Transcript Using mouse genetics to understand human disease

Using mouse genetics to
understand human disease
Mark Daly
Whitehead/Pfizer
Computational Biology
Fellow
What we do
• Genetics: the study of the inheritance of
biological phenotype
– Mendel recognized discrete units of inheritance
– Theories rediscovered and disputed ca. 1900
– Experiments on mouse coat color proved
Mendel correct and generalizable to mammals
– We now recognize this inheritance as being
carried by variation in DNA
Why mice?
• Mammals, much better
biological model
• Easy to breed, feed, and
house
• Can acclimatize to
human touch
• Most important: we can
experiment in many
ways not possible in
humans
What do they
want with me?
Mice are close to humans
Kerstin Lindblad-Toh
Whitehead/MIT Center for Genome Research
Mouse sequence reveals great
similarity with the human genome
Extremely high conservation: 560,000 “anchors”
Mouse-Human Comparison
both genomes 2.5-3 billion bp long
> 99% of genes have homologs
> 95% of genome “syntenic”
Genomes are rearranged copies
of each other
Roughly 50% of bases change in the evolutionary time from mouse to human
Mouse sequence reveals great
similarity with the human genome
Extremely high conservation: 560,000 “anchors”
Anchors (hundreds of bases with >90% identity)
represent areas of evolutionary selection…
…but only 30-40% of the highly conserved
segments correspond to exons of genes!!!
What we can do
• Directed matings
• Inbred lines and crosses
•
•
•
•
Knockouts
Transgenics
Mutagenesis
Nuclear transfer
• Control exposure to
pathogens, drugs, diet, etc.
YIKES!!!
Example: diabetes related mice
available from The Jackson Labs
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•
•
•
•
Type I diabetes (3)
Type II diabetes (3)
Hyperglycemic (27)
Hyperinsulinemic (25)
Hypoglycemic (1)
Hypoinsulinemic (5)
Insulin resistant (30)
• Impaired insulin
processing (7)
• Impaired wound
healing (13)
Inbreeding
• Repeated brothersister mating leads to
completely
homozygous genome
– no variation!
Experimental Crosses
• Breed two distinct inbred lines
• Offspring (F1) are all genetically identical –
they each have one copy of each
chromosome from each parent
• Further crosses involving F1 lead to mice
with unique combinations of the two
original strains
Experimental Cross
Experimental Cross: backcross
• F1 bred back to one of
the parents
• Backcross (F1 x RED)
offspring:
50% red-red
50% red-blue
Experimental Cross: F2 intercross
• One F1 bred to
another F1
• F2 intercross (F1xF1)
offspring:
25% red-red
50% red-blue
25% blue-blue
F2
Trait mapping
F2
100
200
300
Trait mapping
Blue trees = tall, Red trees = short
In the F2 generation, short trees tend to carry “red” chromosomes where the
height genes are located, taller trees tend to carry “blue” chromosomes
QTL mapping use statistical methods to find these regions
How do we distinguish
chromosomes from different strains?
• Polymorphic DNA markers such as Single
Nucleotide Polymorphisms (SNPs) can be
used to distinguish the parental origin of
offspring chromosomes
ATTCGACGTATTGGCACTTACAGG
ATTCGATGTATTGGCACTTACAGG
SNP
Example: susceptibility to Tb
% survival
100
• C3H mice extremely
susceptible to Tb
• B6 mice resistant
B6
50
C3H
0
0
100
200
Days post infection
300
• F1, F2 show
intermediate levels of
susceptibility
One gene location already known
B6
% survival
100
C3H
50
C3H.B6-sst1
0
0
50
100
Survival Time
150
200
• Previous work
identified
chromosome 1 as
carrying a major
susceptibility factor
• Congenic C3H
animals carrying a
B6 chromosome 1
segment were bred
Congenic and consomic mice
• Derived strains of mice in which the
homozygous genome of one mouse strain has a
chromosome or part of a chromosome
substituted from another strain
C3H
Chr 1
Chr 2
Chr 3
Chr 4
Etc.
B6
C3H.B6_chr1
Tb mapping cross
F2 intercross:
C3H.B6sst1
x
x
B6
C3H.B6-sst1 - MTBsusceptible, carrying B6 chr
1 resistance
F1
B6 - MTB-resistant
Trait – survival following
MTB infection
n = 368
…
F2
Results: 3 new gene locations
identified!
