Post-Translational regulation in plant development and abiotic stress response 6-3-2009 Isabel Abreu
Post-Translational regulation in plant development and abiotic stress
response
6-3-2009
Isabel Abreu
Stress
Signal perception
RESPONSE
Tolerance
Survival
Death
Regulating the regulator
Transcriptional Regulation
Alternative splicing– compartmentalization; functional differences
miRNA– leading to gene silencing
Phosphorylation – agonistic
SUMOylation – antagonistic
Mazucotelli et al. (2008) Plant Sci, 174:420
REGULATION
Post-Translational protein modifications (PTMs)
• PTMs result in an increase in complexity from genome to proteome
p.e. 30 000 give rise to 1.8M different protein species (human)
•Involved in protein regulation
•Often reversible, rapid, highly controlled and highly specific
•>300 types of PTMs have been described
•Only affect small percentage of total pool of specific protein
Post-Translational protein modifications (PTMs)
• PTMs result in an increase in complexity from genome to proteome
1. Phosphorylation – reversible attachment of a phosphate group to aa residue (most predominant covalent modification)
• 4 types: N-, S-, acyl-, and O-phosphorylation
• Usually in hydroxyl group of hydroxyamino acids (Ser, Thr,
Tyr)
• Protein kinases/Phosphatases (R-O-PO3)
• Highly dynamic process.
Nakagami et al. (2005) Trend Plant Sci, 10:339
Example of MAPKs
MAPKs are known to be activated by osmotic stresses in Medicago and tobacco;
Arabidopsis: MPK4 and MPK6 are activated by cold, salt, drought, wounding…
MPK3 can be activated by osmotic stress
MEKK1 is transcriptionally induced by salt, drought, cold and wounding
MKK2 is activated by cold and activates MPK4 and MPK6 –overexpression induces cold and salt tolerance and affects expression of 152 genes
Post-Translational protein modifications (PTMs)
• PTMs result in an increase in complexity from genome to proteome
2. S-nitrosylation – addition of NO to a Cys residue
• Role in NO signaling during biotic stress
• Probably as influential as phosphorylation
3. Glycosylation – covalent linkage of an oligosaccharide side chain to a protein
• N-X-S/T
• Starts co-translationally at ER
• Affects proteins involved in stress resistance
Post-Translational protein modifications (PTMs)
• PTMs result in an increase in complexity from genome to proteome
4. Ubiquitination – Covalent addition of Ubiquitin to target proteins
• Often requires phosphorylation
• Polyubiquitination: K48 for proteasome degradation; K63
activity modulation
• Monoubiquitination: leads to changes in localization and
activity
• DUBs are involved in Ubiquitin maturation, recycling and
reversibility.
Polyubiquitination
K63 ubiquitination is involved in
activity modulation
K48 ubiquitination targets proteins for proteasome degradation
UBIQUITINATION pathway
Ubiquitination and Abiotic Stress
• SDIR1 is a positive regulator of ABA signaling
• Up-regulation of SDIR1 enhances ABA-induced stomata closure, inducing drought tolerance
• sdir1-1 has ABA-insensitive phenotype and can be rescued by ABI5, ABF3 and ABF4
1. Regulation of ABA-dependent responses – involvement of RING-type E3 ligases
Ubiquitination and Abiotic Stress
• HOS1 negatively controls cold-stress response
• Promotes degradation of master cold-response regulator: Inducer of CBF Expression 1 (ICE1)
2. Regulation of cold-stress response – involvement of RING-type E3 ligases
HOS1 ICE1Cold-responsive
genes
RUB1: activation and
destabilization of SCF complexes,
transcriptional regulation of p53
HUB1: Pre-mRNA splicing2 targets identified in yeast
ATG8/12: Autophagy, nutrient
recycling in plants
Ub: protein degradation,
localization, protein interaction
Post-translational protein modification by small Ub-like
proteins
UFM1: function unknown
-grasp fold
Diversity in functions for sumoylation
Chromosome segregation
Cell division
DNA replication and repair
Nuclear protein import (RanGAP1)
Protein targeting to
subnuclear structures
Ubiquitination Antagonist
…
Regulation of
inflammatory response
in mammals
Overall, SUMO modifies the activity of proteins; modifies their ability to
interact with other proteins; and modifies their subcellular localization
SUMO
SUMO modification pathway
Novatchkova et al. Planta. 2004;220:1-8
Tempe et al. (2008) Biochem Soc Trans, 36:874
75
150
250
100
50
0 0.5 2 0.5 2 0.5 2 0.5 2 0.5 2 2 4 (h)
control H/S NaCl EthH2O2 Deh
• Several types of abiotic stress
induce sumoylation:
•Heat Shock (37°C);
•50mM H2O2;
•7% ethanol;
•Dehydration
SUMOylation and the response to Abiotic Stress
AP anti-SUMO1
SUMOylation and the response to Drought
SIZ1 (E3 ligase) dependency:
Catala et al. (2007) Plant Cell, 19:2952
SIZ1 has a role in drought tolerance
siz1 is more sensitive to drought
stress.
