Surviving The Deluge: Flooding Stress Flashcards
Plants and submergence
- Water excess, relatively common stress
- several wild species, but few crops thrive in such an environment
Water logging
Only the below-ground part is under water saturating conditions
Flooding
Partial and complete submergence
Partial submergence
Root system and portion of shoot underwater
Complete submergence
Whole plant covered
Economic impact of crop flooding
Largest stressor!
Dynamics of flooding events
- intensity, timing, duration = changing
- frequent: UK, CE, Balkan area
Why is submergence a stress to plants?
- anoxia
- anaerobic activity of roots and rhizosphere
ROS
- affect mitochondria and chloroplasts
- < photosynthesis
Submergence process:
- Soil redox potential «
- Accumulation of toxic compounds (Mn2+, Fe2+, H2S)
- Gases diffuse 10^4 slower in water than air (severely «_space;O2, CO2 availability; ethylene entrapment)
- Fermentation
- Carbon starvation
- «_space;photosynthesis
Anaerobic metabolism
- fermentations necessary to replenish glycolysis NAD+
Fermentation process
- Starch -> soluble sugars
- Soluble sugars -(glycolysis)-> pyruvate (NAD+ -> NADH; ADP -> ATP)
- Pyruvate -lactate dehydrogenase-> lactate (toxic! Acidifies, damages cell) (NADH -> NAD+)
OR - Pyruvate -pyruvate decarboxylase-> acetylaldehyde (toxic!)
- Acetylaldehyde -alcohol dehydrogenase-> ethanol (preferred!) (NADH -> NAD+)
Phytoglobins
- plant Hbs
- v high O2 affinity
- no long distance transport
- nitrate reductase: NAD+ regeneration
Nitrite
Alternative e- acceptor in mETC
A. thaliana ERFVIIs TFs
- 5x total
- 2x hypoxia-inducible (relative expression level across hypoxia time course)
- transient/permanent up regulation
- v conserved N terminus
- cysteine residue followed by 2x glycines
- very rare feature of proteins
N-degron pathway
Determines protein (in)stability depending on exposed aas
Oxygen-dependent oxidation of N-terminal cysteine (+R)
Prepares proteins for degradation
Cysteine in 2nd position
- dangerous!
- exposure R residue; degradation signal
- PTM + R
- 4x possible pathways
N-degron 4x pathway 1
- MC -MetAP-> C
- C -> *C
- *C -> RC
- RC -NO-> degradation
N-degron pathway 2
- MC -> NQ
- NQ -NTAN/NTAQ-> DE
- DE -ART6/VBR1-> R
- R-Ub-> degradation
N-degron pathway 3
- MC -endoproteolytic cleavage-> DE
N-degron pathway 4
- MC -> R
Hiding from N-degron?
Mask/decorate
N-degron ERFVII regulation
- priming effect
N-degron mutant analysis
- mimic hypoxia
- ate1/ate2, prt6 -> mutant genes are core hypoxia responders
RAP2.12
wt = degraded
wt (hypoxia) = stabilised
ate1ate2 = stabilised
erfr11 = less DEGs under hypoxia (1% O2)
High O2
- O2 + HIF-1α tags with Pro
- PH tags with OH
- pVHL targets for degradation
Low O2 sensing in mammals
No PH
PH
- prolyl hydroxylase
- transcriptionally regulated by HIF-1α
Summarising O2 sensing in mammals
- O2 dependent enzymatic proline residue hydroxylation in HIF-1α TF stimulates proteasomal degradation
- stabilised HIF-1α induces metabolic + developmental genes (e.g. angiogenesis)
For O2 sensing, both plants and animals rely on
aerobic degradation of a constitutively produced TF
Can N-cysteine oxidation be enzymatically catalysed in plants?
- localise in cytosol + nucleus (GFP localisation)
- 2x hypoxia inducible
- regulate other nuclear proteins?
Cysteine oxidase in planta
Oxygen sensors! PCOs
Oxygen sensing
Cysteine -O2 dependent cysteine oxidase- cysteine sulfunic acid
HUP29, 43
Cysteinyl dioxygenase activity; PCOs
PCO biochemical characterisation
Mass spectrometry analyses have confirmed that PCO can add 2x O2 atoms to ERFVII N-termini
ERFVII interacting proteins
- RAP2.12 interacts with ACBPs in PM
- shown by biomolecular fluorescence complementation assay; photoconvertible + UV
RAP2.12
Constitutive ERFVII
RAP2.12 behaviour under hypoxia
Nuclear localisation
HRPE
- ERFVII enhancer
- compare hypoxia promotors for signature
- trim + luciferase fusion construct: measure expression under hypoxia
- C9 motif KO: necessary
- 32nt 3x repeat KI: sufficient
ERFVII TF
Relocalisation and stabilisation in buckets to activate hypoxia genes
Can we target ERFVIIs in plant breeding? Logic
- modulating N-degron pathway (prt6) ; does this enhance plant flooding survival ??
- hypoxia tolerance assays in vitro
- submergence assays in soil
Can we target ERFVIIs in plant breeding? Results
- HRE1 hyper-expression = +ve
- hyperstability (masking N-terminal cysteine) severely compromises plant development
- silencing / spontaneous mutation = +ve
Hormone homeostasis under submergence
- phytohormone biosynthesis -> require O2 (+ATP) cosubstrate(s)
- hormone metabolism depends on enzyme oxygen affinity
Ethylene
- gaseous
- biosynthesis enzymes induced in hypoxia in several sp.
- ACS, ACO
Ethylene biosynthesis
Methionine -AdoMet synthetase-> S-Ado-Met -ATS-> amino-cyclopropanone carboxylic acid -ACO (O2->CO2) -> ethylene
Acclimation
- ethylene pretreatment = increased hypoxia tolerance
- ethylene + hypoxia-induced phytoglobin can scavenge NO (uses O2 as substrate)
How are phytoglobins induced?
By ethylene
Ethylene induces ERFVII stabilisation
- CPTIO = NO scavenging partner
- does not accumulate in ethylene treated A. thaliana roots
- stabilises RAP2.13
ANACO13
- early response TF
- hypoxia-induced via promotors
- homologs: -16, -17
Membrane-associated NAC-TFs
- ANACO13: ER-localised
- cleaved under mitochondrial stress
- goes to nucleus
What cleaves ANACO13?
- chemical mutagenesis analysis
- rhomboid proteases cleave substrates inside membranes
- rbl6/rbl2: no ANACO13 nuclear relocalisation
Does ANACO13 contribute to hypoxia tolerance?
silencing/ inhibiting ER release using artificial miRNAS results in < hypoxia tolerance + bleaching
