Chromatin remodelling ATPase Brg1 is an essential factor for the maintenance of the intestinal crypt stem cell and adenoma
formation
Aliaksei Holik
DNA is packed into Chromatin
Nicolas Bouvier, Curie Institute, Paris, France
Chromatin remodelling complexes enable access for
DNA binding proteins
Transcription factors
OFF
ON
Chromatin remodelling
complex
ATP
BRG1 and cancer
BRG1 mutations are found in cancer cell lines and in primary cancers
Heterozygous deletion of Brg1 in mice promotes tumourigenesis due to Brg1 haploinsufficiency
Restoration of BRG1 function leads to the reversion of a cancer phenotype
c-Fos
BRG1
BRCA1 RB/p21
p53
Wnt-pathway
Suggested mechanisms of BRG1 mediated tumourigenesis
Wnt pathway
Willert K., Jones K. A. Genes Dev. 2006;20:1394-1404
Small intestine architecture
Villus
Crypt
Stem cell
Transit amplifying cells
loxP strategyGeneration of conditional knock-out using Cre-loxP techniques
X BRG1
Cre recombinase Ah
loxP loxP
Mouse 1 Mouse 2
Cre recombinase Ah
BRG1
loxP loxP
Cre recombinase Ah
ER
ER
ER
Brg1-deficient cells in small intestine are repopulated with wild-type cells
Timeline of gut repopulation
0
0.1
0.2
0.3
0.4
0.5
0.6
3 5 7 10 14
Days Post Induction
Rat
io o
f re
com
bin
ed c
ryp
ts
Day 3 Day 5 Day 7
Brg1-deficient small intestinal epithelium appears to be normal at day 4 post induction
Brg1fl/fl
Control
-BRG1
Brg1-deficient small intestinal epithelium displays lower apoptosis levels
Apoptosis per Crypt
0.00
0.50
1.00
1.50
2.00
2.50
3.00
Control Brg Hom
BrdU Positive Cells Per Half Crypt
0.00
2.00
4.00
6.00
8.00
10.00
12.00
Control Brg Hom
Inactivation of Brg1 in small intestinal epithelium leads to rapid crypt loss
Brg1fl/fl
Control
Day 3 Day 5 Day 7
Brg1 deletion leads to the failure of the crypt stem cell
Active stem cell
Brg1+ Brg1+Brg1-
Generation of double knock-out using Cre-loxP techniques
X BRG1
Cre recombinase Ah
loxP loxP
Mouse 1 Mouse 2
Cre recombinase Ah
BRG1
loxP loxP
Cre recombinase Ah
ER
ER
ER
APC
loxP loxP
APC
loxP loxP
Brg1/Apc double mutant cells are selectively eliminated in small intestine
Small intestine of Brg1fl/fl/Apcfl/fl mouse at day 7 post induction
-BRG1--catenin
Brg1-deficiency rescues lethal phenotype of Apc deletion in double knock-out mice
Control
Brg1fl/flAPCfl/fl
APCfl/fl
Brg1fl/fl
Median survival APCfl/fl=8 Brg1fl/flAPCfl/fl=14, P=0.0010
Brg1 removes a portion of Apc-deficient cells decreasing tumour burden
Brg1-/Apc-
Brg1- Apc-
Brg1 removes a portion of Apc-deficient cells decreasing tumour burden
Brg1-/Apc-
Brg1- Apc-
Microadenoma
Adenoma
Stem cel
l
Transit amplifying cells
Adenoma progression depends on the cell of origin
Conclusions
Brg1 is a crucial factor for the maintenance of the adult tissue stem cell in small intestine. This is the first evidence of Brg1 requirement for the somatic stem cell maintenance
Targeting the active sub-population of intestinal stem cells provide an attractive concept for the removal of potentially malignant lesions in individuals with genetic predisposition to colorectal cancer
Acknowledgements
The Darwin Trust of Edinburgh
Alan Clarke
Boris Shorning
ARC group