Transcript (bacteria) (Cytokinin signaling in plants)

Hormonal Signaling II: Cytokinin and Ethylene
1. Cytokinin
1) overview: Zeatin (a purine derivative) is most abundant natural cytokinin,
discovered as a DNA breakdown product that stimulate cell
division/growth. It also inhibits aging/senescence, ratio of cytokinin/auxin
regulate organogenesis (higher cytokinin favors shoot formation)
2) Signaling pathways
A) The receptor is similar to the bacterial two-component proteins
The first study was performed using an activation tagging method: the idea
is to activate randomly the genes in the genome by inserting a strong
promoter anywhere in the genome and screen for phenotype that is
related to cytokinin action. IN this case, shoot production from callus
was used (more cytokinin favors shoot formation) to select those
callus that can form shoot without cytokinin!
One such gene called CKI1 (CytoKinin Independent) was isolated—when
this gene is expressed at high level, the callus can form shoot without
cytokinin. The gene encodes a protein with similarity to the histidine
kinase and response regulator in bacteria. This is the most
Popular receptor system in bacteria
to detect environmental signals
receptor
Such as nutrition, temperature,
And chemotaxis signals.
receptor
Action in cytokinin response and the fact that is is similar to a bacterial
receptor suggests that it may be a receptor of cytokinin. Further evidence
came from study of another protein CRE1. The gene was identified by
mutant screening—cre1 loss-of-function mutant was not sensitive to
cytokinin and its callus does not produce shoot even in the presence of
cytokinin! This gene also encode a two component-like receptor very
similar to CKI1. In addition, CRE1expressed in yeast cells can bind
cytokinin.
Further study of function of CRE1 and CKI1 was performed in a protoplast
system: express the receptor and a reporter gene in the protoplast and see if
the receptor activation can turn the reporter gene on or off. Indeed both
CRE1 and CKI1 can serve as a receptor for cytokinin to activate the gene
expression.
The cytokinin receptor appears to be a fused version of
The two component system: the essence of this system is to pass the
signal by transferring the phosphate group from a histidine (H) to
aspartate (D)
(bacteria)
(Cytokinin signaling in plants)
B) Downstream from the receptor
Early response genes of cytokinin turn out to be encoding the response
regulator-like proteins. Further study using the protoplast and transgenic plant
models showed that these RR proteins participate in cytokinin signaling.
Detailed picture is still not clear.
The RR proteins are divided into two families: A type and B type that serve as
negative and positive regulator of cytokinin response. In other words, B type
activate and A type inhibits the cytokinin response.
RR gene expression is activated by cytokinin
Time course after cytokinin treatment
Summary pathway (part I)
1.
2.
3.
Cytokinin binding to
receptor that may
dimerize
Phosphate transfer
from H to D due to
activation of the
histidine kinase
activity
The phosphate transfer
to another protein
called AHP (for
arabidopsis histidine
phosphotransfer)
Summary pathway (part II)
4. Phosphotransfer to
the RR protein,
activating RR
5. B-type RR protein
activate gene expression
6. A-type RR inhibits
gene expression
7. All these events
combine into cytokinin
response at cellular and
whole plant level
2. Ethylene
1) OVERVIEW: gaseous hormone was discovered when growing
plants under gas lamp (triple response), synthesis induced by stress
and pathogen, causing aging and abscision, fruit ripening, etc
Inhibitor of C2H4
synthesis (left) or mutant
of C2H4 response delays
senescence of the flowers
and abscision of leaves
(right).
Arabidopsis
Triple response:
Shorter hypocotyl
Shorter roots
Exaggerated apical
hook
2) Signaling
A) Receptors were identified by genetics and biochemical approaches
The mutant selection: ethylene resistant (or insensitive) and constitutive
ethylene response mutants
Etr1—ethylene resistant-1: mutant plants are not responsive to ethylene. See
all, but one plant, are sensitive to ethylene with triple response. That one
plant is etr1
The ETR1 gene encode a receptor similar to the
cytokinin receptor, the two-component histidine
kinase/response regulator fused together.
Several other genes are identified in similar
genetic screen or simply by comparing the
Sequence similarity. All of them appear to
have extracellular C2H4 binding domain and
transmembrane domains. Some of them have
both the kinase domain and the receiver domain
but others lack one of the domains (See next page)
The ethylene
Receptor family
ETR1 is a true ethylene receptor with the following properties
i) ETR1 wild type protein binds C2H4 when expressed in yeast
but etr1 mutant protein lacks such binding activity
ii) Binding of C2H4 to the ETR1 requires a metal cofactor (Cu).
iii) Dimerization may be conducted by disulfide bonds
B) Downstream from receptor may be a kinase cascade
CTR1—ctr1 mutant is constitutive response mutant—always show triple
response even without ethylene
What is CTR1 gene? Encoding a protein kinase like Raf1 in animal cells
(the kinase that activates MAPK kinase…)
CTR1--Raf-like kinase
MEK-like kinase
MAPK
EIN3—ein3 is a ethylene insensitive mutant –
the gene encoding a transcription factor in the nucleus.
The pathway summary:
The RAN1 protein is essential for
assemble the Cu cofactor with the
receptor for C2H4 binding.
IN the absence of C2H4, the receptor
would activate the downstream kinase
CTR1 that in turn inhibits the
response/gene expression. In the
presence of C2H4, the receptor is
“inhibited” so is the CTR1 kinase. The
response is now “on”. The kinase
cascade may serve as a negative
regulator of the gene expression.
What is the evidence
that receptor is inhibited
by C2H4? And CTR1
is a negative regulator?