Plant Reproduction I (cont.)

1. Plant Reproduction I (cont.) Roses are red, Violets are blue, Some poems rhyme. But this one doesn't. - anonymous

2. You are here Today • Spore formation, • Gamete formation, • Pollination, • Fertilization, • Germination.

3. Nelumbo nucifera (Sacred Lotus) 1288 year old viable seeds Dormant Seeds • Dormancy: slow metabolic rate and suspension of growth and development, • seed coat dormancy (seed coat impermeable to O2 and H2O), • release by scarification (fire, abrasion, digestive tracts, etc.), • internal dormancy (physiological), • the most common internal dormancy is a requirement for cold, • release by stratification (cold treatment). Know these terms.

4. Seed Germinationphase changes • Dormant to metabolically active, • etiolated growth (dark growth), • directional response to gravity, • light, • temperature, • touch, • De-etiolation (dark to light habit), • vegetative growth program.

5. Plant Responses to Signals A weed is a plant that has mastered every survival skill except for learning how to grow in rows. - anonymous

6. Home Stretch • Responses to internal and external signals are studied (to a large extent) by studying the genetics and physiology of... Germination (phase change) Etiolated Growth De-etiolation (phase change) Signal Transduction Hormone Responses Photomorphogenesis Gravitropism others...

7. External Signals • External signals are used by plant cells to alter their physiology, morphology and development, • physical environment, • chemical environment, • biological environment, • sometimes other plants, • Plants receive signals at the cell level, and have no well defined sensory organs, • Except for gravity, all other signals are constantly variable.

8. Internal Signals • Signals can be processed by growing and non-growing cells, • Signals are transduced into biologically meaningful results through numerous and co-ordinated pathways, • changes in ion flux, • regulation of metabolic pathways, • regulation of gene expression, • changes in the cytoskeleton.

9. Signal Transductiongeneral General Signal Transduction Fig. 39.2

10. Network Model Signal Transductionmodels Signal Signal Signal Signals Signal Response Response Response Linear Model

11. Reception • Cell surface receptors, • hydrophilic molecules such as peptides and carbohydrates don’t readily cross the membrane, • are perceived on the cell surface, • Amphiphilic and hydrophobic molecules may pass through the membrane to receptors, • steroid hormones for example, • Light may be perceived at the cell surface, or in the cell

12. De-Etiolation (italics) Plant Receptorsi.e. lightresponses • Where does one look? • no clearly differentiated organs (i.e., eyes, ears, etc.), • sensitive tissues, however, no clearly differentiated cells, • lots of responses. • Germination (+/-) • Stem length (-) • Leaf expansion (+) • Flowering (+/-) • Phototropism (+/-) • Stomatal opening (+) • Chloroplast development (+) • Pigment synthesis (+)

13. De-etiolationgreening Etiolated De-etiolated • Etiolated growth habit, • long stem, • unexpanded closed leaves, • etioplasts vs chloroplasts, • lack of chlorophyll, • apical hook, • short root, • Photomorphogenesis, • Inhibited stem growth, • Expanded leaves, • Pigmentation. • Root development. Nature Reviews Molecular Cell Biology 3; 85-93

14. Phytochromeplant photoreceptor • 1920’s, • researchers observed chlorophyll deficient mutants (albinos) that underwent de-etiolation when given physiologically active light, • 1950’s, • phytochrome discovered, • Molecular switch, signal transducer.

15. N C chromophore kinase domains Phytochrome • Proteins with tetra-pyrrole chromophores, and transmission kinase domains, • Phytochrome gene family contains at least five members, • Gene family members serve different functions.

16. cGMP ? Ca 2+ ? Fig. 39-3 Phytochrome Signal Transductionone approach • aurea (tomato) mutants lack a phytochrome gene and are impaired in greening, • micro-inject second messengers at physiologically relavent concentrations, • look for greening, • cGMP and calcium. Response greening Signal red light

17. The Experiments

18. cAMP cAMP is involved in many signal transduction cascades. cGMP is also involved in signal transduction (i.e. cGMP transduces visual signals in rod cells).

19. …inject cGMP. cGMP and Transcriptional Regulation Fig. 39.9

20. Ca2+ channel Protein is opened. Post Translational Modifications Fig. 39.9

21. Microinject Ca 2+ Calcium as a Second Messenger Fig. 39.9

22. Phytochrome is a greening receptor, cGMP is a second messenger, Calcium is a second messenger. Phytochrome Signal Transductionone approach • What do these experiments show? ? • What don’t these experiments show? ? Response greening Signal red light

23. ...response. ...thousands of genes, phytochromes …hundreds of transcription factors, Nature Reviews Molecular Cell Biology 3; 85-93

24. phytochrome (Pfr) …enhances the expression of MYB, Phytochrome Signal Transductionfor real • Active phytochrome (Pfr) is transported to the nucleus, … binds and inactivates transcriptional repressor (PIF3), …MYB transcription factor is expressed, in turn activates CCA1 transcription, … CCA1 enhances the expression of CAB (chlorophyll A/B) proteins. red light activates phytocrome, active phytochrome (Pfr) is transported to the nucleus, CCA1 expression, in turn, enhances greening proteins.

