AGR2451 - Lecture 4 (M. Raizada)

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AGR2451 - Lecture 4 (M. Raizada)
Notes -questionnaire and hand-out at the
front -this weeks 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.
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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
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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
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Signal Transduction Cascade
C) Example of a receptor kinase adding a
phosphate to activate a cytoplasmic secondary
messenger
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
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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
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A cell must be able to coordinate multiple
environmental inputs. How is this achieved?
Coordinating Multiple Environmental Inputs
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
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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 anothers enzymes or transferring
or removing phosphate molecules. An enzyme that
removes a phosphate phosphatase. There are at
least 70 different phosphatases in Arabidopsis.
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
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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.
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.
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
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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.
From Biochemistry and Molecular Biology of
Plants (W.Gruissem, B. Buchanan and R.Jones
p.972 ASPP, Rockville MD, 2000
Slide 4.6
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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.
From Biochemistry and Molecular Biology of
Plants (W.Gruissem, B. Buchanan and R.Jones
p.851 ASPP, Rockville MD, 2000
Slide 4.7A
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III. Long-distance signalling - The Major Plant
Hormones
Hormone Synthesized where? Function/Notes Auxin
leaf primordia apical dominance young
leaves root induction vascular
tissue development stimulates
fruit development many
others Cytokinins root tips cell
division shoot formation Ethylene rip
ening or fruit ripening senescence
tissues tissue senescence Abscisic acid mature
leaves, stomatal closure especially
after embryogenesis water stress induces
sugar transport from leaves to
seeds induces storage protein
synthesis in seeds Gibberellins young
shoots and elongation of
shoots developing seeds 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
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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.
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
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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 6.9
Slide 4.8
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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