Structural basis for PAS domain heterodimerization in the basic helix--loop--helix-PAS transcription factor hypoxia-inducible factor

Departments of Biochemistry and Pharmacology, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.67). 01/2004; 100(26):15504-9. DOI: 10.1073/pnas.2533374100
Source: PubMed


Biological responses to oxygen availability play important roles in development, physiological homeostasis, and many disease processes. In mammalian cells, this adaptation is mediated in part by a conserved pathway centered on the hypoxia-inducible factor (HIF). HIF is a heterodimeric protein complex composed of two members of the basic helix-loop-helix Per-ARNT-Sim (PAS) (ARNT, aryl hydrocarbon receptor nuclear translocator) domain family of transcriptional activators, HIFalpha and ARNT. Although this complex involves protein-protein interactions mediated by basic helix-loop-helix and PAS domains in both proteins, the role played by the PAS domains is poorly understood. To address this issue, we have studied the structure and interactions of the C-terminal PAS domain of human HIF-2alpha by NMR spectroscopy. We demonstrate that HIF-2alpha PAS-B binds the analogous ARNT domain in vitro, showing that residues involved in this interaction are located on the solvent-exposed side of the HIF-2alpha central beta-sheet. Mutating residues at this surface not only disrupts the interaction between isolated PAS domains in vitro but also interferes with the ability of full-length HIF to respond to hypoxia in living cells. Extending our findings to other PAS domains, we find that this beta-sheet interface is widely used for both intra- and intermolecular interactions, suggesting a basis of specificity and regulation of many types of PAS-containing signaling proteins.

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Available from: Ozgur Karakuzu, May 20, 2015
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    • "The NMR structures of hypoxia-inducible factor 2a (HIF-2a) and the aryl hydrocarbon receptor nuclear translocator (ARNT) PAS-B domains were chosen as the templates for homology modeling. Their coordinate files were obtained from the Protein Data Bank: entries 1P97 for HIF-2a (Erbel et al., 2003) and 1X0O for ARNT (Card et al., 2005). A three-dimensional model of the AaMet PAS-B domain was generated using MODELLER version 9v7 (Sali and Blundell, 1993). "
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    Molecular and Cellular Endocrinology 07/2014; 394(1-2). DOI:10.1016/j.mce.2014.06.021 · 4.41 Impact Factor
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    • "However, the availability of crystal and NMR structures of homologous proteins of the PAS superfamily allowed development of theoretical models for the LBD of the mouse AhR by applying homology modeling techniques (Pandini et al., 2007). The most recent homology model of the mouse AhR LBD (Fig. 3; Pandini et al., 2009) was built using the NMR-determined structures of the PAS B domains of HIF-2a (Erbel et al., 2003) and ARNT (Card et al., 2005) proteins as templates, given their higher degree of sequence identity and similarity with the AhR PAS B domain and their functional relationship to the AhR (Kewley et al., 2004). "
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    ABSTRACT: The Ah receptor (AhR) is a ligand-dependent transcription factor that mediates a wide range of biological and toxicological effects that result from exposure to a structurally diverse variety of synthetic and naturally occurring chemicals. Although the overall mechanism of action of the AhR has been extensively studied and involves a classical nuclear receptor mechanism of action (i.e., ligand-dependent nuclear localization, protein heterodimerization, binding of liganded receptor as a protein complex to its specific DNA recognition sequence and activation of gene expression), details of the exact molecular events that result in most AhR-dependent biochemical, physiological, and toxicological effects are generally lacking. Ongoing research efforts continue to describe an ever-expanding list of ligand-, species-, and tissue-specific spectrum of AhR-dependent biological and toxicological effects that seemingly add even more complexity to the mechanism. However, at the same time, these studies are also identifying and characterizing new pathways and molecular mechanisms by which the AhR exerts its actions and plays key modulatory roles in both endogenous developmental and physiological pathways and response to exogenous chemicals. Here we provide an overview of the classical and nonclassical mechanisms that can contribute to the differential sensitivity and diversity in responses observed in humans and other species following ligand-dependent activation of the AhR signal transduction pathway.
    Toxicological Sciences 09/2011; 124(1):1-22. DOI:10.1093/toxsci/kfr218 · 3.85 Impact Factor
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    • "When the modelled structures were superimposed on the crystal structure of the HIF-2α/Arnt PAS.B dimer, where interaction is through the β-sheet surfaces (49), it was clear that the most severe of our identified mutations in both Arnt and AhR cluster on faces away from the HIF-2α/Arnt PAS.B β-scaffold dimerization interface (Figure 6b). This suggests the PAS.A interactions use a face of the domain, encompassing α-C, β-C and β-D, that is different from the HIF-2α/Arnt PAS.B interface, as well as other examples of PAS domain interactions (9,17–21). Further, AhR and Arnt appear to interact through equivalent regions of PAS.A (Figure 6a). Partner specific Arnt D217 is centrally located in the putative dimerization interface in this model, while the S190P substitution on the opposite face may alter interaction with AhR through effects on packing. "
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