GENE REGULATORY MECHANISMS
Research Focus
The ability of mammalian cells to sense and respond to changes in oxygen levels,
particularly significant decreases (hypoxia), is essential for survival. As
well as playing a key role during development, this response is also important
for many physiological processes in adults, such as angiogenesis, and contributes
to the pathophysiology of major human diseases, including myocardial and cerebral
ischaemia, pulmonary hypertension and cancer. The Hypoxia Inducible transcription
Factors (HIFs) are central to the genomic response to hypoxia, regulating the
expression of a wide array of genes required for maintaining oxygen homeostasis
Known target genes include vascular endothelial growth factor (VEGF) and erythropoietin,
and the major role of HIFs in angiogenesis, particularly during cephalic and
cardiovascular development, has been demonstrated in both gene knockout and
transgenic mouse studies
FIH-1 is a ubiquitously expressed oxygen-dependent asparaginyl
hydroxylase that acts as a cellular oxygen sensor. In normoxia, FIH-1 hydroxylates
a single asparaginyl residue within the Hypoxic Inducible transcription Factors
(HIFs), causing transcriptional repression. In hypoxia the reduced O2-dependent
hydroxylation relieves repression. The HIFs are also regulated at the level
of protein stability by O2-dependent prolyl hydroxylases (PHDs). This mechanism
of oxygen sensing and modulation of protein activity by hydroxylation is both
novel and of particular importance for understanding how and why cells respond
to hypoxia.
Our laboratory has a specific interested in characterising these and other
components involved in the cellular response to hypoxia, specifically their
role during development as well as in adult organisms and human disease.
Novel targets of FIH-1.
As a ubiquitously expressed cellular oxygen sensor it is likely that FIH-1 will
have more substrates in addition to the HIFs. In support, it is well documented
that cellular responses to hypoxia also involve HIF-independent regulation of
multiple proteins. This project aims to identify novel targets of FIH-1, and
characterise the functional consequences of hydroxylation.
Characterisation of Notch1 asparaginyl
hydroxylation by FIH 1.
We have recently demonstrated that Notch1, which plays a crucial role in determining
cell fate, is specifically hydroxylated by FIH-1. The consequence of this hydroxylation,
at the molecular and functional level, as well as the physiological relevance,
are being investigated in this project.
Physiological role of FIH-1 during development.
The importance of the oxygen-sensing asparaginyl hydroxylase FIH-1 in the cellular
response to hypoxia, mediated in part by the regulation of HIF activity, has
been clearly demonstrated. However, the physiological importance of the ubiquitous
FIH-1 during development and in the whole organism has yet to be characterised
and is the focus of this project.
Differential regulation of HIF-1a and HIF-2a.
Although HIF-1a and HIF-2a are structurally very similar, and functionally can
both be regulated in a similar manner by oxygen-dependent prolyl and asparaginyl
hydroxylation, their physiological roles are quite distinct. In this project,
the reasons behind these different physiological roles are being explored, including
cell specific differences, specific interacting/regulating proteins and differential
target genes.
Recent publications
Collaborators & Linkages
- Dr M. Lardelli, School of Molecular
and Biomedical Science, University of Adelaide.
- Dr Grant Booker, School of Molecular and Biomedical Science, University
of Adelaide.Assoc.
- A/Prof. Murray Whitelaw, School of Molecular
and Biomedical Science, University of Adelaide.
- Dr J. Thompson, Department of Obstetrics and Gynaecology, University of
Adelaide.
- Dr Andrew Zannettino, Myeloma & Mesenchymal Research Laboratory, Hanson
Institute, IMVS, Adelaide.
- Prof. Jeff Gorman, Queensland Institute for Medical research, Brisbane,
QLDAssoc
- Prof Michael Raghunath, Departments of Bioengineering and Biochemistry,
National University of Singapore, Singapore.
