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ATP. Ca 2+. ATP. ADP+P i. H +. Ca 2+. Ca 2+. Ca 2+. ADP+P i. Ca 2+ /H+ antiporter. ATP. ATP. Ca 2+. Ca 2+. Ca 2+. Ca 2+. Ca 2+. Ca 2+ channel. Ca 2+ channel. ADP+P i. ADP+P i. 6. Ca 2+ -ATPases , another group of P-type ATPase, are
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ATP Ca2+ ATP ADP+Pi H+ Ca2+ Ca2+ Ca2+ ADP+Pi Ca2+ /H+ antiporter ATP ATP Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ channel Ca2+ channel ADP+Pi ADP+Pi 6. Ca2+-ATPases, another group of P-type ATPase, are distributed among various plant membranes. Ca2+ channel Vacuole Golgi PM Ca2+ channel Nucleus ER
1 4 5 10 1 4 5 10 COOH COOH Ca2+ Ca2+ Auto- inhibitor NH2 Asp-P Asp-P CaM-Binding NH2 ATP-binding ATP-binding Structural Characteristic of Ca2+-ATPases ER-type Calcium ATPases (ECAs) Autoinhibited Calcium ATPases (ACAs)
Autoinhibited Ca2+- ATPase (ACA) ER group ACA7 ACA2 ACA1 ACA11 Vacuole group ACA4 ACA10 ACA8 ACA9 ACA13 ACA12 PM group Unknown group
Localization of ACA11 in Arabidopsis Protoplast GFP RFP DIC Merge Free GFP (Cytosol) 35S-ACA8-GFP (PM) 35S-ACA11-GFP (Vacuole)
Phenotype of vacuole Ca2+ pump mutants (aca4/aca11)
Cell death in aca4/aca11 is a HR-like PCD Lactophenol cleaning (Phenolic compound) Anilline blue staining (callose) WT KO Bright Bright UV UV
EM Photos of aca4/aca11 mutants V V V W.T W.T x60 V V V #316 #316 #316
Recovery phenotype of aca11/aca4 mutant by phosphate Transfer to soil after growth in MSO media for 10 days
Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Pi Pi Pi Pi Pi Pi Pi Pi Pi Pi Pi Pi Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Ca2+ Hypothesis chloroplast chloroplast Nucleus ER Golgi Cell Death Signal??? VPE activation chloroplast membrane collapse Plasma membrane Cell Death
7. Vacuolar and other membranes are energized through vacuolar H+-ATPase V1 sector pH 7.5 V0 sector pH 5.5 (pH 3.0) Structural model of V-type H+-ATPase
Function of V-type H+-ATPases • Plant vacuole contains a highly acidic solution, with pH 5.5. • Proton pumping into the vacuolar lumen not only energizes the membrane for carrier-mediated transport but also generates the low pH of the vacuole. • An acidic lumemal pH is thought to contribute to • vesicle trafficking And protein targeting. • These enzymes are present in membranes from th ER, Golgi, • and coated vesicles of plant cells
The H+-ATPase are potently and specifically inhibited by the macrolide antibiotic bafilomycin A1
8. The plant vacuolar membrane also possesses a unique H+-pumping inorganic pyrophosphatase (H+-ppase) PPi 2 pi
9. ABC-type pumps are emerging as major players in sequestration of amphipathic metabolites and xenotoxics into the vacuole ABC: ATP binding cassette
Transport of a glutathione-conjugated xenotoxic and a chlorophyll catabolite by AtMRP2, and ABC transporter from Arabidpsis.
Cd2+detoxificationpathway in yeast and plant cytoplasm Glu+Cys GCS γ-GluCys +Gly Cd2+ GS Cd(GSH)2 GSH sulfide PCS YCF1 PC sulfide Cd2+ LMW LMW HMW Cd- PC Cd- PC Cd- PC HMT1 vacuole Christopher S.C. et al(2000)