Plant Responses to Signals: Stimuli and Signal Transduction Pathways

1. Chapter 39 Plant Responses to Internal and External Signals

2. Overview: Stimuli and a Stationary Life • Linnaeus noted flowers of different species opened at different times of day/could be used by horologium florae, or floral clock • Reflects time insect pollinators active, one of environmental factors plant senses to compete successfully • Plant’s morphology/physiology tuned to surrounding by complex interactions between environmental stimuli/internal signals • Plants, being rooted to ground, must respond to environmental cues by adjusting individual pattern of growth/development • Must detect changes first • Bending of seedling toward light begins with sensing direction, quantity, and color of light

3. Concept 39.1: Signal transduction pathways link signal reception to response • All organisms received specific signals/respond to them in ways that enhance survival/reproductive success • Plants have cellular receptors that detect changes in their environment (molecule affected by stimulus) • For stimulus to elicit response, certain cells must have appropriate receptor • Stimulation of receptor initiates specific signal transduction pathway

4. Fig. 39-2 (b) After a week’s exposure to natural daylight Begins to resemble typical plant w/broad green leaves, short sturdy stems, long roots (transformation begins w/reception of light by specific pigment, phytochrome) by undergoing changes (de-etiolation) by reception of signal (light) which is transduced into responses (greening) Before exposure to light Tall, spindly stem/nonexpanded leaves (morphological adaptations called etiolation enable shoots to penetrate soil, including short roots due to little need for water absorption from little water loss by shoots) Expanded leaves hindrance as shoots push through soil/chlorophyll waste of energy (underground)

5. Fig. 39-3 Review of a general model for signal transduction pathways CYTOPLASM CELL WALL Transduction Response 1 2 3 Reception Relay proteins and Activation of cellular responses second messengers Receptor Hormone or environmental stimulus Plasma membrane

6. Reception/Transduction • Internal/external signals are detected by receptors, proteins that change in response to specific stimuli • Phytochrome functioning in de-etiolation located in cytoplasm, not built into plasma membrane as most receptors • Second messengers (small molecules/ions in cell transfer and amplify signals from receptors to proteins that cause responses) activate hundreds of molecules of specific enzyme from one activated phytochrome molecule

7. Fig. 39-4-3 An example of signal transduction in plants: the role of phytochrome in the de-etiolation (greening) response Transduction Reception Response 2 1 3 Transcription factor 1 CYTOPLASM NUCLEUS NUCLEUS Specific protein kinase 1 activated Plasma membrane cGMP P Second messenger produced Transcription factor 2 Phytochrome activated by light 2. One pathway uses cGMP as 2nd messenger that activates specific protein kinase. Other pathway involves increase in cytosolic level of Ca2+, which activates different protein kinase P Cell wall Specific protein kinase 2 activated 3. Both pathways lead to expression of genes for proteins that function in de-etiolation (greening) response Transcription 1. Lightsignal detected by phytochrome receptor; phytochrome undergoes change in shape, which then activates at least two signal transduction pathways Translation De-etiolation (greening) response proteins Ca2+ channel opened Ca2+

8. Response • Signal transduction pathway leads to regulation of one or more cellular activities • In most cases, these responses to stimulation involve increased activity of enzymes • This can occur by transcriptional regulation (increases/decreases synthesis of mRNA that codes for enzyme) or post-translational modification (activates existing enzyme molecules)

9. Transcriptional Regulation • Specific transcription factors bind directly to specific regions of DNA and control transcription of genes • In phytochrome-induced de-etiolation, several transcription factors activated by phosphorylation in response to appropriate light conditions • Mechanism by which signal promotes new developmental course may depend on • Positive transcription factors (activators): proteins that increase transcription of specific genes • Negative transcription factors (repressors): proteins that decrease transcription of specific genes • Mutation in Arabidopsis allows for light-grown morphology underground (except for pale color because no chlorophyll grown because no light) because genes activated by light normally blocked by repressor eliminated by mutation