- Prof. Urban Lendahl, Department of Cell and Molecular Biology, Karolinska
Institute, Stockholm, Sweden.
- Prof. Lorenz Poellinger, Department of Cell and Molecular Biology, Karolinska
Institute, Stockholm, Sweden.
- Dr Eric Metzen, Institute of Physiology, University of Luebeck, Luebeck,
Germany.
Recent Publications
Hampton-Smith, R.J., Peet, D.J. (in press) McGraw-Hill Yearbook of Science
and Technology 2007, The McGraw-Hill Editorial Staff, ed. ( McGraw-Hill, USA)
Bilton, R., Trottier, E., Pouyssegur, J., Brahimi-Horn, M.C. (2006) ARDent about
acetylation and deacetylation in hypoxia signalling. Trends in Cell Biology
16: 616-21
Bracken, C.P., Fedele, A.O., Linke, S., Balrak, W., Lisy, K., Whitelaw, M.L.,
Peet, D.J. (2006) Cell-specific regulation of hypoxia-inducible factor (HIF)-1?
and HIF-2? stabilization and transactivation in a graded oxygen environment.
The Journal of Biological Chemistry 281: 22575-85
Peet, D.J. and Linke, S., Regulation of HIF:asparaginyl hydroxylation. (2006)
In Signalling Pathways in Acute Oxygen Sensing, Novartis Foundation Symposium
No. 272, D. J. Chadwick and J.Goode, ed. (Weinheim, Germany: Wiley-VCH Verlag
GmbH)
Bracken, C.P., Whitelaw, M.L., Peet, D.J. (2005) Activity of hypoxia-inducible
factor 2a is regulated by association with the NF- K B essential modulator.
The Journal of Biological Chemistry 280: 14240-51
Linke, S., Stojkoski, c., Kewley, R.J., Booker, G.W., Whitelaw, M.L., Peet,
D.J. (2004) Substrate requirements of the oxygen-sensing asparaginyl hydroxylase
factor-inhibiting hypoxia-inducible factor. The Journal of Biological Chemistry
279: 14391-7
Peet, D.J., Lando, D., Whelan, D.A., Whitelaw, M.L., Gorman, J.J. (2004) Oxygen-dependent
asparagine hydroxylation. Methods in Enzymology 381: 467-87
Bracken, C.P, Whitelaw, M.L., Peet, D.J. (2003) The hypoxia-inducible factors:
key transcriptional regulators of hypoxic responses. Cellular and Molecular
Life Sciences 60: 1376-93
Lando, D., Gorman, J.J., Whitelaw, M.L., Peet, D.J. (2003) Oxygen-dependent
regulation of hypoxia-inducible factors by prolyl and asparaginyl hydroxylation.
European Journal of Biochemistry 270: 781-90
Lees, M.J., Peet, D.J., Whitelaw M.L. (2003) Defining the role for XAP2 in stabilization
of the dioxin receptor. The Journal of Biological Chemistry 278: 35878-88
Fedele, A.O., Whitelaw, M.L., Peet, D.P. Regulation of gene expression by the
hypoxia inducible factors. Molecular Interventions 2:230-43
Lando, D., Peet, D. J., Gorman, J. J., Whelan, D. A., Whitelaw, M. L*., Bruick,
R. K*. (2002) FIH is an asparaginyl hydroxylase enzyme that regulates the transcriptional
activity of Hypoxia Inducible Factor. Genes and Development, 16, 1466-71. (*joint
senior author)
Lando, D., Peet, D.J., Pongratz, I., Whitelaw, M.L. (2002) Mammalian Two-Hybrid
Assay Showing Redox Control of HIF-Like Factor. Methods in Enzymology , Vol.
353
Lando, D., Peet, D.J., Whelan, D.A., Gorman, J.J., Whitelaw, M.L. (2002) Asparagine
Hydroxylation of the HIF Transactivation Domain: A hypoxic Switch. Science 295:858-61
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