Plant defense responses Hypersensitive response Prepare a 10’ talk for Friday March 3 on plant defense responses or describe interactions between plants& pathogens, pests or symbionts Plant defense responses Hypersensitive response Systemic acquired resistance Innate immunity Phytoalexin synthesis Defensins and other proteins Oxidative burst Some possible pests Nematodes Rootworms Aphids Thrips Gypsy moths hemlock woolly adelgid Some possible pathogens Agrobacterium tumefaciens Agrobacterium rhizogenes Pseudomonas syringeae Pseudomonas aeruginosa Viroids DNA viruses RNA viruses Fungi Oomycetes Some possible symbionts N-fixing bacteria N-fixing cyanobacteria Endomycorrhizae Ectomycorrhizae

Converts light to chemical energy Photosynthesis Converts light to chemical energy 6 CO2 + 6 H2O + light energy <=> C6H12O6 + 6 O2

Photosynthesis 2 sets of rxns in separate parts of chloroplast

PSI works by itself in cyclic photophosphorylation PSI gives excited e- to Fd Fd returns e- to P700+ via PQ, cyt b6/f & PC Each e- pumps H+ at cyt b6/f Use PMF to make ATP

PSI and PSII work together in the “Z-scheme” PSII gives excited e- to ETS ending at NADPH Each e- drives cyt b6/f Use PMF to make ATP PSII replaces e- from H2O forming O2

Physical organization of Z-scheme 2 mobile carriers 1) plastoquinone 2) plastocyanin (PC) 3 protein complexes 1) PSI 2) PSII 3) cytochrome b6/f ATP synthase (CF0-CF1 ATPase) is also visible in E/M

Physical organization of Z-scheme Complexes are arranged asymmetrically! PSII is in appressed regions of grana PSI and ATP synthase in exposed regions cytochrome b6/f, PC and PQ are evenly dispersed why PC and PQ must be mobile why membrane must be very fluid

Chemiosmotic ATP synthesis PMF mainly due to ∆pH is used to make ATP -> very little membrane potential, due to transport of other ions thylakoid lumen pH is < 5 cf stroma pH is 8 pH is made by ETS, cyclic photophosphorylation,water splitting & NADPH synth

Chemiosmotic ATP synthesis Structure of ATP synthase CF1 head: exposed to stroma CF0 base: Integral membrane protein

a & b2 subunits form stator that immobilizes a & b F1 subunits a is also an H+ channel c subunits rotate as H+ pass through g & e also rotate c, g & e form a rotor

Binding Change mechanism of ATP synthesis H+ translocation through ATP synthase alters affinity of active site for ATP

Binding Change mechanism of ATP synthesis H+ translocation through ATP synthase alters affinity of active site for ATP ADP + Pi bind to  subunit then spontaneously form ATP

Binding Change mechanism of ATP synthesis H+ translocation through ATP synthase alters affinity of active site for ATP ADP + Pi bind to  subunit then spontaneously form ATP ∆G for ADP + Pi = ATP is ~0 role of H+ translocation is to force enzyme to release ATP!

Binding Change mechanism of ATP synthesis 1) H+ translocation alters affinity of active site for ATP 2) Each active site ratchets through 3 conformations that have different affinities for ATP, ADP & Pi due to interaction with the subunit

Binding Change mechanism of ATP synthesis 1) H+ translocation alters affinity of active site for ATP 2) Each active site ratchets through 3 conformations that have different affinities for ATP, ADP & Pi 3) ATP is synthesized by rotational catalysis g subunit rotates as H+ pass through Fo, forces each active site to sequentially adopt the 3 conformations

Evidence supporting chemiosmosis 1) ionophores (uncouplers) 2) can synthesize ATP if create ∆pH a) Jagendorf expt: soak cp in pH 4 in dark, make ATP when transfer to pH 8

Evidence supporting chemiosmosis Racker & Stoeckenius (1974) reconstituted bacteriorhodopsin and ATP synthase in liposomes Bacteriorhodopsin uses light to pump H+ make ATP only in the light

Evidence supporting “rotational catalysis” Sambongi et al experiment a) reconstituted ATPase & attached a subunits to a slide b) attached actin filament to c subunit & watched it spin

Regulating Light reactions Regulate partitioning of light energy between PSI and PSII by phosphorylating LHCII complex ordinarily is associated with PSII.