10. Post-Translational Modification of Proteins • Post-translational modification involves modification of existing proteins in signal response • Modification often involves phosphorylation of specific amino acids which alters protein’s hydrophobicity/ activity • Many 2nd messengers (including some forms of phytochrome) activate protein kinases directly, often leading to kinase cascades • Many signal pathways ultimately regulate synthesis of new proteins, usually by turning specific genes on or off

11. De-Etiolation (“Greening”) Proteins • Many enzymes that function in certain signal responses • Are directly involved in photosynthesis • Are involved in supplying chemical precursors for chlorophyll production • Affect levels of plant hormones that regulate growth

12. Concept 39.2: Plant hormones help coordinate growth, development, and responses to stimuli • Hormones: chemical signals that coordinate different parts of organism • Signaling molecule produced in tiny amounts by one part of organism and transported to other parts, where it binds to specific receptor and triggers responses in target cells/tissues • Many plant biologists prefer plant growth regulator to describe organic compounds that modify or control one or more specific physiological processes within plant

13. The Discovery of Plant Hormones http://bcs.whfreeman.com/thelifewire/content/chp38/3802001.html • Tropism: any response resulting in curvature of organs toward or away from stimulus (often caused by hormones) • In late 1800s, Charles Darwin/son Francis conducted experiments on phototropism(plant’s response to light) • Observed that grass seedling could bend toward light only if tip of coleoptile was present (maximum exposure of leaves to light for photosynthesis) • Shoot of sprouting grass (enclosed in coleoptile) grows straight upward if seedling kept in dark/illuminated for all sides uniformly • If illuminated form one side, grows toward light (results from differential growth of cells on opposite sides of coleoptile; cells on darker side elongate faster than those on brighter side) • Postulated signal was transmitted from tip to elongating region

14. Darwin/Darwin: phototropic response only when tip is illuminated: Only tip of coleoptile senses light Phototrophic bending occurred at distance from site of light perception (tip)

15. Fig. 39-5c RESULTS Boysen-Jensen: phototropic response when tip is separated by permeable barrier, but not with impermeable barrier Demonstrated that signal was mobile chemical substance Light Tip separated by gelatin (permeable) Phototrophic response occurred Tip separated by mica (impermeable) No phototrophic response occurred

16. Fig. 39-6 http://bcs.whfreeman.com/thelifewire/content/chp38/3802002.html RESULTS Excised tip placed on agar cube • In 1926, Frits Went extracted chemical messenger for phototropism, auxin, by modifying earlier experiments • Auxin later purified/ chemical structure found to be IAA (indoleacetic acid) by Kenneth Thimann Growth-promoting chemical diffuses into agar cube Agar cube with chemical stimulates growth Control (agar cube lacking chemical) has no effect Offset cubes cause curvature Off center blocks caused decapitated coleoptiles to bend away from side with agar block, as though growing toward light Concluded that agar block contained chemical produced in coleoptile tip that stimulated growth as it passed down coleoptile/ which curved toward light because higher concentra- tion of growth-promoting chemical on darker side of coleoptile Control

17. These studies show asymmetrical distribution of auxin moving down from coleoptile tip and causes cells on darker side to elongate faster than cells on brighter side • Studies of organs other than grass coleoptiles provide less support for this, but there is asymmetrical distribution of certain substances that may act as growth inhibitors, and these are more concentrated on lighted side of stem

18. A Survey of Plant Hormones • Plant hormones are produced in very low concentration, but minute amount can greatly affect growth/development of plant organ • Amplified in some way • Hormone may act by altering expression of genes, by affecting activity of existing enzymes, by changing properties of membranes • Any of these actins could redirect metabolism and development of cell responding to small number of hormone molecules • In general, hormones control plant growth/development by affecting division, elongation, and differentiation of cells • Some mediate shorter-term physiological responses to environmental stimuli • Each hormone has multiple effects, depending on site of action, concentration, developmental stage of plant

19. Table 39-1

20. Auxin • Auxinrefers to any chemical that promotes elongation of coleoptiles (multiple functions in flowering plants: cell elongation/lateral root formation) • Indoleacetic acid (IAA) is common natural auxin in plants • Transported directly through parenchyma tissue, from one cell to another • Polar transport: unidirectional transport of auxin (in shoot, moves only from tip to base) • Polarity of auxin movement attributable to polar distribution of auxin transport protein in cells that move hormone from basal end of one cell into apical end of neighboring cell

