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AGR2451 - Lecture 4 (M. Raizada)
Notes:
-questionnaire and hand-out at the front
-this week’s reading on plant hormones on reserve:
Page 545-563 in Biology of Plants
-Review of previous 2 lectures:
1. How an enzyme works - the active site
2. How amino acids build a 3-D, folded enzyme
3. Biological switches- changes in conformation of a protein,
protein-protein-DNA interactions of transcription factors
4. The effect of small charged molecules (Phosphate) on
protein activity.
5. DNA -- mRNA--- protein. Why??
6. Transcription, splicing, mRNA export, translation,
post-translational modifications, protein folding,compartment
export/import.
7. Life is an orchestra of subsets of genes switching on and off to
create diverse cell types/diverse responses.
Lecture 4 -Extracellular & Intracellular Signalling Networks
I. Coordinating gene expression with the environment
To create life, biochemistry had to be placed inside a compartment
sequestered from the environment. However, an organism must sense
and response to its environment to survive. In particular, a plant is
immobile and must respond. The environment is outside of a cell. A
eukaryotic gene is inside the nucleus.
How does the extracellular environment turn a gene on and off? (class)
For Example: cold, pathogen attack, light quality and intensity
A) The signal must be perceived at the plasma membrane/wall surface.
B) The signal must be transmitted across the plasma membrane.
C) The signal must be communicated across the cytoplasm to the nucleus.
D) In the nucleus, the signal must interact with the appropriate
transcription factors to turn genes on and off.
“Signal
Transduction
Cascade”
=a series of
molecular
switches to
communicate
from the cell
surface to a
group of
genes
From Biochemistry and Molecular Biology of Plants
(W.Gruissem, B. Buchanan and R.Jones p.977
ASPP, Rockville MD, 2000
Slide 4.2
Signal Transduction Cascade
A) The signal must be perceived at the plasma membrance/wall surface.
B) The signal must be transduced across the plasma membrane.
How?
A stimulus causes a change in conformation of a protein receptor
embedded in the plasma membrane. A molecular stimulus is called a
ligand. The receptor consists of an extracellular (sensing) domain, a
membrane spanning domain that is hydrophobic, and an intracellular
domain that responds to the change in conformation to signal other
proteins/enzymes:
From Biochemistry and Molecular Biology of Plants
(W.Gruissem, B. Buchanan and R.Jones p.937
ASPP, Rockville MD, 2000
The surface of a pathogen or a molecule released from a pathogen (cell
wall fragment) can act to turn on a plant receptor to activate the plant
disease-resistance/defence response. Hence, plant receptors are crucial
for survival and a key for crop improvement.
C) A common way to communicate the signal from the membrane to
the nucleus is for the activated receptor to transfer a phosphate molecule
onto another enzyme thereby activating it (the phosphate changes its
conformation into a functional state). This enzyme is not anchored to
the membrane but diffusible inside the cytoplasm. It is called a
secondary messenger.
What is a kinase? an enzyme that transfers a phosphate, eg. to another
enzyme.
There are at least 340 kinase proteins in Arabidopsis.
Slide 4.3A
Signal Transduction Cascade
C) Example of a receptor kinase adding a phosphate to
activate a cytoplasmic secondary messenger
QuickTime™ and a
PNG decompressor
are needed to see this picture.
From Biochemistry and Molecular Biology of Plants
(W.Gruissem, B. Buchanan and R.Jones p.943
ASPP, Rockville MD, 2000
One kinase can activate a second kinase which can activate a
third kinase, etc.
Why did evolution build a cascade of second messengers in the
cytoplasm rather than a single secondary messenger from the plasma
membrane to the nucleus?
1. Amplify signal.
2. Many more control points.
Slide 4.3B
Signal Transduction Cascade
D) The signal must interact with the appropriate transcription factors to
turn genes on and off.
How?
