- 1. Mouse Models of Human Disorders Prepared by Rohan Palmer
Institute of Behavioral Genetics University of Colorado at Boulder
[email_address]
2. Goals
- Understanding Genetic Influences on a Trait
-
- Accounting for Variation in an Observed Trait
-
- Response to Selection Study
- Types of Mouse Models of Human Disorders
3. Lecture Goal Exam Question
- You are reading a Times article with a friend and you come
across the
- The study hypothesized that some of these susceptibility
factors may be
- allelic variants of genes that govern embryonic serotonin
neuron
- development and that these alleles may contribute to behavioral
disorders
- by adversely increasing or decreasing serotonin system
activity.
- The article is about finding ways to create mouse models for
human DCGs.
- Your classmate has not yet taken Dr. Careys Behavioral Genetics
course but
- fortunately you have.Describe to him/her what an allelic
variant is and at
- how it might affect a behavioral disorder. Lastly, your
friendlooks at you as if
- you were crazy and says, How does a mouse study, helpUS? Based
on what
- you have taken away from this course, convince your friend
about the
- relevance and validity of mouse models. (20 Points)
4. Heritability
- The extent to which variation on a trait in a
- given population at a given point in time is
- attributable to genetic variation between
5. Comparison Terminologies
- Synteny: The co-localization of genes on
- chromosomes of related species.
- Homolog: The situation where nucleic
- acid or protein sequences are similar
- because they have a common
- evolutionary origin. Oftenused loosely
- to indicate that sequences are very
- Ortholog gene sequences are similar
- Paralog gene sequences are similar
Adapted from the Department of Energy; genomics.energy.gov 6.
Why Create Animal Models?
- No human genetics ethical dilemma
- (Avg. gestational period is 19 days)
- (Females become fertile in 3 weeks)
- (Approximately 40 days for turnover; giving approximately
365/40 generations annually)
- Creating a representative phenotype is difficult
- Ethical dilemma (Animal Rights; Institutional Animal Care and
Use Committee (IACUC)
7. 8. Mouse Genome Informatics 9. Determining Genetic Influences
on a Trait
- Response to 30 generations
- of selection for Open-Field
- Activity in Lab Mice(DeFries et al. 1978)
- Strain A collective of animals from the same species that are
genetically identical.
- Response to selection: DeFries et al. 1978
- It is possible to observe variation within and between strains
of mice.
- Variation on a measured trait within a stain is indicative of
environmental influences on that trait.
- Variation on a measured trait between strains is indicative of
genetic influences on that trait.
- For eg. The A/J inbred strain is susceptible to arthritis while
the C57BL/6J strain is resistant. (Source: Adapted from
Bioinformatics and Functional Genomics, p. 685)
- C57BL/6J mice consume far greater amounts of alcohol than
DBA/2J mice.
10. Why does Emotionality in mice matter?
-
- Includes anxiety and neuroticism
-
- Think about what happens to you when you are anxious?
-
- Exploration, activity, anxiety, fear and
emotionality/reactivity are measured in an open field apparatus
(OFA)
11. Why does Emotionality in mice matter?
-
- Includes anxiety and neuroticism
-
- Think about what happens to you when you are anxious?
-
- (Sweaty, elevated heart rate, preoccupied thoughts, urge to
defecate, involuntary movement, Immobile)
-
- Low activity & high defecation scores define emotionality;
measures are genetically correlated
-
- Animals with high defecation scores are inactive
12. Breeding & Selection Experiment 1 st10 litters H1 line
L1 line 2 nd10 litters H2 line L2 line 3 rd10 litters C1 line 4
th10 litters C2 line BalbF1 F1 F2 F2 B6 F30 x x x The entire
breeding experiment used a total of 14,184 mice 13. Differences
within and between strains 14. Differences btw Selected Lines 15.
Peculiar Association? 16. What Can We Do With Mouse Models 17.
Database of Mouse Models 18. Disease Genes and Polymorphisms
- Typically transgenic mice are used to show how the
over-expression of a gene product affects physiology, behavior,
etc.
- Relatively straightforward procedure.
- Transgenic mouse can be made in 3-6 months.
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- Transgenesis: The transfer of human genes into animals or
cultured cells, esp. to produce molecules with therapeutic
value.
- Mutagenesis: The occurrence or induction of mutation
19. Disease Genes and Polymorphisms
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
Adapted from Hedari Laboratory, Wayne State Univ. 20. Disease
Genes and Polymorphisms
- Alzheimers Transgenic Mice
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
Pictures of Alzheimer's Pathology Stain: Campbell-Switzer Silver
MethodExcert from: Neuroscience Associates Labs
http://www.neuroscienceassociates.com/Stains/alzheimers.htm The APP
transgenic mouse has been the most popularly used model with
neuritic plaques conspicuously displayed by the age of 12-13 months
of age The development of transgenic mice that are doubly
transgenic for APP and presenilin-1 (PS-1) yielded mice that
developed plaques at a much earlier age than the singly transgenic
APP mouse. 21. Disease Genes and Polymorphisms
-
- Integration of transgene is random.
-
-
- It may disrupt the function of another gene
-
-
- May integrate into a part of the genome where gene expression
is suppressed.
-
-
- May integrate into a part of the genome under the control of
alocus control region .
-
-
-
- Number of copies cannot be controlled.
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- Locus Control Region (LCR) region of the chromosome that is
responsible for the expression of linked genes to physiologically
appropriate levels in specific tissues at developmentally
appropriate time points.
