Download Testing the ABC floral-organ identity model

Testing the ABC floral-organ
identity model: expression of A
and C function genes
Objectives:
To test the validity of the ABC model for floral organ identity we will:
1.
Use the model to make predictions concerning the phenotype of double or triple loss-offunction mutants and compare with the actual double mutant phenotypes.
2.
Clone and sequence the ABC genes. Look for similarities with sequenced genes already in
the database.
3.
Determine the time and place of expression for each ABC gene and consider whether
the expression correlates with the functional domain defined by the loss-of-function
phenotype.
4.
Test regulatory interactions between ABC genes by examining how the loss-of-function
of one gene affects the expression domain of another.
5.
Create gain-of-function mutants by generating transgenic plants carrying an ABC gene
cDNA under the control of the CaMV35S promoter.
Arabidopsis Floral Development
Inflorescence SEM
Mature Flower
Flower Development
Arabidopsis Floral Development
Inflorescence SEM
Inflorescence Section
Arabidopsis Floral Development
Inflorescence SEM
Inflorescence Section
in situ hybridization
A Model For Control of Organ
Type
sepal
petal
1
2
stamen
carpel
4
3
B (AP3, PI)
A (AP1, AP2)
C (AG)
AG expression in wild type
Inflorescence SEM
Inflorescence Section
in situ hybridization
AG expression in wild type
AG expression in wild type
Mature Flower
Inflorescence Section
in situ hybridization
A Model For Control of Organ
Type
sepal
petal
1
2
stamen
carpel
4
3
B (AP3, PI)
A (AP1, AP2)
C (AG)
AP1 expression in wild type
Inflorescence SEM
Inflorescence Section
in situ hybridization
AP1 expression in wild type
AG expression
AP1 expression
A Model For Control of Organ
Type
sepal
petal
1
2
stamen
carpel
4
3
B (AP3, PI)
A (AP1, AP2)
C (AG)
AP2 expression in wild type
Inflorescence SEM
Inflorescence Section
in situ hybridization
AP2 expression in wild type
A Model For Control of Organ Type
Ap2 mutant
carpel
stamen
stamen
carpel
1
2
3
4
B (AP3, PI)
C (AG)
Where will AG be expressed?
AG expression in an Ap2 mutant
A Model For Control of Organ Type:
Ag mutant
sepal
petal
petal
sepal
1
2
3
4
B (AP3, PI)
A (AP1, AP2)
Where will Ap1 be expressed?
AP1 expression in an Ag mutant
A Model For Control of Organ Type:
Ag mutant
sepal
petal
petal
sepal
1
2
3
4
B (AP3, PI)
A (AP1, AP2)
Where will Ap2 be expressed?
AP2 expression in an Ag mutant
(unchanged)
A Model For Control of Organ Type:
35SAG
?
sepal
?
petal
1
2
?
stamen
?
carpel
4
3
B (AP3, PI)
A (AP1, AP2)
C (AG)
C (AG)
Structure of wild type and mutant
Arabidopsis flowers
WT
Whorl 1
Whorl 2
Whorl 3
Whorl 4
SEPAL
PETAL
STAMEN
CARPEL
?
STAMEN/
no organ
STAMEN
STAMEN
CARPEL
CARPEL
35S E ?
Ag O SEPAL/
CARPEL
35S E
AP2 O
35S E
AP1 O
A Model For Control of Organ Type:
35SAP2 or 35SAP1
?
sepal
?
petal
1
2
?
stamen
?
carpel
4
3
B (AP3, PI)
A (AP1, AP2)
C (AG)
A (AP1 OR AP2)
Structure of wild type and mutant
Arabidopsis flowers
Whorl 1
Whorl 2
Whorl 3
Whorl 4
SEPAL
PETAL
STAMEN
CARPEL
35S E ?
AG O SEPAL/
CARPEL
35S E SEPAL
AP2 O SEPAL
?
STAMEN/
no organ
PETAL
PETAL
STAMEN
STAMEN
CARPEL
CARPEL
STAMEN
STAMEN
CARPEL
CARPEL
35S E SEPAL
AP1 O SEPAL
PETAL
PETAL
?
STAMEN
?
CARPEL
WT
A Model For Control of Organ Type:
35SAP2 or 35SAP1
?
sepal
?
petal
1
2
?
stamen
?
carpel
4
3
B (AP3, PI)
A (AP1, AP2)
C (AG)
A (AP1 OR AP2)
Review of: Analysis of Class A and C expression.
1.
In wild type plants, AP1 transcript is first observed throughout the stage 1 floral
meristem but disappears from the centre of the stage 3 floral meristem. In later
stages it continues to be expressed in developing sepals and petals.
2.
In wild type plants, AP2 transcript is observed throughout the stage 1 floral meristem
and continues to be found in all whorls in later stages of floral development.
3.
In wild type plants, AG transcript is first observed in the central region of the stage 3
floral meristem. In later stages it continues to be expressed in developing stamens
and carpels.
4.
In Ap2 mutants, AG transcript appears in late stage 2/early stage 3 throughout the
floral meristem although AG is most strongly expressed in the center of the floral
meristem.
5.
In Ag mutants AP1 transcript persists throughout the developing flower past stage
three. The pattern of AP2 transcript in an Ag mutant is the same as in wild type.
6.
Ectopic expression of AP2 or AP1 using the CaMV35S promoter does not affect
floral organ type.
Hypothesis for A-C function interactions
1.
AP1 and AP2 appear early and are needed to specify perianth.
2.
AG appears in stage 3 and negatively regulates expression of AP1 in
inner cells and promotes formation of reproductive organs.
3.
AP1 and AP2 prevent AG expression in perianth.
4.
At least one other regulator must exist to promote AG expression in the
centre at stage 3.