioxidant profiles, whereas GhEPFL9-6 overexpression increased stomatal density but exhibited improved physiological performance in controlled drought assays, likely through secondary changes in shoot architecture and osmotic adjustment rather than stomatal traits alone. Protein–protein interaction assays indicated that both peptides can interact with the receptor-like kinase GhERECTA1, suggesting their involvement in a shared signaling framework. Together, these findings reveal functional divergence of EPF/EPFL peptides in cotton and provide a basis for further evaluation of their roles in drought adaptation using stable genetic systems.
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Claims 13
Claim · model-written summary
GhEPF2-2 and GhEPFL9-6 are expressed preferentially in leaves.
- Taxon
- cotton
Gossypium hirsutumt0 - Organ / stage
- leafe0
- Outcome
- expressione0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Transcript profiling revealed leaf-preferential expression of both genes”
t0“in cotton”
Claim · model-written summary
The two genes show opposite transcriptional responses to dehydration.
- Taxon
- cottonGossypium hirsutumt0
- Treatment
- dehydratione0
- Outcome
- transcriptional responsese0
- Direction
- oppositee0
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“opposite transcriptional responses to dehydration”
t0“in cotton”
Claim · model-written summary
Virus-induced silencing of GhEPF2-2 increased stomatal density in cotton.
- Taxon
- cottonGossypium hirsutumt0
- Treatment
- Virus-induced gene silencing (VIGS) of GhEPF2-2e1
- Gene
- GhEPF2-2e1
- Outcome
- stomatal densitye1
- Direction
- increasede1
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Virus-induced gene silencing (VIGS) of GhEPF2-2 increased stomatal density and aggravated oxidative stress”
t0“in cotton”
Claim · model-written summary
Virus-induced silencing of GhEPF2-2 aggravated oxidative stress in cotton.
- Taxon
- cottonGossypium hirsutumt0
- Treatment
- Virus-induced gene silencing (VIGS) of GhEPF2-2e1
- Gene
- GhEPF2-2e1
- Outcome
- oxidative stresse1
- Direction
- aggravatede1
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Virus-induced gene silencing (VIGS) of GhEPF2-2 increased stomatal density and aggravated oxidative stress”
t0“in cotton”
Claim · model-written summary
Silencing GhEPFL9-6 reduced stomatal density in cotton under PEG-induced dehydration.
- Taxon
- cottonGossypium hirsutumt0
- Treatment
- silencing GhEPFL9-6e2
- Gene
- GhEPFL9-6e2
- Outcome
- stomatal densitye2
- Direction
- reducede2
- Qualifier
- under PEG-induced dehydratione2
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e2“silencing GhEPFL9-6 reduced stomatal density and improved antioxidant status under PEG-induced dehydration”
t0“in cotton”
Claim · model-written summary
Silencing GhEPFL9-6 improved antioxidant status in cotton under PEG-induced dehydration.
- Taxon
- cottonGossypium hirsutumt0
- Treatment
- silencing GhEPFL9-6e2
- Gene
- GhEPFL9-6e2
- Outcome
- antioxidant statuse2
- Direction
- improvede2
- Qualifier
- under PEG-induced dehydratione2
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e2“silencing GhEPFL9-6 reduced stomatal density and improved antioxidant status under PEG-induced dehydration”
t0“in cotton”
Claim · model-written summary
GhEPF2-2 overexpression decreased stomatal density in Arabidopsis thaliana.
- Taxon
- Arabidopsis thaliana
Arabidopsis thalianae3 - Treatment
- GhEPF2-2 overexpressione4
- Gene
- GhEPF2-2e4
- Outcome
- stomatal densitye4
- Direction
- decreasede4
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e3“Heterologous expression in Arabidopsis thaliana”
e4“GhEPF2-2 overexpression decreased stomatal density and altered antioxidant profiles”
Claim · model-written summary
GhEPF2-2 overexpression altered antioxidant profiles in Arabidopsis thaliana.
- Taxon
- Arabidopsis thalianaArabidopsis thalianae3
- Treatment
- GhEPF2-2 overexpressione4
- Gene
- GhEPF2-2e4
- Outcome
- antioxidant profilese4
- Direction
- alterede4
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e3“Heterologous expression in Arabidopsis thaliana”
e4“GhEPF2-2 overexpression decreased stomatal density and altered antioxidant profiles”
Claim · model-written summary
GhEPFL9-6 overexpression increased stomatal density in Arabidopsis thaliana.
- Taxon
- Arabidopsis thalianaArabidopsis thalianae3
- Treatment
- GhEPFL9-6 overexpressione5
- Gene
- GhEPFL9-6e5
- Outcome
- stomatal densitye5
- Direction
- increasede5
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e3“Heterologous expression in Arabidopsis thaliana”
e5“GhEPFL9-6 overexpression increased stomatal density but exhibited improved physiological performance in controlled drought assays, likely through secondary changes in shoot architecture and osmotic adjustment rather than stomatal traits alone”
Claim · model-written summary
GhEPFL9-6 overexpression improved physiological performance in controlled drought assays in Arabidopsis thaliana.
- Taxon
- Arabidopsis thalianaArabidopsis thalianae3
- Treatment
- GhEPFL9-6 overexpressione5
- Gene
- GhEPFL9-6e5
- Outcome
- physiological performancee5
- Direction
- improvede5
- Qualifier
- controlled drought assayse5
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e3“Heterologous expression in Arabidopsis thaliana”
e5“GhEPFL9-6 overexpression increased stomatal density but exhibited improved physiological performance in controlled drought assays, likely through secondary changes in shoot architecture and osmotic adjustment rather than stomatal traits alone”
Claim · model-written summary
The improved drought performance of GhEPFL9-6 overexpression plants may arise through secondary changes in shoot architecture and osmotic adjustment rather than stomatal traits alone.
- Taxon
- Arabidopsis thalianaArabidopsis thalianae3
- Gene
- GhEPFL9-6e5
- Outcome
- physiological performancee5
- Qualifier
- secondary changes in shoot architecture and osmotic adjustmente5
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e3“Heterologous expression in Arabidopsis thaliana”
e5“GhEPFL9-6 overexpression increased stomatal density but exhibited improved physiological performance in controlled drought assays, likely through secondary changes in shoot architecture and osmotic adjustment rather than stomatal traits alone”
Claim · model-written summary
Both peptides interact with the receptor-like kinase GhERECTA1 in protein-protein interaction assays.
- Taxon
- cottonGossypium hirsutumt0
- Gene
- GhERECTA1e6
- Outcome
- interacte6
- Qualifier
- both peptidese6
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e6“Protein–protein interaction assays indicated that both peptides can interact with the receptor-like kinase GhERECTA1”
t0“in cotton”
Claim · model-written summary
GhEPF2-2 and GhEPFL9-6 may act in a shared GhERECTA1 signaling framework.
- Taxon
- cottonGossypium hirsutumt0
- Gene
- GhERECTA1e6
- Outcome
- shared signaling frameworke7
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e6“Protein–protein interaction assays indicated that both peptides can interact with the receptor-like kinase GhERECTA1”
e7“suggesting their involvement in a shared signaling framework”
t0“in cotton”
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