Course PlanWe will study effects of soil and stresses on plant secondary products and see where it leads us1.Learn more about plant secondary products•Why do they make them?•When do they make them?•Where do they make them?

Pick some plants to study•Capsicum (Chiles) = capsaicin•Nicotiana (Tobacco) = nicotine•Onions (Allium cepa)?= syn-Propanethial-S-oxide•Garlic (Allium sativum)?= Alliin•Radishes (Raphanus sativus) = glucosinolates•Mustard (Brassica juncea) = glucosinolates•Basil (Ocimum basilicum) = eugenol (and others)•Cilantro (Coriandrum sativum) = lots of candidates!•Peppermint (Mentha × piperita) = menthol + many•Catnip (Nepeta cataria) = nepetalactone•Dill = dillapiole•Purslane (Portulaca oleracea)= omega-3 fatty acids•Brassica rapa = glucosinolates

Plant StressWon Senator Proxmire’s “Golden Fleece” award for wasteful government spending1.Water?2.Nutrients?3.Environment?• Temp?• Pollution?• Ozone, other gases?• Herbicides, eg Round-Up, Atrazine?

• Insects and other herbivores? • Pathogens = bacteria, viruses, fungi

Plant StressNext assignment: presenting a plant stressor, what is known about it, and why it might affect plant 2˚ compounds in an ~ 10 minute presentation?

Alternative: presenting another good plant/stressor response to study and why we should choose it over the ones already presented.

Mineral NutritionMacronutrients: CHOPKNSCaFeMgBNaCl

• Ca: signaling, middle lamella, cofactor• Fe: cofactor• Mg: cofactor• mobile in plant, so shows first in old leaves

Mineral NutritionMicronutrients: BNaCl others include Cu, Zn, Mn

• B: cell elongation. NA metabolism• Na: PEP regeneration, K substitute• Cl: water-splitting, osmotic balance• Cu: cofactor• immobile in plant, so shows first in young leaves

Plant food• 2.6% ammoniacal nitrogen • 4.4% nitrate nitrogen• 9% phosphorus• 5% potassium• calcium (2%)• magnesium (0.5%)• sulfur (0.05%)• boron (0.02%)• chlorine (0.1%)• cobalt (0.0015%)• copper (0.05%)• iron (0.1%)• manganese (0.05%)• molybdenum (0.0009%)• nickel (0.0001%)• sodium (0.10%) • zinc (0.05%)

Mineral NutritionSoil nutrients•Amounts & availability varyPSU extension Text

Mineral NutritionSoil nutrients•Amounts & availability vary•Many are immobile, eg P, Fe

Mineral NutritionSoil nutrients• Amounts & availability vary• Many are immobile, eg P, Fe• Mobile nutrients come with soil H2O

Mineral NutritionSoil nutrients• Amounts & availability vary• Many are immobile, eg P, Fe• Mobile nutrients come with soil H2O

• Immobiles must be “mined”• Root hairs get close

Mineral NutritionImmobile nutrients must be “mined”

• Root hairs get close• Mycorrhizae get closer

Mineral NutritionImmobile nutrients must be mined

• Root hairs get close• Mycorrhizae get closer

Solubility varies w pH

Mineral Nutrition•Solubility varies w pH•5.5 is best compromise

Mineral NutritionSolubility varies w pH•5.5 is best compromise•Plants alter pH @ roots to

aid uptake

Mineral NutritionNutrients in soil•Plants alter pH @ roots to aid uptake• Also use symbionts

• Mycorrhizal fungi help: especially with P

Mineral NutritionAlso use symbionts•Mycorrhizal fungi help: especially with P• P travels poorly: fungal hyphae are longer & thinner

Mineral NutritionAlso use symbionts•Mycorrhizal fungi help: especially with P• P travels poorly: fungal hyphae are longer & thinner

• Fungi give plants nutrients

Mineral NutritionAlso use symbionts•Mycorrhizal fungi help: especially with P• P travels poorly: fungal hyphae are longer & thinner

• Fungi give plants nutrients• Plants feed them sugar

Mineral NutritionAlso use symbionts•Mycorrhizal fungi help: especially with P• P travels poorly: fungal hyphae are longer & thinner

• Fungi give plants nutrients• Plants feed them sugar

• Ectomycorrhizae surround root: only trees, esp. conifers

Mineral NutritionEctomycorrhizae surround root: only trees, esp. conifers• release nutrients into apoplast to be taken up by roots

Mineral NutritionEctomycorrhizae surround root: trees•release nutrients into apoplast to be taken up by rootsEndomycorrhizae invade root cells: Vesicular/Arbuscular

