1. Functional biology of and limitations to Photosynthesis Lecture 17 03/29/05

2. Last Time Biomechanics and allometry Biomechanics Euler buckling equation How the Euler equation has constrained and guided plant evolution Different ways to be a tree Allometry Allometric growth and biomass partitioning

3. Today Photosynthesis Part II -Carbon reactions -What regulates photosynthesis (carbon reactions) -Controls over CO 2 diffusion -Light reactions review -Carbon fixation limits -Biochemical control over carbon fixation -Triose-P transporter -A/C i Curves

4. Central Question What have been the important constraints which have shaped the evolution of plant form and function? Last Lecture (2nd lecture in class) –Evolution of Photoautotrophy –Basic form of the photosynthetic apparatus This Lecture –Controls over photosynthesis - evolutionary implications / important variation

5. www.tiscali.co.uk/.../ hutchinson/m0030839.html users.rcn.com/.../ BiologyPages/L/Leaf.html

6. A group of palisade mesophyll cells in a spinach leaf, in confocal stereo. The numerous mitochondria (light green) lie between the chloroplasts (light green) (which are only very dimly fluorescent under the optical conditions used here). Some are round particles, others long branched filaments. Some are slightly blurred because the cells were still alive and the mitochondria were moving while the image was being recorded. Chloroplasts are light green and essentially fill most of the cell. Fluorescently vital stained with Rhodamine-123. Palisade cells are packed with chlorophyll and mitochondria

7. www.kensbiorefs.com/ cellstructure.html Where do these reactions take place?? http://www.daviddarling.info/images/chloroplast.jpg

8. Membrane Harvest Light 3 Major Functions of Light Reactions

9. Where does the ATP and NADPH go from the light reactions? The Calvin cycle (PCR cycle) Turns out there is plenty of light energy, most of the time, what regulates photosynthesis is carboxylation! Calvin Cycle or Photosynthetic Carbon Reduction (PCR Cycle)

10. Overview - Photosynthetic Apparatus Light harvesting reactions Resources: Light (photons) and Water (electrons) Products: ATP (energy), H+ (potential energy) and NADPH (reducing power) Carbon reactions

11. Carbon fixation reactions (C3 Photosynthesis) Use ATP and NADPH from light harvesting reactions to reduce CO 2 to sugars

12. Chloroplast inner membranes Chloroplast Initial product is A 3 carbon sugar (PGA) Phosphoglyceric acid Carbon fixation catalyzed by enzyme Ribulose-bisphosphate carboxylase-oxygenase Rubisco

13. Structure of Rubisco showing its four-fold symmetry. From the University of Hamburg site. http://www.biologie.uni-hamburg.de/b-online/fo24_1/e1rxoe.htm

14. Rate of Rubisco mediated PCR cycle is mediated by Light concentration Starch is a chain of sugars

15. All reactions are enzyme mediated [A] + [B] C + D [enzyme] Substrates Catalyst Products Rate of production of A* will depend upon many things -Most importantly on concentrations of players If [enzyme] is low then reaction is enzyme (binding site) limited If [A] and or [B] is low then reaction is substrate limited -Efficiency of enzyme

16. Notable Features of Carbon Fixation Reactions (i) Large nitrogen requirement for Rubisco and other photosynthetic enzymes (Nitrogen limitation) (ii) Dependence on the products of the light-harvesting reactions (which depends on amount of light hitting leaf) (iii) Dependent upon CO 2 supply to the chloroplast What influences CO 2 supply? Ability of CO 2 to reach reactions...

17. Limits to Photosynthesis Consider one important limit - diffusion of CO 2 into chloroplasts from atmosphere How can we make sense of CO 2 limitation of photosynthesis?

18. Fick’s Law of Diffusion In order to exchange ‘resources’ with the environment plants must follow diffusion laws Flow of certain resource per unit area per unit time Diffusion coefficient (varies with temp and concentration) Concentration Gradient (change in resource concentration, c j with distance, x)

19. Resistance Pathway Plants regulate CO 2 uptake and water loss by changing the size of stomatal opening (which regulates Stomatal conductance - the flux of water vapor or CO 2 per unit driving force (a given concentration gradient)

20. Elaborating Fick’s Law - Anatomy of the System (since we identify CO 2, we remove the diffusivity term – but it will return when we begin to consider water versus CO 2 ) Conversion – Fick’s Law J CO 2 = D x /  r J CO 2 is the flux of CO 2 D x is the concentration gradient  r is the sum of all resistances to the flux of CO 2 along that path

21. Elaborating Fick’s Law - Concentration gradient C a is the external CO 2 concentration (external to the leaf) C i is the concentration at the site of carboxylation (we call this “intercellular CO 2 concentration)

22. Resistances to CO 2 diffusion 1.Boundary layer air-phase resistance 2.Stomatal resistance (=1/conductance) 3.Internal air spaces aqueous-phase resistance 4.Diffusional resistance across mesophyll cell wall 5.Resistance at the site of carboxylation (chloroplast resistance Resistance Pathway Should a plant always minimize resistance to CO 2 diffusion?

