Ca2+ signaling in plant
LightCold stressHeat shockMechanical stresses(Touch, wind &Wounding)Pathogen invadingPhytohormones(Auxin, ABA, GA)Gravity
[Ca2+]cyt
AM, FM, DU
DiverseCellularResponses
PrimaryCa2+-SignalDecoder
CalmodulinsCDPKsOther CBPs
Amplitude (AM)
Duration (DU)
Frequency (FM)
Model of Ca2+/calmodulin-mediated network in plants
• Calcineurin is a Ca2+, CaM-dependent protein phosphatase that is highly conserved in eukaryotes from yeast to mammals. Calcineurin is composed of two subunits. - catalytic subunit, calcineurin A (CaM-binding domain)- regulatory subunit, calcineurin B
• CaM and CBL(calcineurin B–like) protein are small proteins that contain multipleCa2+ binding domains but lack other effector domains, such as the kinase domain in CDPKs.CBLs interact with target proteins and regulate their activity. CBLs are encoded by a multigene family of at least 10 members in Arabidopsis.Unlike CaMs, which interact with a large variety of target proteins, CBLs appear to interact with a single family of protein kinases.
• CBL-interacting protein kinase (CIPK) all contain a unique C-terminal region that is both required and sufficient for interaction with the AtCBL-type but not calmodulin-type Ca2+ binding proteins from plants. Interactions between the kinases and AtCBLs require micromolar concentrations of Ca2+,suggesting that increases in cellular Ca2+ concentrations may trigger the formation of AtCBL–kinase complexes in vivo.
Calcineurin, CBL proteins, CIPK
A schematic presentation of calcineurin to CBL–CIPK
• Calcineurin B(CNB)• Calmodulin (CaM)• Calcineurin A (CNA)• Calcineurin B-like protein (CBL)
AtCBL4
AtCBL5
AtCBL9
AtCBL1
AtCBL2
AtCBL3
AtCBL6
AtCBL7
AtCBL8
Phylogenetic tree of AtCBL proteins in Arabidopsis thaliana
Overall structure of the AtCBL2–AtCIPK14 complex
CBL1, CBL2, CBL3, and CBL9 interacted with CIPK6, CIPK16, and CIPK23
Regulatory features of CBL–CIPK interactions
• CBLs interact with CIPKs through the C-terminal non kinase domain, which contains a conserved region among different CIPK members.-hydrophobic interaction with CBL proteins
• Regulatory feature concerns the role of Ca2+ in the CBL–CIPK interaction. Initial analysis of interaction between CBL1 and CIPK1 showed a requirement for micromolar levels of Ca2+.
• Regulatory C-terminal domain of CIPKs include the motif for interacting with a group of type-2C protein phosphatases (PP2Cs).
• Activating CIPK kinase activity, certain structural features of CBLs also suggest that these Ca2+-sensors can change cellular localization of the CBL–CIPK complexes.
Regulation of ion homeostasis by the SOS pathway during salt stress
SOS : Salt overly sensitive
SOS2 : CIPK24
SOS3 : CBL4
Hypothetical model of alternative targeting of CIPK24/SOS2 to plasmamembrane (PM) or tonoplast by CBL4/SOS3 or CBL10.
Stress and ABA Induction of CBL1 Protein Expression Patterns of the CBL9 Gene
Hypothetical Model of CIPK3 Function
Abiotic stress signaling through the CBL-CIPK network
A working model of the Ca2+-dependent pathway for potassium (K)channel activation in low-K response
Signaling pathways that regulate the expression and activities of ion transporters to maintain a low cytoplasmic concentration of Na+ under salt stress
1. Calcineurin B-like proteins (CBLs) represent a unique family of plant calcium sensors that relay signals by interacting with a family of protein kinases (CIPKs).
2. SOS3, a Ca2+ sensor, transduces the signal downstream after activating and interacting with SOS2 protein kinase.
3. This SOS3-SOS2 complex activates the Na+/H+ antiporter activity of SOS1 thereby reestablish cellular ion homeostasis.
SOS : Salt Overly Sensitive gene
SOS 1 : Na+/H+ andtiporter of plasma membrane
SOS 2 : CBL-interacting protein kinases (CIPKs_24)
SOS 3 : Calcineurin B-like protein, (CBL4 _ calcium sensor like calmodulin)
Expression pattern of the CBL10 gene.
F : flower
S: stems
L : rosette leaves
R : roots
SH : shoots
SL : siliques
The cbl10 mutant is hypersensitive to NaCl
175mM NaCl
Mature cbl10 mutant plants are hypersensitive to NaCl
300mM NaCl
NaCl hypersensitivity of the cbl10 mutant is caused by defects in ion homeostasis.
NaCl hypersensitivity of the cbl10 mutant is caused by defects in ion homeostasis
CBL10 interacts with CIPK24
CBL10+
CIPK24GST CIPK CBL10
CBL10 is localized to punctate structures and vacuolar membranes
CBL10 co-localizes with endosomal and tonoplast marker proteins
ARA6CBL10
CBL10 ARA7
CBL10
CBL1
FM4-64
FM4-64
CBL10
CBL1
TPC1
TPC1
ARA6, ARA7 : endosomesFM4-64 : plasma membraneTPC1 : vacuolar membrane
CBL10 interacts with CIPK24 at the tonoplast
CBL10::CIPK24
DAPI/FM4-64
TPC1
CBL1::CIPK24
Hypothetical model of CBL10 function
Summary
1. The CBL protein CBL10 functions as a crucial regulator of salt tolerance in Arabidopsis.
2. Cbl10 mutant plants exhibited significant growth defects and showed hypersensitive cell
death in leaf tissues under high-salt conditions.
3. The Na+ content of the cbl10 mutant was significantly lower than in the wild type under
either normal or high-salt conditions, suggesting that CBL10 mediates a novel Ca2+- signaling pathway for salt tolerance.
4. The CBL10 protein physically interacts with the salt-tolerance factor CIPK24 (SOS2), and
CBL10-CIPK24 (SOS2) complex is associated with the vacuolar compartments that are
responsible for salt storage and detoxification in plant cells.
5. These findings suggest that CBL10 and CIPK24 (SOS2) constitute a novel salt-tolerance
pathway that regulates the sequestration/compartmentalization of Na+ in plant cells.
6. These results identified CIPK24 as a multi-functional protein kinase that regulates different
aspects of salt tolerance by interacting with distinct CBL calcium sensors.