Light Controls of Plant Development 1
Electromagenetic spectrum 2
Light It is vital for photosynthesis and is also necessary to direct plant growth and development. It acts as a signal to initiate and regulate photoperiodism and photomorphogenesis. 3
Plant photoreceptors Phytochromes are primarily responsible for perception of red (620-700 nm) and far-red light (700-800 nm). Cryptochrome and NPH1 (phototropin) mediate responses to UV-A (320-380 nm) and blue light (380-500 nm). One or more unidentified UV-B receptor(s) are responsible for UV-B (280-320 nm) perception. 4
Blue light responses Plant responses regulated by the blue light include phototropism, stomatal opening and chlorophyll synthesis. 5
Phytochrome responses Plant responses regulated by the phytochrome include seed germination, flowering, and photomorphogenesis (i.e., leaf expansion, stem elongation and chloroplast development). 6
Phytochrome It is a photoreceptor that absorbs primarily red light and far-red light. It is a soluble protein with a molecular mass of ~ 250 kda. It occurs as a dimer made up of two equivalent subunits. Each subunit consists of two component: a light-absorbing pigment called the chromophore, and a polypeptide chain called the apoprotein (molecular mass ~ 125 kda). Together, the apoprotein and its chromophore made up the haloprotein. In higher plants, the chromophore of phytochrome is a linear tetrapyrrole termed phytochromobilin. 7
The chromophore portion of the phytochrome can exist in one of two forms, Pr or Pfr. 8
Phytochrome (cont.) There is only one chromophore per monomer of apoprotein, and it is attached to the protein through a thioether linkage to a cysteine residue. One part of the protein acts as the photoreceptor which bears the chromophore, and the other as a kinase which triggers cellular responses. When the chromophore absorbs light, it isomerizes from one form to the other. This changes in configuration results in a slight change in the kinase portion of the protein. The kinase is the biologically active region of the molecule, and its interaction with other molecules elicits a physiolgical response. 9
Phytochrome can interconvert between Pr and Pfr forms In dark-grown or etiolated plants, phytochrome is synthesized in a red-light absorbing form, referred to as Pr. Pr (blue pigment) is converted by red light to a far-red light-absorbing form called Pfr (blue-green pigment). Pfr in turn, can be converted back to Pr by far-red light. This conversion/reconversion is known as photoreversibility. The action spectrum of light needed for these responses shows a peak in the red at about 660 nm. These responses can be reversed by an application of far-red light (peak at 730 nm) soon after the red treatment. 10
Phytochrome can interconvert between Pr and Pfr forms (cont.) 11
The Pr form: It absorbs at a peak of 660 nm. It is the form synthesized in dark-grown seedlings. When Pr absorbs red light, it is converted to the Pfr form. The Pfr form: It absorbs at a peak of 730 nm. The Pfr form is the active form that initiates biological response. When Pfr absorbs far-red light, it is converted to the Pr form. Pfr can also spontaneously revert to the Pr form in the dark over time. This is called dark reversion. 12
Photostationary state Pfr absorbs some red light, so in red light, there is an equilibrium of 85% Pfr and 15% Pr. Pr absorbs very little far-red light, so in far-red light, there is an equilibrium of 97% Pr and 3% Pfr. Both forms of phytochrome can also absorb blue light. 13
Photoreversibility responses induced by phytochromes 14
Phytochrome genes and proteins There are five phytochrome genes in dicots, corresponding to Arabidopsis termed PHYA, PHYB, PHYC, PHYD and PHYE. Monocots, like duckweeds, have a fewer phytochrome genes, homologs of PHYA, PHYB and PHYC. There are two types of phytochrome. Type I phytochrome is light-labile. The only member is phya. Type II phytochrome is light-stable. The members are phyb, phyc, phyd and phye. Note: The haloproteins are written by phya, phyb and so on. 15