21. The Role of Auxin in Cell Elongation • One of chief functions of auxin is to stimulate elongation of cells within young developing shoots • Apical meristem of shoot major site of auxin synthesis, moving down to region of cell elongation • According to acid growth hypothesis, auxin stimulates proton pumps in plasma membrane  • Proton pumps lower pH in cell wall, activating expansins (enzymes that loosen wall’s fabric by breaking hydrogen bonds between cellulose microfibrils and other cell wall constituents) • With the cellulose loosened, cell can elongate • Also rapidly alter gene expression, causing cells in region of elongation to produce new proteins within minutes

22. Figure 39.8 Cell elongation in response to auxin: the acid growth hypothesis 3 Expansins separate microfibrils from cross- linking polysaccharides. Cell wall–loosening enzymes Cross-linking polysaccharides Expansin CELL WALL 4 Cleaving allows microfibrils to slide. Cellulose microfibril H2O Cell wall Cell wall becomes more acidic. 2 Plasma membrane 1 Auxin increases proton pump activity. Nucleus Cytoplasm Plasma membrane Vacuole CYTOPLASM 5 Cell can elongate. http://bcs.whfreeman.com/thelifewire/content/chp38/3802003.html

23. Lateral and Adventitious Root FormationAuxins as Herbicides/Other Effects of Auxin • Auxin is involved in root formation and branching • Used in vegetative propagation of plants by cuttings • Overdose of synthetic auxins can kill eudicots • Monocots rapidly inactivate these synthetic auxins • Auxin affects secondary growth by inducing cell division in vascular cambium and influencing differentiation of secondary xylem • Developing seeds produce auxin which promotes fruit growth (can induce normal fruit development without pollination for commercially grown seedless tomatoes)

24. CytokininsControl of Cell Division and Differentiation • Cytokininsare so named because they stimulate cytokinesis (cell division) • Produced in actively growing tissues (roots, embryos, and fruits) • Work together w/auxin to control cell division/differentiation • When concentration of both at certain levels, mass of cells continues to grow, but remains cluster of undifferentiated cells (callus) • If cytokinin levels increase, shoot buds develop • If auxin level increase, roots form

25. Control of Apical Dominance • Cytokinins, auxin, and other factors interact in control of apical dominance, terminal bud’s ability to suppress development of axillary buds • Direct inhibition hypothesis says auxin/cytokinins act antagonistically in regulating axillary bud growth • Auxin transported down shoot from apical bud directly inhibits axillary buds from growing, causing stem to elongate • Cytokinins entering shoot system from roots counter action by signaling axillary buds to grow • Does not account for all experimental findings

26. Fig. 39-9 Lateral branches “Stump” after removal of apical bud (b) Apical bud removed, enables lateral branches to grow (removes inhibition) Axillary buds Apical bud intact (primary source of auxin) Inhibition of growth of axillary buds, possibly influenced by auxin from apical bud, favors elongation of shoot’s main axis (c) Auxin added to decapitated stem prevents lateral branches from growing

27. Anti-Aging Effects • Cytokinins retard aging of some plant organs by inhibiting protein breakdown, stimulating RNA and protein synthesis, and mobilizing nutrients from surrounding tissues • If leaves removed from plant dipped in cytokinin solution, stay greener much longer • Also slows deterioration of leaves on intact plants (used to spray on cut flowers to keep fresh)

28. GibberellinsStem Elongation/Fruit Growth • Gibberellinshave variety of effects, such as stem elongation, fruit growth, and seed germination • Major sites of gibberellin production are roots/young leaves • Stimulate growth of leaves/stems, little effect on roots • In stems, they stimulate cell elongation and cell division • Both auxin/gibberellins must be present for some fruit to ripen • Used in spraying of Thompson seedless grapes which makes individual grapes grow larger/ make internodes elongate, allowing more space for individual grapes (reduces yeasts/other microorganisms from infecting fruit)