25. Plant Responses to Signals IIPhytohormones No one can look at the plants growing on a bank or on the borders of a thick wood, and doubt that the young stems and leaves place themselves so that the leaves may be well illuminated...they are extremely heliotropic; and this probably serves...as a guide (for) the buried seeds through fissures in the ground or through overlying masses of vegetation, into the light and air. - Charles Darwin “The Power of Movement in Plants” (1880) • Charles Darwin was Plant Physiologist, • Phototropism, • Introduction to the plant hormone Auxin.

26. Phytohormones …a plant product that is able to stimulate physiological responses at very low concentrations, • either in the tissue in which it is synthesised, • or in other regions of the plant to which it is transported, ...do not operate in isolation from one another, but often act in co-ordination to produce subtle responses, …affect gene expression, enzyme activity and membrane function.

27. Tropism: a growth response in plants that results in curvature toward, or away from a stimulus. Phototropism Charles Darwin and his son Francis localized the location of perception for blue light phototropism.

28. Positive Phototropism: growth toward a light stimulus Peter Boysen-Jensen demonstrated that a diffusable substance was involved .

29. Natural auxins... Peter Went demonstrated that the difusable substance resulted in cell expansion. Went isolated the active compound. IAA

30. Oat Coleoptile (2 cm) Oat Coleoptile (> 2 cm) Bioassays • Bioassay: identification (or quantitation) of a biologically active substance by measuring the effect the substance has on living material. auxin

31. Synthesized in the SAM Auxin Transportpolar Auxin moves basipetally (from apical ends to basal ends).

32. Note pH Polar Transpotrequires energy • IAA is in anionic form at cytosolic pH levels, • the plasma membrane is relatively impermeable to IAA- , • IAA- builds up in the cell, • IAA- carrier proteins are differentially expressed at the basal ends of cells, • IAA- exits via efflux carriers once the concentration builds, • IAA- is protonated to become a neutral compound IAAH.

33. IAAH H+ Polar Transpotrequires energy • IAAH enters the next cell passively, through the plasma membrane, • IAAH is also imported by secondary active transport (not shown in the book’s diagram), • Once in the next cell, IAAH returns to IAA- (at the higher cytosolic pH), • H+-ATPases pump protons out of the cell, • driving down the pH outside, • and driving in IAAH in via secondary active transport (down the proton gradient, • IAA- builds up in the cell.

34. IAAH H+ Polar Transpotrequires energy H+-ATPases set up a membrane potential, (positive on the outside), IAA- efflux is facilitated.

35. “Leader” Auxin…induces apical dominance, SAM intact SAM removed basipetal movement of auxin inhibits axillary buds, axillary buds are released.

36. SAM intact SAM removed basipetal movement of auxin inhibits axillary buds, axillary buds are released, Apical Dominance

37. Auxins …induce vascular differentiation hydathode …xylem differentiates between hydathodes and leaf vasculature ...following the path of basipetal auxin transport. …xylem differentiation occurs around the wound, ...following the path of auxin diffusion.

38. Auxins …induces lateral and adventitious root formation, Honeysuckle cut stems Rooting compounds. [ auxin ]

39. Auxins …promotes fruit development, seeds removed seeds removed + auxin seed Normal …embryo produces auxin that stimulates fruit development. Strawberry

40. Fig. 39.7 Auxin…promotes cell expansion, Auxin activates the H+-ATPase. Acidification of the cell wall activates expansin (cell wall proteins).

41. Mode of action (hypothesis) Acid Growth Hypothesis ABP + Auxin …activates the pump. AUXIN BINDING PROTEIN Time course of action.

42. Auxin Binding Proteinauxin receptor? over-express ABP1 gene • Auxin receptor has not conclusively been identified, • a protein that is found to bind auxin, (Auxin Binding Protein, ABP) has been proposed as a candidate receptor, • A common test for receptors, is to over-express (express more than the normal amount of protein), and test to see if the signal is amplified, • ABP passes this test. add auxin

43. wild type abp mutant Auxin Binding Proteinauxin receptor? dead ABP controls cell division and elongation in embryogenesis.

44. Auxins …promote gene expression, 5 - 50 minutes Genes Products

45. Concept Map Receptor Discovery Auxin Function(s) Transport Note: Ch 39, pp 817 - 824…quiz style questions.