• Most angiosperms, especially in nutrient-poor soils

Mineral NutritionEndomycorrhizae invade root cells: Vesicular/Arbuscular

• Most angiosperms, especially in nutrient-poor soils• May deliver nutrients into symplast

RhizosphereEndomycorrhizae invade root cells: Vesicular/Arbuscular

• Most angiosperms, especially in nutrient-poor soils• May deliver nutrients into symplast• Or may release them when arbuscule dies

RhizosphereEndomycorrhizae invade root cells: Vesicular/Arbuscular

• Most angiosperms, especially in nutrient-poor soils• Deliver nutrients into symplast or release them

when arbuscule diesAlso find bacteria, actinomycetes, protozoa associated with root surface = rhizosphere

RhizosphereAlso find bacteria, actinomycetes, protozoa associated with root surface = rhizosphere• Plants feed them lots of C!

RhizosphereAlso find bacteria, actinomycetes, protozoa associated with root surface = rhizosphere• Plants feed them lots of C!• They help make nutrients available

RhizosphereAlso find bacteria, actinomycetes, protozoa associated with root surface = rhizosphere• Plants feed them lots of C!• They help make nutrients available• N-fixing bacteria supply N to many plant spp

N assimilation by N fixersExclusively performed by prokaryotesDramatically improve the growth of many plants

N assimilation by N fixersExclusively done by prokaryotesMost are free-living in soil or water

N assimilation by N fixersExclusively done by prokaryotesMost are free-living in soil or waterSome form symbioses with plants

N assimilation by N fixersExclusively done by prokaryotesMost are free-living in soil or waterSome form symbioses with plantsLegumes are best-known, butmany others including mosses,ferns, lichens

N assimilation by N fixersExclusively done by prokaryotesMost are free-living in soil or waterSome form symbioses with plantsLegumes are best-known, butmany others including mosses,ferns, lichensAlso have associations where N-fixers form films on leaves orroots and are fed by plant

N assimilation by N fixersExclusively done by prokaryotesAlso have associations where N-fixers form films on leaves orroots and are fed by plantAll must form O2-free environment for nitrogenase

N assimilation by N fixersAll must form O2-free environment for nitrogenaseO2 binds & inactivates electron-transfer sites

N assimilation by N fixersO2 binds & inactivates electron-transfer sitesHeterocysts lack PSII, haveother mechs to lower O2

N assimilation by N fixersHeterocysts lack PSII, haveother mechs to lower O2 Nodules have specialstructure + leghemoglobinto protect from O2

Nodule formationNodules have special structure + leghemoglobin to protect

from O2 Bacteria induce the plant to form nodules

Nodule formationBacteria induce the plant to form nodules1. Root hairs secrete chemicals that attract N-fixers

Nodule formationBacteria induce the plant to form nodules1. Root hairs secrete chemicals that attract N-fixers2. Bacteria secrete Nod factors that induce root hair to

coil up. Nod factors determine species-specificity

Nodule formation1. Root hairs secrete chemicals that attract N-fixers2. Bacteria secrete Nod factors that induce root hair to

coil up. Nod factors determine species-specificity3. Nod factors induce degradation of root cell wall

Nodule formation3. Nod factors induce degradation of root cell wall4. Plant forms "infection thread"=internal protusion of

plasma membrane that grows into cell5. When reaches end of cell bacteria are released into

apoplast and repeat the process on inner cells

Nodule formation5. When reaches end of cell bacteria are released into

apoplast and repeat the process on inner cells6. Cortical cells near xylem form a nodule primordium

Nodule formation5. When reaches end of cell bacteria are released into

apoplast and repeat the process on inner cells6. Cortical cells near xylem form a nodule primordium7. When bacteria reach these cells the infection thread

breaks off, forming vesicles with bacteria inside

Nodule formation7. When bacteria reach these cells the infection thread

breaks off, forming vesicles with bacteria inside8. Vesicles fuse, form the peribacteroid membrane and

bacteria differentiate into bacteroids.

Nodule formation8. Vesicles fuse, form the peribacteroid membrane and

bacteria differentiate into bacteroids.9. Plant cells differentiate into nodules

Nodule formation8. Vesicles fuse, form the peribacteroid membrane and

bacteria differentiate into bacteroids.9. Plant cells differentiate into nodules: have layer of cells

to exclude O2 & vasculatureto exchange nutrients

Nodule formationPlant cells differentiate into nodules: have layer of cells to

exclude O2 & vasculature to exchange nutrientsComplex process that is difficult to engineer: 21 non-

legume plant genera have N-fixers

Nitrogen fixationN2 + 8H+ + 8e− + 16 ATP → 2NH3 + H2 + 16ADP + 16 Pi

Catalysed by nitrogenase, a very complex enzyme!