PHYA: The PHYA gene is transcriptionally active in dark-grown seedlings, but its expression is strongly inhibited in the light in monocots. In dark-grown oat, treatment with red light reduces phytochrome synthesis because PrA (phya in Pr form) inhibits the expression of its own gene. In addition, the PHYA mrna is unstable, so once darkgrown seedling are transferred to the light, PHYA mrna rapidly disappears. The inhibitory effect of light on PHYA gene transcription is strong in monocots but is less dramatic in dicots. 16
PHYA: (cont.) Protein destruction also regulates amount of phya. PfrA is unstable. PfrA may become marked for destruction by the ubiquitin system. Note: Ubiquitin is a small poly peptide that binds covalently to proteins and serves as a recognition site for large proteolytic complex, the proteasome. Therefore, monocots rapidly lose most of their phya in the light as a result of a combination factors: inhibition of transcription, mrna degradation, and proteolysis. In dicots, phya levels also decline in the light as a result of proteolysis, but not as dramatically. 17
PHYA: (cont.) Lost of phya in the light: 18
PHYB-E: phyb through phye are synthesized at the same rate in both darkness and light, and their concentration is about the same in both cases, with phyb being the predominant in light-grown plants. The expression of mrnas of PHYB through PHYE genes is not significantly changed by light, and the encoded phyb through phye proteins are more stable in the Pfr form than is PfrA. 19
Localization of phytochrome In etiolated seedlings the highest phytochrome levels are usually found in meristematic regions or in regions that were recently meristematic, such as the bud and first node of pea, or the tip and node regions of the coleoptile in oat. 20
Light-induced seed germination (1) Phytochrome mutants (2) Gibberellins 21
(1) Phytochrome mutants Newly imbibed Arabidopsis seeds display some germination in darkness, however germination is promoted by low amounts (fluences) of red light and inhibited by low amounts of far-red light. Note: The amount of light is referred to as the fluence, which is defined as the number of photons impinging on a unit surface area. Units are moles of quanta per square meter (mol m-2). After several days in darkness, imbibed seeds become more sensitive to light: they will germinate in response to a broad spectrum of radiation, from UV-B to far-red, provided at very low fluences. These contrasting responses suggest that more than one photoreceptors control seed germination. 22
(1) Phytochrome mutants (cont.) In phyb mutants, germination in darkness or after treatment with low fluences of red light is substantially reduced compared to wild type. This suggests that phyb is largely responsible for mediating germination under these conditions. The induction of germination by red light, and its inhibition by far-red light, suggest that phyb promotes germination when it is in the Pfr form. Other studies in phyc, phyd and phye mutants also supports that the phytochromes encoded by these genes regulate germination in red light. 23
(1) Phytochrome mutants (cont.) In phya mutants, they are specifically impaired in the highly light sensitive response, which develop when imbibed seeds are kept in total darkness. This suggests that phya mediates germination under these conditions. The extreme light sensitivity of imbibed seeds develops because of the accumulation of high quantities of phya in the Pr form during prolonged darkness. Under these conditions, germination is promoted even when a tiny fraction of the accumulated PrA is converted to PfrA. Since PrA displays some absorption over the whole spectrum, this allows exposure to a very low fluence of any wavelength to induce germination. 24
Seed germination in wild-type and phytochrome mutants of Arabidopsis in different light environments. (a) Some wild-type seeds germinate even in darkness. This response is reduced in phyb mutants, suggesting that it requires phyb. (b) Germination of wild-type seeds is promoted by low fluence red light. This response is also reduced in phyb mutants, suggesting that it requires phyb. (c) When imbibed for several days in darkness, wild-type seeds become extremely light-sensitive and will germinate in response to very low fluence light across a broad spectrum. This response is lost in phya mutants, suggesting that it 25 requires phya.