•The secondary messengers must travel through the nuclear pore into
the nucleus, and there they directly bind to specific transcription factors
and/or add a phosphate to the transcription factors thereby changing
their conformation, causing them to be activated or repressed
(causing whole sets of genes to be transcriptionally turned on or off).
•Those genes that commonly share the DNA sequence (promoter or
enhancer) to which the activated transcription factor binds become
coordinately regulated.
From Biochemistry and Molecular Biology of Plants
(W.Gruissem, B. Buchanan and R.Jones p.962
ASPP, Rockville MD, 2000
Slide 4.3C
Coordinating Multiple Environmental Inputs
A cell must be able to coordinate multiple environmental inputs.
How is this achieved?
QuickTime™ and a
PNG decompressor
are needed to see this picture.
From Biochemistry and Molecular Biology of Plants
(W.Gruissem, B. Buchanan and R.Jones p.933
ASPP, Rockville MD, 2000
A) Combinatorial control of gene regulatory regions.
What is this and how is it achieved?
One gene may have a regulatory region consisting of multiple enhancer
sequences each of which binds a different transcription
factors, from a using a unique signal transduction cascade.
Light- Nitrogen- Drought- Responsive
Growth gene
6-20 base DNA enhancer sequences
Slide 4.4A
Coordinating Multiple Environmental Inputs
B) Cross-talk between signal transduction cascades.
The different signalling pathways may enhance or interfere
with one another by binding to one another’s enzymes or
transferring or removing phosphate molecules.
An enzyme that removes a phosphate = phosphatase. There are
at least 70 different phosphatases in Arabidopsis.
QuickTime™ and a
PNG decompressor
are needed to see this picture.
From Biochemistry and Molecular Biology of Plants
(W.Gruissem, B. Buchanan and R.Jones p.984
ASPP, Rockville MD, 2000
Why is such cross-talk between signal transduction enzymes
useful to the cell?
Can respond to a stimulus (eg. Pathogen attack) by turning
on/off different responses in a coordinated way.
Slide 4.4B
II. Cell-to-cell local coordination
When multicellular organisms developed, evolution needed to
allow cells to send information to neighboring cells to give them
positional information.
A) Cell-cell receptor-ligands. An adjacent cell exports a small protein
(peptide) or a chemical to its cell surface which binds to the receptor of
an adjacent cell to turn on/off a specific set of genes or activate/repress
specific enzymes.
QuickTime™ and a
PNG decompressor
are needed to see this picture.
B) What are plasmodesmata? They are direct cytoplasmic connections
between adjacent cells. This allows small molecules such as Ca++
and even transcription factors to move between adjacent cells in a
very regulated manner.
QuickTime™ and a
PNG decompressor
are needed to see this picture.
From Biochemistry and Molecular Biology of Plants
(W.Gruissem, B. Buchanan and R.Jones p.751and p.1021
ASPP, Rockville MD, 2000
Slide 4.5
AGR2451 - Lecture 6 - October 2nd
III. Long-distance signalling
As multicellular organisms became very complex, with distinct
organs, then gene expression in distant cells needed to be
coordinated in order for the organism to develop properly and to
respond to the environment.
A) This can be mediated by a wave of diffusible Ca++ ions.
These ions then bind to other enzymes to activate/repress them.
Similar to phosphate, Ca++ can also be used as a messenger
inside a cell: in this case, the Ca++ may be released from
the vacuole where it is stored, into the cytoplasm.
Cold-induced
wave of
Ca++ in a
tobacco leaf.
QuickTime™ and a
PNG decompressor
are needed to see this picture.
From Biochemistry and Molecular Biology of Plants
(W.Gruissem, B. Buchanan and R.Jones p.972
ASPP, Rockville MD, 2000
Slide 4.6
III. Long-distance signalling
B) The main method, however, are hormones, long-distance
signalling molecules.
C) Each cell may respond to the same signal in a different way
(for example: in drought, the leaves may stop expanding to
decrease evaporation while the roots might extend to find water).