29. Fig. 39-11 Mobilization of nutrients by gibberellins during the germination of grain seeds such as barley 1 After water is imbibed, gibberellins (GA) from embryo signals to aleurone to germinate Sugars and other nutrients are consumed. 2 3 Aleurone secretes -amylase and other enzymes. Aleurone Endosperm -amylase Sugar GA GA Water Radicle Scutellum (cotyledon) Germination

30. Brassinosteroids • Brassinosteroidsare steroids, chemically similar to sex hormones of animals • Induce cell elongation/division in stem segments and seedlings • Reduce leaf abscission (leaf drop) • Promote xylem differentiation

31. Abscisic Acid • Abscisic acid (ABA)slows growth • Antagonistic to growth hormones and ratio determines final physiological outcome • Two of many effects of ABA • Seed dormancy • Drought tolerance

32. Seed Dormancy • Seed dormancy ensures that seed will germinate only in optimal conditions • In some seeds, dormancy is broken when • ABA is removed by heavy rain, light, or prolonged cold • Light/prolonged exposure to cold to inactivate ABA • Precocious (early) germination is observed in maize mutants that lack transcription factor required for ABA to induce expression of certain genes

33. Drought Tolerance • ABA is primary internal signal that enables plants to withstand drought • Plant begins to wilt  ABA accumulates in leaves  rapid closing of stomata  reduces transpiration/prevents further water loss • ABA causes potassium channels in guard cells to open  loss of potassium ions from cells  osmotic loss of water  guard cell turgor reduction  closing of stomatal pores

34. Ethylene (H2C=CH2) The Triple Response to Mechanical Stress • Plants produce ethylenein response to stresses such as drought, flooding, mechanical pressure, injury, and infection • Effects of ethylene include response to mechanical stress, senescence, leaf abscission, fruit ripening, in response to high concentrations of externally applied auxin • Ethylene induces triple response, which allows growing shoot to avoid obstacles (rock in ground) • Consists of slowing of stem elongation, thickening of stem (making it stronger), and horizontal growth (curvature) • Resumes vertical growth when obstacle gone

35. Ethylene-insensitive mutants fail to undergo triple response after exposure to ethylene because they lack functional ethylene receptor (ein or ethylene-insensitive mutants) • Others undergo triple response in air without physical obstacles and have regulatory defect that causes them to produce ethylene at rates 20x normal (eto or ethylene overproducing mutants) • Can be restored to wild-type by treating seedlings with inhibitors of ethylene synthesis • Other mutants undergo triple response in air but do not respond to inhibitors of ethylene synthesis (ctr, constitutive triple-response mutants) • Do not respond to inhibitors of ethylene synthesis

36. Fig. 39-14 ein mutant ctr mutant ein mutant fails to undergo triple response in presence of ethylene (b) ctr mutant undergoes triple response even in absence of ethylene

37. Senescence • Senescence: programmed death of plant cells or organs or entire plant • Starts w/onset of apoptosis (programmed cell death) where newly formed enzymes break down many chemical components (chlorophyll, DNA, RNA, proteins, membrane lipids) to be salvaged by plant • Burst of ethylene associated w/programmed destruction of cells, organs, entire plant

38. Leaf Abscission • Change in balance of auxin and ethylene controls leaf abscission (process that occurs in autumn when leaf falls) • Essential elements salvaged/stored in stem parenchyma cells • Nutrients recycled back to developing leaves next spring • After leaf falls, protective layer of cork becomes leaf scar that prevents pathogens from invading plant

39. Fruit Ripening • Immature fleshy fruit (tart, hard, green) protect them from herbivores • Ripe fruit attracts animals to disperse seeds • Burst of ethylene production (positive feedback) infruit triggers ripening process (enzymatic breakdown of cell wall softens fruit, conversion of starches/acids to sugars makes them sweet) • Moving air prevents ethylene accumulation/ carbon dioxide prevents ethylene production  slows ripening of stored fruits