What about plant size?
Catala et al. (2007) Plant Cell, 19:2952
siz1:
Significantly smaller than WT plants
Deficiencies in leaf elongation and enlargement
siz1 is affected in growth
Smaller epidermal and mesophyll cells
Catala et al. (2007) Plant Cell, 19:2952
SIZ1 has a role in drought tolerance
siz1 is more sensitive to drought
stress.
Catala et al. (2007) Plant Cell, 19:2952
SIZ1 has a role in drought tolerance
siz1 seedlings loose water faster than WT
Catala et al. (2007) Plant Cell, 19:2952
Transcriptome analysis of siz1
Among the ~1700 drought-inducible genes, 262 are positively regulated by SIZ1.
• Not involved in ABA-independent drought response pathways (DREB2A & ERD1)
• Needed for basal expression of ABA-dependent genes involved in drought response
• Role in regulation of anthocyaninbiossintetic pathway
• Role in regulation of of expression of several enzymes involved in hormone biosynthetic pathways (brassinosteroids; jasmonate)
Catala et al. (2007) Plant Cell, 19:2952
Diverse functions of sumoylation
Chromosome segregation
Cell division
DNA replication and repair
Nuclear protein import (RanGAP1)
Protein targeting to
subnuclear structures
Ubiquitination Antagonist
…
…
SUMO
Mediates stress response (SIZ1):
•General Sumoylation (ABA
independent)
•Regulates ABA dependent drought-
stress response
Mazucotelli et al. (2008) Plant Sci, 174:420
UBC9 Example
• Ubiquitination: 10 E1 + 100 E2 + 1000 E3• SUMOylation: 1 E1 + 1 E2 + <10 E3
UBC9
How is regulation done?
Knipscheer et al. (2008) Mol Cell, 31:371
UBC9 Example
Knipscheer et al. (2008) Mol Cell, 31:371
ICE1 Example
• ICE1 is constitutively expressed• Activates CBF genes expression upon cold stress only• Is important for cold tolerance
ICE1 Example
• ICE1 is constitutively expressed• Activates CBF genes expression upon cold stress only
Miura et al. Plant Cell, 19:1403
ICE1 Example
• ICE1 is constitutively expressed• Activates CBF genes expression upon cold stress only
Miura et al. Plant Cell, 19:1403
SUMOylation induces CBF activation by ICE1
ICE1 Example
• ICE1 is constitutively expressed• Activates CBF genes expression upon cold stress only
Miura et al. Plant Cell, 19:1403
SUMOylation induces CBF activation by ICE1Ubiquitination leads to ICE1 degradation
ICE1 Example
• ICE1 is constitutively expressed• Activates CBF genes expression upon cold stress only
Phosphorylation
Miura et al. Plant Cell, 19:1403
SUMOylation induces CBF activation by ICE1Ubiquitination leads to ICE1 degradation
ICE1 Example
• ICE1 is constitutively expressed• Activates CBF genes expression upon cold stress only
Phosphorylation
Miura et al. Plant Cell, 19:1403
SUMOylation induces CBF activation by ICE1Ubiquitination leads to ICE1 degradationPhosphorylation can induce SUMOylation
ICE1 Example
• ICE1 is constitutively expressed• Activates CBF genes expression upon cold stress only
Phosphorylation
Miura et al. Plant Cell, 19:1403
SUMOylation induces CBF activation by ICE1Ubiquitination leads to ICE1 degradationPhosphorylation can induce SUMOylationHOS1 phosphorylation upon cold stress leads to nuclear import (?)
ICE1 Example
• ICE1 is constitutively expressed• Activates CBF genes expression upon cold stress only
Phosphorylation
Miura et al. Plant Cell, 19:1403
Complex interplay between post-translational protein modification in transcriptional control