Lecture 21 - environmental regulation of development Flashcards
(21 cards)
How does the environment regulate animal development?
YES (e.g. sex determination in amphibians by temp. of temp. of eggs & maturation Human height
BUT - no average change in pattern (no extra legs etc.) - different in plants
How does the environment regulate plant development?
-Plants cannot move so must adapt
- Changes are more dramatic (but often repetition of pattern)
- More amenable to genetic analysis - easy to quantify response & then perform mutational analysis
- Molecular programmes (at the chromatic level) dictate actions, such as when leaves grow - e.g. temperatures
What is the Light Environment?
Photoperiod - the day length
Quantity - the amount of light reaching a surface
Quality - the wavelengths & balance of wavelengths
Red - 600-700 nm (wavelength)
Far-red - 700-800 nm (wavelength)
Red:far-red ratio is important in regulating a number of responses in plants
Why is the sky blue?
Because small molecule scatter blue light, as light travels through the atmosphere. Sunset leads to different colour due to different levels of blue & red light (short & long wavelength). LIGHT IS VARIABLE
FULL SUN - shorter wavelengths (blue) more easily scattered
LOW SUN (dusk/dawn) - longer wavelengths (red) - enhanced scattering
At what wavelength do Chlorophyll a, b & caretenoids levels drop?
Green light (550-600)
How is light not just about photosynthesis?
As light passes through leaves Red/BLue preferentially absorbed - enriched in far-red (Ft)
Light reflected from leaves (also enriched in Fr)
UNDER A CANOPY - reduced quantity - low R:Fr
NEAR COMPETITORS - high quantity - low R:Fr
OPEN - high quantity + low R:Fr
Plants in open environment are exposed to high in blue & high R:Fr ratio light.
Plants in environments closer to other plants - exposed to less light, as well as less blue & R:Fr light.
What is phototropism?
- differential growth in response to stimulus
- perception at different site to growth
- ‘influence’ moves from site of perception to site of growth
- plant hormone auxin causes tropic growth
- becomes asymmetrically distributed on stimulus
STEM PHOTOTROPISM IS BLUE LIGHT DEPENDENT
plants are able to perceive unidirectional light- at the tip of the plant. There was a factor that become asymmetrically distributed within the stem, that resulted in differential growth of the stem.
Auxin was a hormone that was identified - derived from tryptophan. Asymmetric growth due to unidirectional BLUE light was due to a redistribution of auxin through the stem.
How was phototropism explored in experiments?
- Grew plants in the dark. Some were placed with a cap (apex couldn’t light). They were then exposed to blue light. This concluded that perception of the blue light did occur in this region.
- Tips were placed in agar blocks & exposed them to unidirectional blue light - plants were able to bend. He measured amount of auxin that diffused into agar block. Separated the agar block & measured the amount of auxin in each part.
Determine that auxin distribution drove the elongation growth of the plant.
What did exposure to blue light cause?
Phosphorylation of a membrane protein
~120kDa protein in pea membrane
- phosphorylated specifically in response to blue light
- Followed same dynamics as phototropism
- found in numerous plants
Extracted proteins plants in dark or low levels of blue light. Proteins found in cytoplasm or membrane-bound proteins. Looked for proteins undergoing phosphorylation & found that there was a protein in the membrane fraction that underwent phosphorylation specifically in response to blue light. It has the same action spectrum of something responding to phototropism. This was found in multiple plant species, meaning that it could be a key protein in regulating the blue light phototropic responses.
How do genetics lead to phototropism
Multiple different genes appear to regulate the response to unidirectional light. We know some of the mutations were from the same gene through a COMPLEMENTATION TEST - crosses & look at progeny. If you have mutants in the same gene - progeny should all have mutant phenotype, whereas mutants in different genes - all plants have a phototropic response
- nph1 mutants are defective in phototropism
How is genetics used to identify Phototropins?
Map based cloning strategy - laborious
Determine rough map position on chromosome 3 & identified closely linked PCR marker.
Screen Yeast artificial chromosome library with PCR markers
Narrowed down to 7kb fragment
Complemented nph1 mutant with fragment - so contained NPH1 gene
Used to screen cDNA library
Identified a 3.3kb transcript
What is NPH1?
A blue light photoreceptor
(apoprotein - the protein encoded component)
Made of:
LOV1 - light oxygen or voltage domain
LOV2 - light oxygen or voltage domain
STK - serine-threonine kinase domain
Photoreceptor can initiate responses to different wavelengths of light
What is the LOV domain?
Light oxygen or voltage domain:
- binds chromophore
- flavin mononucleotide
LOV2 - regulates STK domain
Each of the LOV domains bind chromophore. Conformation in the dark where the STK domain is brought close to the LOV2 domain, which inactivates it. In response to light, the FMN (flavin mononucleotide) binds to the LOV domains & causes a change in conformation of the protein component of the photoreceptor, releasing the STK domain, which auto-phosphorylates the phototropins & other targets.
Phototropins are the photoreceptors for blue light.
What is the Serine/Threonine Kinase?
- Blue light autophosphorylation
- conformational change
What are events downstream of phototropins?
- NPH3 is required for phototropism
- Phosphorylated by PHOT1
- mediates auxin gradient
What is seen in the wild-type?
Change in auxin, with higher concentration of auxin in shaded area, not seen in NHP3 mutant.
How does NPH3 localization change in light?
DARK - NPH3 localised to plasma membrane
LIGHT - internalized & forms gradient across bending tissue
NPH3 transgene fused with GFP. NPH3 is localized to the membrane in the dark. In light, NPH3 becomes internalized, so it is no longer associated with the membrane.
You can also start to see an NPH3 gradient across the stem - more on the light side than the shaded side. The light side isn’t going to undergo bending, but the shading side will.
What are differences between DARK & LIGHT?
DARK:
- PHOT1 inactive
- NPH3 membrane bound
- Auxin evenly distributed
LIGHT:
Shaded side:
- PHOT1 inactive
- NPH3 internalized
- High auxin
Lit side:
- PHOT1 active
- NPH3 internalized
- Low auxin
What do PIN proteins control?
PIN proteins control auxin distribution
- Efflux carriers that direct auxin transport
- Light changes PIN protein localization to direct auxin flow
Auxin can’t easily freely diffuse between cells. If it needs to move there must be transporters - PIN proteins are transporters that move auxin out of the cell. They are often associated with the outer membrane & distribution can change - redirect flow of auxin. Within a cell, THE LOCALIZATION OF PIN PROTEINS CAN CHANGE in response to signals, which can lead to a change in the flow of auxin, within an organ.
Exposure to unidirectional light leads the PIN proteins to become more localized to the shaded side. This means that more auxin is pushed out towards the shaded side.
Auxin distribution is dictated by the efflux carriers & the localization of the carries change in response to unidirectional blue light. This could mean that mutants of the PIN proteins are affected in phototropism.
What are PIN mutants?
PIN mutants are not phototropic
- pin3, pin4 & pin7 mutant does not show phototropism
PIN protein act redundantly to control a number of processes. Looked into single, double & triple mutants.
Summarise environmental regulation of development
- light is not just about photosynthesis
- light is an information source
- plants use photoreceptors to decode light & initiate responses
- phototropism involves stem bending towards light
- it is mediated by Phototropin blue light receptors
- Blue light activates PHOTs & in stems leads to NPH3 phosphorylation
- gradient of PHOT/NPH3 across stem
- Inversely related to auxin distribution
- Auxin redistribution requires PIN efflux carriers
