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Neurogenesis and cell fate specification: the role of
transcriptional regulatory networks
Joe Corbo
Washington University School of Medicine
([email protected])
Unicellular organism
Unicellular organism
Multicellular organism
Proliferation and differentiation
Fertilized egg
Diverse cell types
Cellular differentiation on an epigenetic landscape
Fertilized
egg
Differentiated cell types
Why do mammals have so few cell types?
- A cell type is defined by the complement of genes it expresses.
- ~25,000 genes in the mammalian genome.
- Let’s assume that each gene can be either ON or OFF.
- A cell type consists of a specific combination of ON and OFF genes.
- Thus, the genome permits 225,000 possible cell types.
- But mammals have only several hundred cell types. Why?
Cell type #1
Cell type #2
Cell type #3
The number of cell types is constrained by the
structure of transcriptional regulatory networks
cis-regulatory element
coding sequence
Today’s lecture
Part 1:
Neurogenesis and
cell fate specification
Part 2:
Transcriptional regulatory
networks
Photoreceptors are the entry
point of our visual system
The retina has seven basic cell classes
Rod
Cone
Horizontal cell
Bipolar cell
Amacrine cell
Ganglion cell
How to define a cell type?
-molecules
-morphology
-connectivity
(Mueller glia)
not depicted
The retina contains as many as 80
different cell types
Data for mouse retina
Cell classes
cell types
Rods
Cones
Horizontal cells
Bipolar cells
Amacrine Cells
Ganlion cells
Mueller glia
total # of cell types:
1
2
1
15
30
30
1
80
retina
Generation of neuronal diversity
in the developing retina
pupil
ONL
INL
GCL
Embryonic retina
Adult retina
light
sensory
information
Proliferation and differentiation
Retinal progenitor
Seven retinal cell classes
ONL
INL
GCL
Embryonic retina
Adult retina
light
sensory
information
The cell cycle in the developing retina
Interkinetic nuclear migration
Three types of
cell division:
Symmetric
mitotic
Asymmetric
Symmetric
post-mitotic
Proliferation in the developing retina
Q fraction: proportion of
cell division progeny exiting
the cell cycle (“Quitting fraction”)
P fraction: proportion of
cell division progeny continuing
To proliferate
Proliferation in the developing brain
Q fraction: proportion of
cell division progeny exiting
the cell cycle (“Quitting fraction”)
P fraction: proportion of
cell division progeny continuing
To proliferate
Retinal cell types are born in overlapping waves
Cell number
Rod
Bipolar
Amacrine
Gang.
Cone
Müller glial
P11
P9
P7
P5
P3
P1
E18
E16
E14
E12
E10
Horiz.
Birth date
Adapted from Young, 1985.
Cell lineage in the retina
Three types of
cell division:
Symmetric
mitotic
Asymmetric
Example cell lineage:
Symmetric
post-mitotic
Final output:
2 ganglion cells
1 horizontal cell
5 rods
1 bipolar cell
The competence model of retinal development
Competence model: progenitors pass through a
series of competence states, during each of
which the progenitors are competent to produce
a subset of retinal cell types.
Environmental factors:
(1) Small molecules
(2) Secreted proteins
(3) Cell surface molecules
Progenitor
Etc.
Competence
•
Time
Differentiated cell types
Today’s lecture
Part 1:
Neurogenesis and
cell fate specification
Part 2:
Transcriptional regulatory
networks
What is a transcription factor?
A transcription factor is a protein with two parts:
(1) A sequence-specific DNA-binding domain
(2) An activation or repression domain
Activation
domain
Transcription
factor
DNA-binding
domain
DNA
What is a transcription factor?
A transcription factor is a protein with two parts:
(1) A sequence-specific DNA-binding domain
(2) An activation or repression domain
Activation
domain
Transcription
factor
DNA-binding
domain
DNA
CTAATCCC
A transcription factor can bind a family of related sequences
CRX:
CTAATCCC
CAAATCCC
CTAATCGC
CTAAGCCC
CAAATCCC
CTAAGCGC
etc.