22. Disease Genes and Polymorphisms
-
- Localization of expression cannot be controlled
completely.
-
- Over time, the transgene is frequently shut off.
-
- These issues can be overcome by generating several transgenic
lines using the same construct and comparing the data across
lines.
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
23. Disease Genes and Polymorphisms
- Targeted mutant mice Alteration of gene of interest through
homologous recombination
- The gene of interest can be eliminated (knock-out or null
mutant) or altered (knock-in).
- Loss of gene function or alteration of function is typically
evaluated in KO and KI mice
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- Mutagenesis: The occurrence or induction of mutation
24. Disease Genes and Polymorphisms
- The gene of interest can be conditionally deleted or
altered.
- Very labor intensive and technically challenging methods are
involved.
- Typical time to generate a null-mutant mouse is 1-3 years.
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
25. Disease Genes and Polymorphisms
- Knock-out: Remove gene or exon of gene that renders the gene
product non-functional.
- Knock-in: Replace the natural gene or exon with an altered gene
or exon that alters the function of the gene product.
- Congenic*:Introgression of a locus of interest from another
strain; selecting for a given marker in the donor strain.
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- -Knock-out/ Knock-in / Conditional
- Knock-out/ Inducible expression /
- Introgression: Infiltration of the genes of one species into
the gene pool of another through repeated backcrossing of an
interspecific hybrid with one of its parents.
26. Disease Genes and Polymorphisms
- Conditional KO: Gene is modified so that it is deleted only
under specific conditions.
- Inducible/Tissue specific expression: Gene expression is
regulated by a drug supplement (typically a chemical not endogenous
to the mouse).
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- -Knock-out/ Knock-in/ Conditional
- Knock-out/ Inducible expression
27. Disease Genes and Polymorphisms
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- -Knock-out/ Knock-in / Conditional
- Knock-out/ Inducible expression
- Chimera - an animal that has two or more different populations
of genetically distinct cells that originated in different
zygotes
Adapted from the Fred Hutchinson Cancer Research Center 28.
Disease Genes and Polymorphisms
- Gene targeting in the mouse
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- -Knock-out/ Knock-in / Conditional
- Knock-out/ Inducible expression
29. Disease Genes and Polymorphisms
- Problems with KOs and KIs
- Targeting vectorcomponents (such as the marker used) may affect
the expression of genes close to the mutated gene.
- Potential developmental effects, such as:
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- -Knock-out/ Knock-in / Conditional
- Knock-out/ Inducible expression
30. Disease Genes and Polymorphisms
- Problems with KOs and KIs
- Genetic background: No behavioral trait is determined by a
single gene.
- It is expensive and labor intensive.
- Mice take a long time to create.
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- - Knock-out/ Knock-in / Conditional
- Knock-out/ Inducible expression
31. Disease Genes and Polymorphisms
- Conditional targeted mutation: Use of Cre recombinase to delete
genes in a temporal or spatial
- Cre recombinase is an enzyme that when in contact with LOXP
sites flanking a region of DNA, it can alter the DNA between
them.
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- - Knock-out/ Knock-in/Conditional
- Knock-out/ Inducible expression
32. Disease Genes and Polymorphisms
- Inducible Transgenic Mice
- The inserted gene is activated at a desired developmental
stage.
- An novel transcription factor designed to bind to the gene and
cause expression in the presence of an activator compound is
used.
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- - Knock-out/ Knock-in/Conditional
- Knock-out/ Inducible expression
33. Disease Genes and Polymorphisms
- Problems of conditional KOs and inducible transgenics
- Limited availability of tissue-specific mice to knock-out or
induce expression in a tissue specific fashion.
- Conditional knockout may not delete the gene in all of the
desired cells.
- Gene induction/repression may not be efficiently
regulated.
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- - Knock-out/ Knock-in/Conditional
- Knock-out/ Inducible expression
34. Disease Genes and Polymorphisms
- Metabolic Syndrome( Vartanian et al, 2006 )
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- -Knock-out/ Knock-in / Conditional
- Knock-out/ Inducible expression
- Dyslipidemia abnormal concentrations for lipoproteins in the
blood.
Deletion of the DNA glycosylase NEIL1 gene in mice results in
severe obesity, fatty liver, dyslipidemia, and insulin resistance;
symptoms that are consistent with metabolic syndrome in humans (a
collective title for obesity, insulin resistance, dyslipidemia, and
hypertension in humans) 35.
- Voluntary Ethanol Consumption inChrna6andChrnb3Knockout
Mice(Vidable,. L, et al. 2008; Ehringer Lab)
Human studies suggest that 6 & 3 nAch receptors are
associated with alcohol. Alcohol modulates binding. Types &
Forms of nAch Receptors Two Bottle Paradigm: Alcohol vs. Water
36.
- Voluntary Ethanol Consumption inChrna6andChrnb3Knockout Mice
(Ehringer Lab, Summer 08)
Chrna6 Knockouts Results Chrnb3 Knockouts Results Blue Male;
Green - Female 37. Disease Genes and Polymorphisms
- Examples of transgenic mice:
- Molecular defects of Alz.
- Targeted Transgenesis in Mice
- Targeted Mutagenesis in Mice
- - Types of mutagenic mice
- - Knock-out/ Knock-in/ Conditional
- Knock-out/ Inducible expression
38. Alternate Resources