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Computational Biology 15
THREE EXAMPLES: INTEGRATIVE APPROACH
TO COMPLEX TRAIT ANALYSIS IN MICE
Commonly used genetic mapping tools identify chromosomal regions
affecting complex traits in rodent models of human disease-related
raits. However, identification of the causative genetic factor within a
linked chromosomal region is essential for obtaining new information
about a disease or biological process. The process of identifying genetic
loci within linked chromosomal regions is difficult and often
unproductive, which has been a source of frustration for many (50).
However, the following three examples demonstrate how the combined
use of whole genome gene expression profiling and QTL analysis of
mouse genetic models enables the efficient identification of causative
genetic variants within linked regions. 3.1. Murine Experimental Model
of Asthma An integrative approach was utilized to analyze a wellcharacterized murine genetic model that mimics the pathophysiology of
human allergic asthma (Fig.2). In this model, allergen exposure results
in airway hyperresponsiveness,Fig. 2. Diagrammatic representation of
an integrated genetic and genomic approach for analyzing a murine
experimental genetic model of allergic asthma. increased airway
epithelial mucus content, antigen-specific IgE in serum, and pulmonary
eosinophilia (51,52). Inbred mouse strains vary markedly in their
susceptibility to disease induction in this model. Two strains with
markedly different susceptibilities to experimental allergen-induced
asthma were used:
theA/Jstrainishighlysusceptibletoallergeninducedairwayhyperresponsiv
eness and the C3H/HeJ strain is highly resistant. Analysis of the
inheritance pattern of the asthmatic response in intercross progeny led
to the identification of regions on chromosomes 2 and 7 that regulated
asthma susceptibility in this experimental model. To identify gene
candidates, pulmonary gene expression wasprofiledusingoligonucleotide
microarrays. After phenotypical assessment, lungs were harvested from
parental (A/J, C3H/HeJ) and F1 mice and from eight first-generation
backcross progeny (BC1) that exhibited phenotypically extreme
allergen-induced airway responsiveness. As indicated in Fig. 3, 2718 of
the 7350 genes on the microarray were expressed in the lungs of the
parental strains. Atotal of 739 genes were differentially expressed in the
lungs16 Peltz
Fig. 3. Identification of a gene regulating susceptibility in an
experimental murine genetic model of allergic asthma. The number of
differentially expressed genes after comparison of gene expression
profiles was determined after examining the number of genes: on the
oligonucleotide array (all); expressed in the lungs of the two parental
mouse strains (present); differentially expressed between the two
strains according to the criteria provided by the manufacturer ();
computationally determined to be over threefold different (3) between
the two strains; or differentially expressed among the eight BC1
intercross progeny examined. The gene expression profile of five pairs
of phenotypically extreme BC1 progeny was compared as described in
the text. The number of differentially expressed genes when three (3)
or four (4) of the five comparisons indicated that the gene was
differentially expressed is shown. allergen-induced airway
hyperresponsiveness. C5 transcript levels in whole lungs of ovalbuminsensitized A/J, C3H, and F1 (A/J C3H) mice are indicated; six
highresponder BC1 and six low-responder BC1 mice are shown in
comparison to their dynamic airway hyperresponsiveness. BC1 mice
with C5 deficiency are represented by diamonds and BC1 mice that are
C5 heterozygous are shown as circles. (Reproduced with permission
from ref. 30.) C5 mRNA expression revealed that it was homozygous for
the A/J allele at the C5 locus. All of the other intercross progeny that
were resistant to experimental asthma induction had a C3H-derived C5
allele. Subsequent analysis revealed that the presence of a deletion in
the coding sequence of C5 in susceptible mice leads to the absence of C5
protein and susceptibility to the asthmatic trait (30). The mechanism by
which a genetic deficiency in C5 could lead to susceptibility to an
asthmatic trait could then be characterized. Subsequent experimental
analysis revealed that C5 deficiency effected the production of cytokines
regulating the asthmatic response. IL-12 is a cytokine with potent
effects on T-cell differentiation. When produced in the airways, it can
prevent or reverse allergic asthma. Inhibition of C5-mediated signaling
by blockade of the C5a receptor rendered human monocytes unable to
produce IL-12 in vitro. This provided a plausible mechanism for the
regulation of susceptibility to asthma by alleles of C5 in mice.