A transcription factor can bind a family of related sequences
CRX:
CTAATCCC
CAAATCCC
CTAATCGC
CTAAGCCC
CAAATCCC
CTAAGCGC
etc.
~1,000 TFs in human genome and they fall into various families
http://www.bioguo.org/AnimalTFDB/images/pic_family.jpg
How does transcriptional
regulation work?
brain
cis-regulatory
elements (CREs)
are in blue
retina
coding sequence
limbs
= transcriptional
activators
300-600 bp
Transcription factors control:
-spatial pattern
-timing
-levels
= transcriptional
repressor
Transcriptional regulatory networks
cis-regulatory element
coding sequence
Photoreceptors are the entry
point of our visual system
Fertilized
egg
Photoreceptor
precursor cell
rod
cone
“The complex system of interactions
underlying the epigenetic landscape”
photoreceptor
Crx
Nrl
Nr2e3
photoreceptor
Gene networks in photoreceptor development: Otx2
Crx probe
•
•
•
Otx2 is a homeodomain-containing
transcription factor (TF)
One of the first TFs to be expressed in
developing photoreceptors
When mutated, Crx expression is lost and
photoreceptors fail to differentiate
Wt
Wt
Otx2-/-
Otx2-/Otx2
Crx
Furukawa et al.,
Nat. Neurosci. 6: 1255-1263
Gene networks in photoreceptor development: Crx
Otx2
•
•
•
•
Crx is a homeodomain-containing
transcription factor (TF) similar to Otx2
One of the first TFs to be expressed in
developing rods and cones
When Crx is mutated photoreceptors fail to
differentiate normally; causes several forms
of blindness in humans
Expression of both rod-specific and conespecific genes is affected
Crx
Rod-specific
genes
Cone-specific
genes
Furukawa et al.,
Nat. Genet. 23: 466-470
Gene networks in photoreceptor development: Nrl
•
•
•
Otx2
Nrl is a leucine zipper transcription
factor
Nrl is expressed only in rods, not in
cones
It is required for activation of rod
genes and repression of cone genes
Crx
Nrl
Rod-specific
genes
Cone-specific
genes
Furukawa et al.,
Nat. Genet. 23: 466-470
Wild-type rod
ON
OFF
Nrl mutant “rod”
OFF
ON
Gene networks in photoreceptor development: Nrl regulation
•
Otx2
Nrl is directly regulated by at least
three upstream transcription factors:
Otx2, Crx and Rorb
Rorb
b
Crx
Nrl
Rod-specific
genes
Cone-specific
genes
Furukawa et al.,
Nat. Genet. 23: 466-470
Gene networks in photoreceptor development: Nr2e3
•
•
•
•
•
Otx2
Nr2e3 a member of the nuclear hormone
receptor superfamily
Nr2e3 is only expressed in rods, not cones
Nrl activates expression of Nr2e3
Nr2e3 represses cone genes in rods
It also activates rod genes
Rorb
b
Crx
Nrl
Nr2e3 probe
Wt
Nrl-/Pnr
Nr2e3
Rod-specific
genes
Cone-specific
genes
Wild-type rod
ON
OFF
Nr2e3 mutant “rod”
ON
ON
Transcriptional networks as the driving force behind
the generation of neuronal diversity
Otx2
Rorb
b
Crx
Nrl
Retinal
progenitor
Pnr
Nr2e3
Retinal
Cell types
Rod-specific
genes
Cone-specific
genes
Differentiated cell types occupy defined position in the ‘state space’
Of the cell’s transcriptional regulatory network
Embryonic
stem cell
Photoreceptor
precursor cell
rod
cone
Today’s lecture
Part 1:
Neurogenesis and
cell fate specification
Part 2:
Transcriptional regulatory
networks
[email protected]
http://pathology.wustl.edu/~corbolab/