23. When plants reduce stomatal conductance (or Increase resistance) water is conserved BUT photosynthesis declines. Reduce the efficiency with which plants converts light energy to carbohydrates Remember.... Also, when water stress is too great stomates will close

24. (i) Large nitrogen requirement for rubisco and other photosynthetic enzymes (Nitrogen limitation) (ii) Dependence on the products of the light-harvesting reactions (which depends on amount of light hitting leaf) (iii) CO 2 supply to the chloroplast (iv) Phosphate Translocator – mediates the ‘supply’ and ‘demand’ for triose-P in the cell –“Source-Sink” theory and carbohydrate backup Notable Features of Carbon Fixation Reactions

25. Summary - Biochemical regulation of photosynthesis Light Reactions – mediate inputs of energy Rubisco (Ribulose bisphosphate carboxylase / oxygenase) – mediates inputs of carbon –Extensive research has shown that Rubsico controls the reaction rates of the whole Calvin cycle reaction complex Phosphate Translocator – mediates the ‘supply’ and ‘demand’ for triose-P in the cell –“Source-Sink” theory and carbohydrate backup

26. Chloroplast Production of sucrose Triose Phosphate/ Inorganic Phosphate Translocator (TPT)

27. Transporters - exchange between Chloroplast and Cytosol Outer membrane of chloroplast has pore-forming proteins (porins) - allow substances to diffuse freely Inner chloroplast membrane is the permeability barrier - transport is usually by specific translocators Rate of transport depends upon demand for Sucrose

28. Take-home from the P i regulation The translocator is ultimately controlled by sink strength of the plant (the ability of plant’s sinks to utilize sucrose) If sucrose builds up in the leaf due to lack of sink strength, it negatively feeds back on reactions in the cytosol (which in turn slows down the generation of P i ) P i must be available to exchange with triose-P out of the chloroplast. Lack of triose-P transport results in triose-P converted into starch in the chloroplasts. –(slows RuBP regeneration)

30. Sharkey Table – limitations of photosynthesis Rubisco activity limits photosynthesis under high light, low C i conditions, so there is more RuBP (ribulose 1,5 bisphosphate) than binding sites on Rubisco. RuBP regeneration limits photosynthesis under low light and high CO 2, so there are open binding sites on Rubisco because electron transport capacity is inadequate to regenerate enough RuBP Triose-P utilization limits photosynthesis under high light and high CO 2, also resulting in more RuBP than available Rubisco binding sites.

31. How do we make sense of the Limits to Photosynthesis? The A/C i Curve A = CO 2 Assimilation Rate C i = Internal [CO 2 ]

32. Internal CO 2 concentration (  mol mol -1 ) Assimilation rate (  mol m -2 s -1 )

33. Internal CO 2 concentration (  mol mol -1 ) Demand Function Assimilation rate (  mol m -2 s -1 ) RuBP saturated region CaCa

34. What sets this intercept? *key feature that has driven evolutionary diversification in land plants!

35. Internal CO 2 concentration (  mol mol -1 ) Demand Function Assimilation rate (  mol m -2 s -1 ) RuBP saturated region CaCa

36. Internal CO 2 concentration (  mol mol -1 ) Demand Function Assimilation rate (  mol m -2 s -1 ) RuBP saturated region CaCa ?

37. Internal CO 2 concentration (mmol mol -1 ) Demand Function Assimilation rate (mmol m -2 s -1 ) RuBP saturated region CaCa

38. Internal CO 2 concentration (mmol mol -1 ) Demand Function Assimilation rate (mmol m -2 s -1 ) Supply Function RuBP saturated region CiCi Supply function Rate at which CO 2 is supplied to rubisco sites and is determined by [CO 2 ] in atmosphere and Stomatal conductance (1/resistance) Plant Example #1 CaCa Rates of RuBP regeneration limiting to photosynthesis

39. Internal CO 2 concentration (mmol mol -1 ) Assimilation rate (mmol m -2 s -1 ) Supply Function CaCa CiCi Demand Function How does plant #2 differ from plant #1? Plant Example #2

40. Controls over photosynthesis Supply function – controlled by stomatal responses to: What Controls Demand and Supply Functions?

41. Controls over photosynthesis Demand function – controlled by:

42. Questions