Hence, there must be cell-specific receptors or secondary
messengers.
QuickTime™ and a
PNG decompressor
are needed to see this picture.
From Biochemistry and Molecular Biology of Plants
(W.Gruissem, B. Buchanan and R.Jones p.851
ASPP, Rockville MD, 2000
Slide 4.7A
III. Long-distance signalling - The Major Plant Hormones
Hormone
Auxin
Synthesized where?
leaf primordia
Function/Notes
•apical dominance
young leaves
•root induction
•vascular tissue
development
•stimulates fruit
development
•many others
Cytokinins
root tips
•cell division
•shoot formation
Ethylene
ripening or
senescence tissues
•fruit ripening
•tissue senescence
Abscisic acid
mature leaves,
especially after
water stress
•stomatal closure
•embryogenesis
•induces sugar
transport from
leaves to seeds
•induces storage
protein synthesis in
seeds
Gibberellins
young shoots and
developing seeds
•elongation of shoots*
•seed germination
•stimulates flowering
*Why was a mutation that altered Gibberellin synthesis (allele
Norin10) enormously responsible for the Green Revolution?
Shortened plant height in response to added fertilizer, preventing
plants from falling over and losing seeds.
Slide 4.7B
Hormones - Example Ethylene
Ethylene is a diffusible gas. One of its functions is to promote fruit
ripening. Ripening fruit releases this gas (hence this is why one bad
apple spoils the whole bag).
Ethylene = C2H4
Ethylene binds to a receptor and the
signal is transmitted via a phsphorylation
kinase cascade to the nucleus where the
signal activates transcription factors which
turn on genes involved in fruit ripening
such as cell wall degrading enzymes,
pigment activation and sugar release.
QuickTime™ and a
PNG decompressor
are needed to see this picture.
•Cell wall
degrading enzymes
•Pigment activation
From Biochemistry and Molecular Biology of Plants
(W.Gruissem, B. Buchanan and R.Jones p.980 and 1075
ASPP, Rockville MD, 2000
Slide 6.7C
Protein-Protein Interactions are Complex
Biologists are slowly determining which proteins interact with other
proteins for signalling, for biochemical pathways, and to create higher
order structures (such as microtubule cables). The result is the creation of
protein-interaction maps that display entire networks of proteins
working together. These maps illustrate that:
A) many proteins are required for every process, not single proteins.
B) It illustrates that signals from the environment
play a major role in the molecular events inside a cell.
C) In the future, such maps will be at the forefront of advances in both
agriculture and medicine.
Example of a
Yeast protein
Interaction map
Can plan
new herbicides
and pesticides
Eisenberg et al. (2000) Nature 405, 823-826
Nature Publishing Company, UK.
Slide 4.8
6.9
IV. Evolution by altering “Master Switches”
What is a "Master Switch"??
It is a single molecule (Transcription factor, receptor, messenger) which
switches on/off large numbers of genes or enzymes.
For example, there is a Master Switch to make eyes, switches to make
flowers, etc. Changes in the intensity of the switch or when and where
the switch acts can thus profoundly alter how an organism interacts with
its environment or how it develops. These small changes have likely led
to new species and dramatic changes during evolution. Similarly,
changes in transcription factors have also resulted in large changes in
phenotype with only a small change in genotype.
V. Summary of Key Concepts
•Biology has protein-based switches, including transcription factors
(on/off) and changes in protein conformation by Calcium/Phosphate
•The environment interacts with an organism by affecting one
or more of these switches.
•The environment turns genes on/off using a plasma membrane
receptor that activates a signal transduction cascade ultimately
resulting in the switching on/off of gene expression.
•There are also receptors and molecules that allow adjacent cells
and distant cells to communicate.
•Hormones allow an organism to have a coordinated response,though
different cells may turn on/off different genes in response to the same
hormone.
•A single gene may be turned on/off by multiple environmental
stimuli (combinatorial control) and different signalling cascades may
interfere with one another.
Slide 4.9