Nitrogen fixationN2 + 8H+ + 8e− + 16 ATP → 2NH3 + H2 + 16ADP + 16 Pi

Catalysed by nitrogenase, a very complex enzyme!Also catalyzes many other reactionsUsually assayed by acetylene reduction

Nitrogen fixationN2 + 8H+ + 8e− + 16 ATP → 2NH3 + H2 + 16ADP + 16 Pi

Usually assayed by acetylene reductionSequentially adds 2 H per cycle until reach NH3

Nitrogen fixationSequentially adds 2 H per cycle until reach NH3

May then be exported to cytosol & assimilated by GS/GOGAT or assimilated inside bacteroid

Nitrogen fixationSequentially adds 2 H per cycle until reach NH3

May then be exported to cytosol & assimilated by GS/GOGAT or assimilated inside bacteroidAre then converted to amides or ureides& exported to rest of plant in the xylem!

Nutrient uptakeMost nutrients are dissolved in water

Nutrient uptakeMost nutrients are dissolved in water

• Enter root through apoplast until hit endodermis

Nutrient uptakeMost nutrients are dissolved in water

• Enter root through apoplast until hit endodermis• Then must cross plasma membrane

Crossing membranesA) Diffusion through bilayer B) Difusion through protein poreC) Facilitated diffusionD) Active transportE) Bulk transport

1) Exocytosis2) Endocytosis

Selective

Active

Nutrient uptakeThen must cross plasma membrane

• Gases, small uncharged & non-polar molecules diffuse

Nutrient uptakeThen must cross plasma membrane

• Gases, small uncharged & non-polar molecules diffusedown their ∆ [ ]• Important for CO2, auxin & NH3 transport

Nutrient uptakeThen must cross plasma membrane

• Gases, small uncharged & non-polar molecules diffusedown their ∆ [ ]• Polar chems must go through proteins!

Selective Transport

1) Channels

integral membrane proteins with pore that specific ions diffuse through

Selective Transport

1) Channels

integral membrane proteins with pore that specific ions diffuse through• depends on size & charge

Channelsintegral membrane proteins with pore that specific ions diffuse through• depends on size & charge• O in selectivity filter bindion (replace H2O)

Channelsintegral membrane proteins with pore that specific ions diffuse through• depends on size & charge• O in selectivity filter bindion (replace H2O)• only right one fits

ChannelsO in selectivity filter bindion (replace H2O)• only right one fits• driving force?electrochemical

Channelsdriving force : electrochemical “non-saturable”

Channelsdriving force : electrochemical “non-saturable”regulate by opening & closing

Channelsregulate by opening & closingligand-gated channels open/close when bind specific chemicals

Channelsligand-gated channels open/close when bind specific chemicalsStress-activated channels open/close in response to mechanical stimulation

ChannelsStress-activated channels open/close in response to mechanical stimulationvoltage-gated channels open/close in response to changes in electrical potential

Channels• Old model: S4 slides up/down• Paddle model: S4 rotates

Channels• Old model: S4 slides up/down• Paddle model: S4 rotates• 3 states

1.Closed2.Open3. Inactivated

Selective Transport1) Channels2) Facilitated Diffusion (carriers)

Carrier binds molecule

Selective TransportFacilitated Diffusion (carriers)

Carrier binds moleculecarries it through membrane& releases it inside

Selective TransportFacilitated Diffusion (carriers)

Carrier binds moleculecarries it through membrane& releases it insidedriving force = ∆ [ ]

Selective TransportFacilitated Diffusion (carriers)

Carrier binds moleculecarries it through membrane& releases it insidedriving force = ∆ [ ]Important for sugar transport

Selective TransportFacilitated Diffusion (carriers)Characteristics

1) saturable 2) specific 3) passive: transports down ∆ []

Selective Transport1) Channels2) Facilitated Diffusion (carriers)Passive transport should equalize [ ]Nothing in a plant cell is at equilibrium!