(1) Phytochrome mutants (cont.) The responses of Arabidopsis to different light regime allow seed germination in a wide range of environments. Some seeds germinate in darkness when temperature and soil moisture allow. Promotion of germination by red light (mediated largely by phyb) will enhance the germination on the soil surface in sunlight (high R : FR). The extremely light-sensitive response mediated by phya will promote germination (1) in buried seeds after exposure to flashes of light during soil disturbance; (2) of seeds buried just below the soil surface; and (3) of seeds on the soil surface just beneath a heavy canopy (low R : FR ratio). 26
(2) Gibberellins The biosynthesis of gibberellins is required for germination in most species. Arabidopsis gibberellic acid 1 (ga1) mutants are not able to synthesize gibberellins and fail to germinate even in the light unless gibberellins are provided exogenously. Phytochromes control germination by influencing gibberellin biosynthesis and sensitivity. Red light induces transcription of GA4 and GA4H genes of Arabidopsis. These genes encode enzyme gibberellin 3βhydroxylase which catalyses the final step in the synthesis of biologically active gibberellins. In contrast far-red light represses the transcription of these genes. 27
(2) Gibberellins (cont.) Furthermore, ga1 mutants require less exogenous gibberellin to germinate when grown in red light, indicating that phytochrome signalling also increases the seed s sensitivity to gibberellins. The interaction between phytochrome and gibberellins during germination. Phytochrome in the Pfr form increases gibberellin synthesis by promoting transcription from the GA4 and GA4H genes. These genes encode gibberellin 3β-hydroxylase, which catalyses the final step in the synthesis of bioactive gibberellins. Pfr signalling also increases the sensitivity of the seed to gibberellins. Arrows accompanied by a plus sign indicate positive regulation. 28
Seedling etiolation and photomorphogenesis (1) Skotomorphogenesis (2) Photomorphogenesis (3) Light perception by the seedling (4) Negative regulators of photomorphogenesis (5) Hormones 29
(1) Skotomorphogenesis (etiolation) Angiosperm seedlings become etiolated in constant darkness. This is characterized by reduced root development; a hook-shaped shoot; rapid stem or hypocotyl elongation; fold, unexpanded leaves and/ or cotyledons; an inactive shoot apical meristem; and etioplasts (chloroplast precursors that lack chlorophyll). Etiolation is an adaptation to germination below the soil surface. Resources are directed towards rapid upward growth through the soil, with the hook-shaped shoot protecting the shoot apical meristem. 30
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(2) Photomorphogenesis (de-etiolation) Light-grown seedlings display photomorphogenesis, sometimes called de-etiolation. This is characterized by extensive root development; slow stem elongation; straightening of the shoot; unfolding and expansion of leaves and cotyledons; the expression of genes necessary for pigment biosynthesis and the photosynthetic machinery; development of etioplasts into chloroplasts; and activation of the shoot apical meristem. 32
(3) Light perception by the seedling Photomorphogenesis can be induced by a broad spectrum of illumination. Arabidopsis seedlings de-etiolate in response of UV-A, blue, red or far-red light. Some aspects of photomorphogenesis, such as changes in gene expression or the inhibition of hypocotyl elongation, can be induced by brief pulses of light. However, full deetiolation requires continuous illumination. Cryptochromes and phytochromes are the photoreceptors that mediate photomorphogenesis. The phenotypes of cry1 and cry2 mutants of Arabidopsis indicate that both are required for full de-etiolation in response of UV-A or blue wavelengths. 33
(3) Light perception by the seedling (cont.) The phenotypes of phytochrome mutants show that phyb is needed for de-etiolation under continuous red light, while phya mediates de-etiolation under continuous far-red light. phyb mutants show residual photomorphogenesis under continuous red light, indicating that other light stable phytochromes mediates de-etiolation. Signal transduction downstream of these photoreceptors induces photomorphogenesis in two ways. Firstly, photosynthetic genes may be activated by direct positive regulation downstream of light perception. Secondly, light-induced signalling inactivates negative regulators of photomorphogenesis. 34
Photomorphogenesis is promoted by continuous red light, continuous far-red light and continuous UV-A/ blue light in wild-type seedlings of Arabidopsis. The phenotypes of photoreceptor mutants indicate that the response to red light requires phyb (a); the response to far-red light requires phya (b); and the response to UV-A/ blue light requires cry1 (c). (d) in darkness, wild type seedlings are etiolated and photomorphogenesis is suppressed. The pleiotropic cop/ det/ fus mutants have photomorphogenic phenotype in darkness, indication that the COP/ DET/ FUS genes are required to suppress photomorphogenesis in darkgrown seedlings. 35