40. Systems Biology and Hormone Interactions • Interactions between hormones and signal transduction pathways make it hard to predict how genetic manipulation will affect plant • Systems biology seeks comprehensive understanding that permits modeling of plant functions

41. Concept 39.3: Responses to light are critical for plant success • Light cues many key events in plant growth and development, photosynthesis, measuring passage of days and seasons • Photomorphogenesis: effects of light on plant morphology • Plants detect not only presence of light but also direction, intensity, and wavelength (color) • Graph (action spectrum)depicts relative effectiveness of different wavelengths of radiation in driving particular process • Two peaks (red/blue light) for photosynthesis • Action spectra are useful in studying any process that depends on light (phototropism) • Two major classes of light receptors: blue-light photoreceptorsand phytochromes

42. Fig. 39-16 Action spectrum for blue-light-stimulated phototropism in maize coleoptiles 1.0 436 nm 0.8 0.6 Phototropic effectiveness 0.4 0.2 0 500 400 550 450 650 700 600 Wavelength (nm) (a) Action spectrum for blue-light phototropism Light Time = 0 min Time = 90 min (b) Coleoptile response to light colors Phototropic bending toward light controlled by phototropin (photoreceptor sensitive to blue/violet light, particularly blue light

43. Blue-Light PhotoreceptorsPhytochromes as Photoreceptors • Various blue-light photoreceptors control hypocotyl elongation, stomatal opening, and phototropism • Phytochromes are pigments that regulate many of plant’s responses to light throughout its life • These responses include seed germination and shade avoidance

44. Phytochromes and Seed Germination • Many seeds remain dormant until light conditions change • In 1930s, scientists at U.S. Department of Agriculture determined action spectrum for light-induced germination of lettuce seeds • Red light increased germination, while far-red light inhibited germination • Final light exposure was determining factor • Effects of red/far-red light reversible

45. Fig. 39-18 Photoreceptor responsible for opposing effects of red/far-red light are phytochromes Two identical subunits, each consisting of polypeptide component covalently bonded to nonpolypeptide chromophore Chromophore Photoreceptor activity Kinase activity

46. Phytochromes exist in two photoreversible states • Depend on color of light provided • Converts Pr (inhibits germination) to Pfr, which triggers many developmental responses (germination) • Though light contains both red and far red light, conversion to Pfr faster than conversion to Pr so ratio of Pfr to Pr increases in light, triggering germination

47. Fig. 39-19 http://highered.mcgraw-hill.com/sites/9834092339/student_view0/chapter41/animation_-_phytochrome_signaling.html Pfr Pr Red light Responses: seed germination, control of flowering, etc. Synthesis Far-red light Slow conversion in darkness (some plants) Enzymatic destruction

48. Phytochromes and Shade Avoidance • Phytochrome system also provides plant with information about quality of light • Sunlight contains both red/far-red radiation • During day, Pr⇄ Pfr interconversion reaches dynamic equilibrium, with ratio of two phytochrome forms indicating relative amounts of red/far-red light • Allows plants to adapt to changes in light conditions • Shaded plants receive more far-red than red light • In “shade avoidance” response, phytochrome ratio shifts in favor of Pr when tree is shaded, inducing tree to allocate more resources to growing taller • Direct sunlight stimulates branching/inhibits vertical growth

49. Biological Clocks and Circadian Rhythms • Phytochrome helps plant keep track of passage of days/seasons • Many plant processes (transpiration, synthesis of certain enzymes) oscillate during the day • Response to changes in light levels, temperature, and relative humidity accompanying 24-hour cycle of day/night • Many legumes lower their leaves in evening and raise them in morning, even when kept under constant light or dark conditions

50. Circadian rhythms: cycles that are about 24 hours long and are governed by internal “clock” rather than daily responses to environmental cycle • Can be entrained (set) to exactly 24 hours by day/night cycle from environment • Deviations (free-running periods of 21-27 hours) occur when organism kept in constant environment (not erratic because still keep perfect time, just not synchronized with outside world) • We can interfere with biological rhythm, but will reestablish position for time of day when interference removed • Clock may depend on synthesis of protein regulated through feedback control and may be common to all eukaryotes