Gene identified on chromosome 12
100
100
bh
% survival
% survival
bb
hh
50
Mice engineered to be
missing a critical component
of the immune system
Days after infection
located in
this region are
C57Bl/6J
B.likewise more susceptible, C.
B6-Igh6
validating that particular
gene as involved in Tb
susceptibility
0
0
0
100
200
days post infection
A.
50
Chi square
18.99
df
2
P value
P<0.0001
300
0
25
50
75
100
125
150
100
% survival
At the end of chr 12 – mice
inheriting two C3H copies
survive significantly longer
than those
with one or two
B6-IL12-/B6 copies
50
0
0
25
50
75
100
125
150
Days after infection
Chi square
30.02
Chi square
20.17
df
2
df
1
P value
P<0.0001
P value
P<0.0001
BALB/cBJ
BALB/c-mMT-/-
Mouse History
• Modern
“house mice”
emerged from
Asia into the
fertile crescent
as agriculture
was born
Mouse history
Recent mouse history
Fancy mouse breeding - Asia, Europe
(last few centuries)
Retired schoolteacher Abbie Lathrop
collects and breeds these mice
Granby, MA – 1900
Castle, Little and
others form most
commonly used
inbred strains
from Lathrop stock
(1908 on)
W.E. Castle
C.C. Little
Mouse history
Mouse history
• Asian musculus and European domesticus
mice dominate the world but have evolved
separately over ~ 1 Million years
• Mixing in Abbie Lathrop’s schoolhouse
created all our commonly used mice from
these two distinct founder groups
Genetic Background of the
inbred lab mice
musc
C3H
DBA
domest
domest
domest
cast
musc
musc
domest
{
C57BL/6
domest
musc
Avg segment size ~ 2 Mb
<1 SNP/10 kb
{
{
Comparing two inbred strains –
frequency of differences in 50 kb segments
~40 SNP/10 kb
Finding the genes responsible for
biomedical phenotypes
20 Mb
C3H (susceptible)
B6 (resistant)
Traditionally: positional cloning is painful
(e.g., generating thousands of mice for fine mapping, breeding congenics) –
As a result, countless significant QTLs have been identified in mapping
crosses but only a small handful have thusfar resulted in identification
of which gene is responsible – the critical information that will advance
research into prevention and treatment!
Using DNA patterns to find genes
20 Mb
C3H (susc.)
B6 (res.)
Critical Region
Using DNA patterns to find genes
20 Mb
C3H (susc.)
B6 (res.)
DBA (susc.)
Critical Region
Example: mapping of albinism
Critical region
First genomic region mapped
129S1
T
A
*
C
C
C
*
C
G
G
T
A
C
G
A
G
G
G
AKR
A
G
T
T
T
A
A
T
G
G
T
A
C
G
A
G
G
G
A_J
A
G
T
T
T
A
A
T
G
G
T
A
C
G
A
G
G
G
BALB_c
T
A
*
C
C
C
G
C
G
G
T
A
C
G
A
G
G
G
C3H
A
G
T
T
T
A
A
T
G
G
T
A
C
G
A
G
G
G
C57B6
A
G
T
T
T
A
A
T
C
T
A
G
T
A
C
C
C
A
CBA
A
G
T
T
T
A
A
T
C
T
A
G
T
A
C
C
C
A
DBA2
A
G
T
T
T
A
A
T
C
T
A
G
T
A
C
C
C
A
FVB
A
G
T
T
T
A
A
T
C
T
A
G
T
A
C
C
C
A
I
A
G
T
T
T
A
A
T
G
G
T
A
C
G
A
G
G
G
NOD
A
G
T
T
T
A
A
T
G
G
T
A
C
G
A
G
G
G
NZB
*
A
C
C
C
C
*
C
C
T
*
G
T
A
C
C
C
A
SJL
A
G
T
T
T
A
A
T
C
T
A
G
T
A
C
C
C
A
SWR
A
G
T
T
T
A
A
T
C
T
A
G
T
A
C
C
C
A
Chr 4
(Mb)
35.7
37.6 37.9
39.4
Future Genetic Studies
Mapping
Expression
Pathways
Model Systems
Thanks to
(Whitehead Institute)
Claire Wade
Andrew Kirby
(MIT Genome Center)
EJ Kulbokas
Mike Zody
Eric Lander
Kerstin Lindblad-Toh
Funding:
Whitehead Institute
Pfizer, Inc.
National Human Genome Research Institute