Regulating Light reactions Regulate partitioning of light energy between PSI and PSII by phosphorylating LHCII complex ordinarily is associated with PSII. if PSI falls behind PSII LHCII is kinased

Regulating Light reactions if PSI falls behind PSII LHCII is phosphorylated increased negative charge forces it out of appressed stacks

Regulating Light reactions if PSI falls behind PSII LHCII is phosphorylated increased negative charge forces it out of appressed stacks it then associates with PSI & boosts PSI cyclic activity

Regulating Light reactions if PSI falls behind PSII LHCII is phosphorylated increased negative charge forces it out of appressed stacks it then associates with PSI & boosts PSI cyclic activity sensor is PQ: when highly reduced it indirectly activates a protein kinase that kinases LHCII

Regulating Light reactions sensor is PQ: when highly reduced it indirectly activates a protein kinase that kinases LHCII elevated PQH2 means PSI is falling behind

Regulating Light reactions sensor is PQ: when highly reduced it indirectly activates a protein kinase that kinases LHCII elevated PQH2 means PSI is falling behind Allows plants to adjust relative ATP & NADPH syn

Regulating ATP synthase in dark, ATP synthase could run backwards and consume ATP to make a PMF

Regulating ATP synthase in dark, ATP synthase could run backwards and consume ATP to make a PMF 2 mechanisms prevent this 1) ATP synthase needs a pH gradient to be active

Regulating ATP synthase in dark, ATP synthase could run backwards and consume ATP to make a PMF 2 mechanisms prevent this 1) ATP synthase needs a pH gradient to be active 2) ATP synthase must be reduced by ferredoxin (via thioredoxin) to be active becomes oxidized (therefore inactive) in the dark

Photoinhibition Most plants are saturated at 1/4 full sunlight

Photoinhibition Most plants are saturated at 1/4 full sunlight Excess will damage photosystems

Photoinhibition Most plants are saturated at 1/4 full sunlight Excess will damage photosystems D1 of PSII is fuse: first item to break in high light

Photoinhibition Most plants are saturated at 1/4 full sunlight Excess will damage photosystems D1 of PSII is fuse: first item to break in high light= photoinhibition

Photoinhibition D1 of PSII is fuse: first item to break in high light= photoinhibition Avoidance?

photoinhibition Avoidance? Carotenoids"quench" excited electrons

photoinhibition Avoidance? Carotenoids"quench" excited electrons Dissipate excess energy as heat

photoinhibition Avoidance? Carotenoids"quench" excited electrons Dissipate excess energy as heat Violaxanthin sends exciton to PS

photoinhibition Avoidance? Carotenoids"quench" excited electrons Dissipate excess energy as heat Violaxanthin sends exciton to PS Zeaxanthin dumps energy as heat

photoinhibition Violaxanthin sends exciton to PS Zeaxanthin dumps energy as heat Convert Violaxanthin to Zeaxantin under high light

photoinhibition Violaxanthin sends exciton to PS Zeaxanthin dumps energy as heat Convert Violaxanthin to Zeaxantin under high light Use NADPH to revert Zeaxanthin to Violanthin in low light

photoinhibition Violaxanthin sends exciton to PS Zeaxanthin dumps energy as heat Convert Violaxanthin to Zeaxantin under high light Revert Zeaxanthin to Violanthin in low light Other mechs : paraheliotropism, anthocyanins

Light-independent (dark) reactions The Calvin cycle

Light-independent (dark) reactions occur in the stroma of the chloroplast (pH 8) Consumes ATP & NADPH from light reactions regenerates ADP, Pi and NADP+

Light-independent (dark) reactions Overall Reaction: 3 CO2 + 3 RuBP + 9 ATP + 6 NADPH = 3 RuBP + 9 ADP + 9 Pi + 6 NADP+ + 1 Glyceraldehyde 3-P

Light-independent (dark) reactions 1) fixing CO2 2) reversing glycolysis 3) regenerating RuBP

fixing CO2 1) RuBP binds CO2

fixing CO2 RuBP binds CO2 2) rapidly splits into two 3-Phosphoglycerate therefore called C3 photosynthesis

fixing CO2 1) CO2 is bound to RuBP 2) rapidly splits into two 3-Phosphoglycerate therefore called C3 photosynthesis detected by immediately killing cells fed 14CO2

fixing CO2 1) CO2 is bound to RuBP 2) rapidly splits into two 3-Phosphoglycerate 3) catalyzed by Rubisco (ribulose 1,5 bisphosphate carboxylase/oxygenase) the most important & abundant protein on earth

fixing CO2 1) CO2 is bound to RuBP 2) rapidly splits into two 3-Phosphoglycerate 3) catalyzed by Rubisco (ribulose 1,5 bisphosphate carboxylase/oxygenase) the most important & abundant protein on earth Lousy Km

fixing CO2 1) CO2 is bound to RuBP 2) rapidly splits into two 3-Phosphoglycerate 3) catalyzed by Rubisco (ribulose 1,5 bisphosphate carboxylase/oxygenase) the most important & abundant protein on earth Lousy Km Rotten Vmax!

Reversing glycolysis converts 3-Phosphoglycerate to G3P consumes 1 ATP & 1 NADPH

Reversing glycolysis G3P has 2 possible fates 1) 1 in 6 becomes (CH2O)n

Reversing glycolysis G3P has 2 possible fates 1) 1 in 6 becomes (CH2O)n 2) 5 in 6 regenerate RuBP