| Collaborative studies with the Allergy
and Inflammation Research Group and the Gene Targeting Laboratory on the
role of IL-5 in allergic lung disease have continued. The present emphasis
is to define the roles played by IL-5 and eotaxin in regulating eosinophilic
inflammation and airways hyperresponsiveness using mice deficient in one
or both cytokines. The results indicate that eosinophils play a key role
in the orchestration of allergic inflammation and airways dysfunction.
These and other studies carried out by the Cytokines and Inflammation
Group, have formed the basis of our NH&MRC Program Grant on the Molecular
Mechanisms in the Regulation of Allergy and Inflammation which starts
in 2003.
Studies have also continued on the mechanisms regulating IL-5 gene
expression in T lymphocytes. This expression is both tissue-specific
and inducible and is very relevant to the eosinophil-mediated tissue
damage which occurs in asthma and allergy. Jun Wang established transient
expression systems in both mouse and human T cells and carried out comprehensive
transactivation studies to identify the transcription factors and the
binding sites involved in the regulation of the mouse and human IL-5
genes. The results suggest the formation of an enhanceosome type complex
in the proximal promoter region involving the transcription factors
GATA-3, AP-1 and Ets and possibly also HMGI(Y) which regulates inducible
and tissue-specific gene expression. The involvement of MAP kinase pathways
in IL-5 induction has also been studied.
The receptor system for IL-5 is shared with two related cytokines,
IL-3 and GM-CSF, which are also involved in the regulation of blood
cell formation and inflammation. One of the major problems in cell signalling
is to understand how these cytokines regulate blood cell growth and
function by binding and activating their cell-surface receptors. In
a collaborative project with Paul Carr and David Ollis (Research School
of Chemistry) using X-ray crystallography we have determined the structure
of the complete extracellular domain of the beta common receptor which
is the major signalling entity of the IL-5 receptor and is central to
the signalling of all three cytokines. The receptor was expressed in
insect cells and its crystallization and derivatization involved extensive
use of site-directed mutagenesis to improve crystal quality and to solve
the phase problem. The novel dimer configuration of the receptor gives
new insights into receptor activation. James Murphy is using site-directed
mutagenesis to define the residues of the beta common receptor which
are involved in forming the activated receptor complex. Peter Fineran,
Alice Church, Sally Ford, Janine Inggs and Jane Olsen have prepared
the activated complex of the closely related beta-IL-3 receptor. Further
structural studies should give a better understanding of the process
of receptor activation and provide opportunities to develop drugs capable
of controlling this important receptor system. Such drugs could be useful
in treating asthma, allergy or cancer.
The group has also participated in another project in the area of functional
genomics. with Hugh Campbell (Research School of Biological Sciences)
and the Gene Targeting Laboratory. In this work, the functions of the
mammalian homologues of two interesting genes, flightless and small
optic lobes, with functions in development and behaviour in the fruit
fly Drosophila are being investigated by gene targeting in mice. Investigations
to date have shown a function for mammalian flightless in early embryonic
development, remarkably analogous to its role in Drosophila. This cross-organism
approach uses information gained from studies with the simpler fruit
fly to better understand development and brain function in mammals.
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