SFB796 - Sub project A6

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Functional selectivity as a result of viral chemokine receptor heteromerization

 

Project summary

The hijacking of a host signalling machinery is a fundamental part of viral host-evasion strategy. Certain viruses like human cytomegalovirus (HCMV), Epstein-Barr virus (EBV) and Kaposi's sarcoma herpes virus (KSHV) encode viral G-protein coupled receptors (vGPCRs). These vGPCRs couple very efficiently to signalling networks of a host. The expression of vGPCRs augments virus survival, host invasion and, in some cases, oncogenesis or cardiovascular disease by exploiting favoured signalling cascades. Numerous mammalian GPCRs function as homo- or heteromers. Reciprocal allosteric modulation of different protomers often results in a distorted signalling outcome compared to the signalling of homodimers. This phenomenon is described as a form of functional selectivity. Heterodimers composed of different HCMV vGPCRs have been described, but the molecular mechanisms governing heteromerization process and its functional consequences remain scarcely characterized. Also, heteromerization of UL33 and UL78 vGPCRs with the host's chemokine receptors CCR5 and CXCR4 has been reported to have a profound effect on the host's receptor signalling and thus consequently on the re-programming of central cellular processes. The concept of the physical association and cross-receptor interactions thus represents an unexpected mechanism for the GPCR regulation and function, and opens great perspectives for the development of pharmaceuticals that exclusively modulate heteromers.

The main goals of this project are the identification of molecular mechanisms governing heteromerization processes and the characterization of changes in the functional response as a consequence of receptor heteromerization. To identify molecular mechanisms involved in a heteromerization event, the combination of co-immunoprecipitation, pull-down techniques and mass spectrometry will be applied. This approach will allow us to determine heteromerization partners of US28 (vGPCR of HCMV) and the identification of epitopes (e.g., intracellular loops, C-terminus) governing receptor heteromerization. The characterization of changes in the functional response of G proteins upon heteromerization will be undertaken in complemented donor-acceptor resonance energy transfer assays, which will allow us to achieve the control over the identity of the components comprising the signalling unit. This technique combines the features of protein complementation and resonance energy transfer, which enables the time-resolved study of defined receptor heteromers in living cells. Our recently described small molecules that act as inverse agonists of the US28 receptor, and endogenous ligands will be used in these assays. By understanding the precise mechanisms of signal transduction and negative regulation of vGPCRs, it should be possible to devise strategies to block the actions of vGPCRs (e.g. by small molecule inverse agonists) and to develop practical therapies for diseases.

 

Project relevant publications

Bernat, V., Heinrich, M., Baumeister, P., Buschauer, A., Tschammer, N.   (2012).   Synthesis and application of the first radioligand targeting the allosteric binding pocket of chemokine receptor CXCR3.   ChemMedChem.   accepted , doi:cmdc.201200184 .

Kralj A, Wetzel A, Mahmoudian S, Stamminger T, Tschammer N*, Heinrich M*.   (2011).   Identification of novel allosteric modulators for the G-protein coupled US28 receptor of human cytomegalovirus.   Bioorg. Med. Chem. Lett. 21(18): 5446-5450   * - corresponding author.

Koschatzky S, Tschammer N, Gmeiner P.   (2011).   Cross-receptor interactions between dopamine D2L and neurotensin NTS1 receptors modulate binding affinities of dopaminergics.   ACS Chem. Neurosci. 2(6): 308-316.

Tschammer N, Bollinger S, Kenakin T, Gmeiner P.   (2011).   Histidine 6.55 is a major determinant of ligand biased signaling in dopamine D2L receptor.   Mol. Pharmacol. 79(3): 575-585.

Tschammer N, Elsner J, Götz A, Ehrlich K, Schuster S, Ruberg M, Kühhorn J, Thompson D, Whistler J, Hübner H, Gmeiner P.   (2011).   Highly potent 5-aminotetrahydropyrazolopyridines: Enantioselective dopamine D3 receptor binding, functional selectivity, and analysis of receptor-ligand interactions.   J. Med. Chem. 54(7): 2477-2491.