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The targeting of US28 is dependent on GASP-1

Background

The human cytomegalovirus (HCMV) encodes the seven transmembrane (7 TM)/G protein-coupled receptor (GPCR) US28. US28 is able to endocytose and signal in a constitutive - i.e. ligand-independent - manner and is located predominantly in the membranes of intracellular organelles, especially late endosomes/lysosomes and multivesicular bodies (MVBs). It was suggested that the virions of HCMV may be budding into the membranes of these MVBs, where the viral receptors are then incorporated into the viral membranes during the final stages of virus assembly.

Methods

Protein-protein interaction between US28 and GASP-1 (G protein-coupled receptor-associated sorting protein-1) was investigated by GST-fusion protein-binding assay, MBP protein competition binding assay and co-immunoprecipitation. In HEK293 cells endogenously expressing GASP-1 the interaction between US28 and GASP-1 was disrupted by i) overexpression of dominant negative cGASP-1 or by ii) shRNA knock-down of endogenous GASP-1. The role of GASP-1 in the post-endocytic trafficking of US28 was analyzed by means of immunocytochemistry and biotin protection degradation assay.

Results

Here we show that GASP-1, which sorts many GPCRs to the lysosomes, also plays an important role in the post-endocytic targeting of US28. We find that disruption of the GASP-1/US28 interaction by either i) overexpression of dominant negative cGASP-1 or by ii) shRNA knock-down of endogenous GASP-1 is sufficient to alter the lysosomal targeting of US28 and also its post-endocytic degradation.

Conclusion

Our data suggest that the interaction of US28 and GASP-1 has important implications for the post-endocytic fate of US28.

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Correspondence to Maria Waldhoer.

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Open Access This article is published under license to BioMed Central Ltd. This is an Open Access article is distributed under the terms of the Creative Commons Attribution License ( https://creativecommons.org/licenses/by/2.0 ), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Moser, E., Tschische, P., Platzer, W. et al. The targeting of US28 is dependent on GASP-1. BMC Pharmacol 9, A1 (2009). https://doi.org/10.1186/1471-2210-9-S2-A1

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Keywords

  • HEK293 Cell
  • Human Cytomegalovirus
  • Virus Assembly
  • Multivesicular Body
  • Intracellular Organelle