(4) Negative regulators of photomorphogenesis Genetic studies in Arabidopsis indicate that etiolation is a consequence of the inhibition of photomorphogenesis. A number of Arabidopsis mutants have been identified that display photomorphogenesis when grown in continual darkness. This phenotype is described either as de-etiolated (det) or constitutively photomorphogenic (cop). det and cop mutations are recessive and result in loss of gene function. The wild-type function of the DET and COP is, therefore, to suppress photomorphogenesis in darkgrown seedlings. Additional mutants with the det/ cop phenotype were recovered in screens for purple seedlings and these mutants are called fusca (fus). 36
(5) Hormones Several aspects of etiolation and/ or photomorphogenesis can be altered by hormone treatments and mutations affecting hormone synthesis and/ or response. Two recessive mutants in Arabidopsis, det2 and constitutive photomorphogenesis and dwarfism (cpd), result in partially de-etiolated phenotype in dark-grown seedlings. In darkness, det2 and cpd mutants have short hypocotyls no apical hook and express light-inducible genes. DET2 and CPD encode enzymes that catalyse steps in the biosynthesis of brassinosteroid hormone, brassinolide, and det2 and cpd mutants have very low brassinolide content. Therefore, brassinolide is required for at least some aspects of etiolation. 37
(5) Hormones (cont.) Addition of cytokinin or genetic perturbation of auxin signalling can cause partial de-etiolation in the dark. Gibberellin-deficient mutants are dwarfed relative to wild type, indicating that gibberellin is required for hypocotyl and stem elongation. It is possible that the increase in gibberellin concentration in germinating embryos allows the subsequent rapid upward growth of etiolated seedlings. 38
Shade escape (1) Shading regulates plant development (2) R : FR ratio and shading 39
(1) Shading regulates plant development The change in light quality due to shading by other plants also regulates plant development. One of the main different between sunlight and light that has passed through leaves is a fall in the ratio of red to farred wavelengths (R : FR) due to preferential absorption of red light by chlorophyll. A drop in R : FR produces different responses in different plants. Most non-angiosperms are shade tolerant and respond by developing a morphology suited to shaded conditions, for example producing large but thin shade leaves. 40
(1) Shading regulates plant development (cont.) In contrast, angiosperms usually respond with rapid internode and petiole elongation, a decrease in leaf area and thickness, enhance apical dominance (i.e. reduced branching) and early flowering. This response is called shade escape. 41
(2) R : FR ratio and shading R : FR is defined as the ratio in intensity of wavelengths from 655-665 nm (red) and 725-735 nm (far-red). Note: R : FR = photon fluence rate in 10 nm band centered on 660 mn photon fluence rate in 10 nm band centered on 730 mn In open environments, the day time R : FR is about 1.15 : 1 regardless of whether the sky is clear or overcast. Due to the presence of chlorophyll and other pigments, leaves strongly absorb wavelengths below 700 nm. Therefore, R : FR falls dramatically beneath a canopy due to the depletion of red wavelength, reaching values as low as 0.5 : 1. 42
(2) R : FR ratio and shading (cont.) R : FR also falls in the proximity of vegetation due to low R : FR in the light reflected from leaves. This allows some species to initiate shade escape response before actual shading occurs by reacting to light reflected from neighbouring plants. 43
The ratio of red light to far-red light regulates the shade escape response. (a) Comparing spectral fluence rates (light intensity at different wavelengths) in sunlight and canopy shade shows that the ratio R : FR is much higher in sunlight. Therefore, R : FR is an indicator of the degree of shading by neighbouring plants. (b) If red is high relative to far red, as in sunlight, the Pr will convert to Pfr. High Pfr inhibits the transcription of the ATHB-2 gene and hence inhibits shade escape. 44
(2) R : FR ratio and shading (cont.) Phytochromes enable plants to sense shading by other plants. As shading increase, the R : FR ratio decrease. The greater proportion of far-red light converts more Pfr to Pr, and the ratio of Pfr to total phytochrome (Pfr/Ptotal) decreases. When stimulated natural radiation was used to vary the far-red content, it was found that for so-called sun plants (plants that normally grown in an opened-field habitat), the higher the far-red content (i.e., the lower Pfr : Ptotal ratio) the higher the rate of stem extension. 45
(2) R : FR ratio and shading (cont.) In other words, stimulated canopy shading (high levels of far-red light) induces sun plants or shade-avoiding plants to allocate more of resources to grow taller. For shade plants which normally grow in the shaded environment, they showed little or no reduction in their stem extension rate as they were exposed to higher R : FR ratio. Thus there appears to be a systemic relationship between phytochrome-controlled growth and species habitat. 46
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