~REGULATION & CONTROL~
TABLE OF CONTENTS:
II REGULATION
III CONCLUSION
Hypoxia-Inducible Factor 1 Complex
Hypoxia Inducible Factors are proteins that serve to increase the transcription of specific proteins that (1), increase the flow of oxygen to the tissue area and (2), to stabilize the tissue to hypoxic conditions. These proteins are binary DNA-binding complexes that control the hypoxia responses of several genes, and regulate the adaptive responses to the lack of oxygen (Srinivas). These protein structures follow a basic-helix-loop-helix-PAS motif. This complex is formed from two protein subuints - an alpha (HIF-alpha) and a beta (HIF-beta) subunit. While both the HIF-alpha and HIF-beta proteins are continuously synthesized, the HIF-alpha is continuously degraded under normal, normoxic conditions. It is under hypoxic conditions that the HIF-alpha subunit is stabilized, and the dimer complex forms (though this stabilization may also be induced with iron chelators, transition metals, and antioxidants). The sequences and mechanisms involved in this stabilization are largely unknown. In this paper, HIF-1 will be the basic model discussed. HIF-2 and HIF-3 have also been identified, however, they differ only in the HIF-alpha subunit, share an approximate 55-75% homology with HIF-1alpha, and the basic stabilization mechanisms appear to be identical (Srinivas).
REGULATION
The
overall method of general regulation is somewhat basic -- both subunits
are continuously transcribed in the cells, and only the alpha is rapidly
degraded under normoxic conditions. Several cellular processes have
been linked to the degredation of HIF-alpha, all of which involve an enzymatic
cleavage using oxygen. As oxygen is depleated, the pH of the cellular
environment drops, stabilizing the alpha subunit, and triggering the formation
of the alpha+beta complex. This complex acts as a transcription promoter,
known to bind with at least 14 different sequences, all of which are G/C
rich, and correspond with hypoxia-related proteins, and the alpha subunits.
Several species
of mammal have been studied so far, including humans, ferrets, mice, and
several primate species, all of which exhibit HIF-1, 2 and 3. Each
of these alpha units display what appear to be the same basic mechanisms
of both stabilization and degradation. ìAll three alpha members appear
to be regulated in a similar way, predominantly by post translational mechanismsî
(Srinivas). Under normoxic conditions, the alpha subunit is continually
synthesized, but quickly is degraded by the ubiquitin- proteasome cycle.
Hypoxia, transition metals, iron chelators, and several antioxidants decrease
the degradation of the HIF-alpha sub-units, allowing dimerization with
HIF-1alpha and formation of the HIF-DNA binding complex. The signals, mechanisms,
and sequences of events involved in the hypoxia-regulated degradation of
the HIF-alpha proteins are not fully understood, and not extensively studied.
To confuse matters further, several different research groups have reported
conflicting results.
Stabilization appears to be largely dependent on a carefully maintained ratio of reduced glutathione (GSH) to oxidized glutathione (GSSH). Several studies link the hypoxic induction of HIF-alpha with this GSH/GSSH equilibrium (Haddad). There is currently no published evidence that the glutathione redox equilibrium, or glutathione itself is in any way directly responsable for the actual degrataion of the HIF-alpha subunit or the alpha+beta complex.
Phosphorylation
does not appear to be a deactivating mechanism of HIF complexes, as no
serines or threonines exist within the universal regions of HIF-alpha,
and mutations in all possible phosphorylation sites on HIF-1-alpha have
not affected the functionality of the protein (Pugh).
This
protein complex has only fairly recently been discovered, and deemed important
enough to gtudy in great detail. In late 1995, the protein was first
discovered, and first published in '96. Since then, hundreds of papers
have been released, growing exponentally every year. Little is known
mechanically about this complex, and especially it's degredation pattern.
Possible degredation mechanisms all appear to be dependent on pH, oxygen-scavenging
enzymes and NADPH availability, but actual pathways and mechanisms are
far from understood. Experiments have defined several domains in
HIF-alphas which can independently grant the hypoxia-inducible properties,
and may suggest mechanisms of activation dependent on the concentration
of HIF-alpha subunits, but these are currently hypothetical explanations
that are being investigated.
REFERENCES:
1. "Characterization of an Oxygen/Redox-Dependent Degradation Domain of Hypoxia-Inducible Factor [Alpha] (HIF-[alpha]) Proteins" Vickram Srinivas, et al. Cerdeza Foundation for Hematologic Research, Department of Medicine, Thomas Jefferson University. 17 May 1999. Biochemical and Biophysical Research Communications, 260:557-561.
2. "ATP=stimulated c-fos and zif268 mRNA expression is inhibited by chemical hypoxia in a rat brain-derived type 2 astrocyte cell lune, RBA-2." A.C. Hung, H.M. Huang, et al. Journal of Cell Biochemistry, March 2000. 77(2):323-32.
3. "Regulation of mammalian O2 homeostasis by hypoxia-incucible factor 1." Gregg L. Semenza. Annual Review of Cellular Developmental Biology, 1999. 15:551-78.
4. "Oxygen sensing, hypoxia-inducible factor-1 and the regulation of mammalian gene expression." Peter J. Ratcliffe, John F. O'Rourke, et al. Journal of Experimental Biology, August 1998. 201(8):1153-62.
5. "Hypoxia Response Element of the Human Vascular Endothelial Growth Factor Gene Mediates Transcriptional Regulation by Nitric Oxide: Control of Hypoxia-Inducible Factor-1 Activity by the Nitric Oxide." Hideo Kimure, Alessandro Weisz, et al. Blood, January 2000. 95(1):189-97.
6. "Reciprocal Positive Regulation of Hypoxia-Inducible Factor-1[alpha] and Insulin-Like Growth Factor 2." David Feldser, Fayton Agani, et al. Cancer Research, 1999. 59(16):3915-18.
7. "Interleukin-1[beta]
and tumor Necrosis Factor-[alpha] Stimulate DNA Binding of Hypoxia-Inducible
Factor-1." Thomas Hellwig-Burgel, Karen Rutkowski, et al. Blood,
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