Selective TransportPassive transport should equalize [ ]Nothing in a plant cell is at equilibrium!Solution: use energy to transport specific ions against their ∆ [ ]

Active TransportIntegral membrane proteins • use energy to transport specific ions against their ∆ [ ]• allow cells to concentrate some chemicals, exclude others

Active TransportCharacteristics1) saturable

102-104 molecules/s105-106 ions/s

Active TransportCharacteristics1) saturable2) specific

Active TransportCharacteristics1) saturable2) specific

3) active: transport up ∆ [ ] (or ∆ Em)

4 classes of Active transport ATPase proteins1) P-type ATPases (P = “phosphorylation”)

• Na/K pump• Ca pump in ER & PM• H+ pump in PM

• pumps H+ out of cell

4 classes of Active transport ATPase proteins1) P-type ATPases (P = “phosphorylation”)2) V-type ATPases (V = “vacuole”)

• H+ pump in vacuoles

4 classes of Active transport ATPase proteins1) P-type ATPases (P = “phosphorylation”)2) V-type ATPases (V=“vacuole”)3) F-type ATPases (F = “factor”) a.k.a. ATP synthases

• mitochondrial ATP synthase• chloroplast ATP synthase

4 classes of Active transport ATPase proteins1) P-type ATPases (P = “phosphorylation”)2) V-type ATPases (V = “vacuole”)3) F-type ATPases (F = “factor”)4) ABC ATPases (ABC = “ATP Binding Cassette”)

• multidrug resistance proteins

4 classes of Active transport ATPase proteins1) P-type ATPases (P = “phosphorylation”)2) V-type ATPases (V = “vacuole”)3) F-type ATPases (F = “factor”)4) ABC ATPases (ABC = “ATP Binding Cassette”)

• multidrug resistance proteinspump hydrophobic drugs out of cellsvery broad specificity

Secondary active transportUses ∆ [ ] created by active transport to pump something else across a membrane against its ∆ [ ]

Secondary active transportUses ∆ [ ] created by active transport to pump something else across a membrane against its ∆ [ ] Symport: both substances pumped same way

Secondary active transportUses ∆ [ ] created by active transport to pump something else across a membrane against its ∆ [ ] Symport: both substances pumped same wayAntiport: substances pumped opposite ways

Secondary active transportUses ∆ [ ] created by active transport to pump something else across a membrane against its ∆ [ ] Symport: both substances pumped same wayAntiport: substances pumped opposite ways

Nutrient uptakeGases enter/exit by diffusion down their ∆ [ ]

Nutrient uptakeGases enter/exit by diffusion down their ∆ [ ]Ions vary dramatically!

Nutrient uptakeGases enter/exit by diffusion down their ∆ [ ]Ions vary dramatically!H+ is actively pumped out of cell by P-type H+ -ATPase

Nutrient uptakeGases enter/exit by diffusion down their ∆ [ ]Ions vary dramatically!H+ is actively pumped out of cell by P-type H+ -ATPaseand into vacuole by V-type!

Nutrient uptakeH+ is actively pumped out of cell by P-type H+ -ATPaseand into vacuole by V-type!• Main way plants make membrane potential (∆Em)!

Nutrient uptakeH+ is actively pumped out of cell by P-type H+ -ATPaseand into vacuole by V-type!• Main way plants make membrane potential (∆Em)!• K+ diffuses through channels down ∆Em

Nutrient uptake• K+ diffuses through channels down ∆Em• Also taken up by transporters

Nutrient uptake• K+ diffuses through channels down ∆Em• Also taken up by transporters

• some also transport Na+

Nutrient uptake• K+ diffuses through channels down ∆Em• Also taken up by transporters

• some also transport Na+

• why Na+ slows K+ uptake?

Nutrient uptake• K+ diffuses through channels down ∆Em• Also taken up by transporters

• some also transport Na+

• why Na+ slows K+ uptake?

Na+ is also expelled by H+ antiport

Nutrient uptake• K+ diffuses through channels down ∆Em• Also taken up by transporters

• some also transport Na+

• why Na+ slows K+ uptake?

Na+ is also expelled by H+ antiport•Enters through channels

Nutrient uptakeNa+ is also expelled by H+ antiport•Enters through channelsCa2+ is expelled by P-typeATPases in PM

Nutrient uptakeNa+ is also expelled by H+ antiport•Enters through channelsCa2+ is expelled by P-typeATPases in PM& pumped into vacuole by H+ antiport

Nutrient uptakeNa+ is also expelled by H+ antiport•Enters through channelsCa2+ is expelled by P-typeATPases in PM& pumped into vacuole by H+ antiport• enters cytosol via channelsPO4, SO4, Cl & NO3 enter by H+ symport

Nutrient uptakePO4, SO4, Cl & NO3 enter by H+ symport• also have anion channels of ABC type