Identification of a highly conserved valine-glycine-phenylalanine amino acid triplet required for HIV-1 Nef function

Retrovirology

Bruno Verhasselt³  Oliver T Fackler⁶  Kevin K Arien¹  Guido Vanham⁴  Matthias Geyer²  Kalle Saksela⁵  Evelien Naessens³  Jolien Vermeire³  Veronica Iannucci³  Bettina Stolp⁶  Pieter J Meuwissen³ 

[1] Present Address: Department of Biomedical Sciences , Virology Unit, Institute of Tropical Medicine, Antwerp, Belgium;Max Planck Institute for Molecular Physiology, Dortmund, (D-44227), Germany;Department of Clinical Chemistry, Microbiology, and Immunology, Ghent University, Ghent, (B-9000), Belgium;Department of Biomedical Sciences, Virology Unit, Institute of Tropical Medicine, Antwerp, (B-2000), Belgium;Department of Virology, Haartman Institute, University of Helsinki and Helsinki University Central Hospital, Helsinki, (FIN-00014), Finland;Department of Infectious Diseases, Virology, University Hospital Heidelberg, INF 324, Heidelberg, (D-69120), Germany

关键词: Replication;    Infectivity;    Receptor downregulation;    Lck;    Cytoskeleton;    SH3 domain binding;    Sequence motifs;    Nef;    HIV;   

Others  :  1209311
DOI  :  10.1186/1742-4690-9-34

 received in 2011-12-16, accepted in 2012-04-27,  发布年份 2012

【 摘 要 】

Background

The Nef protein of HIV facilitates virus replication and disease progression in infected patients. This role as pathogenesis factor depends on several genetically separable Nef functions that are mediated by interactions of highly conserved protein-protein interaction motifs with different host cell proteins. By studying the functionality of a series of nef alleles from clinical isolates, we identified a dysfunctional HIV group O Nef in which a highly conserved valine-glycine-phenylalanine (VGF) region, which links a preceding acidic cluster with the following proline-rich motif into an amphipathic surface was deleted. In this study, we aimed to study the functional importance of this VGF region.

Results

The dysfunctional HIV group O8 nef allele was restored to the consensus sequence, and mutants of canonical (NL4.3, NA-7, SF2) and non-canonical (B2 and C1422) HIV-1 group M nef alleles were generated in which the amino acids of the VGF region were changed into alanines (VGF→AAA) and tested for their capacity to interfere with surface receptor trafficking, signal transduction and enhancement of viral replication and infectivity. We found the VGF motif, and each individual amino acid of this motif, to be critical for downregulation of MHC-I and CXCR4. Moreover, Nef’s association with the cellular p21-activated kinase 2 (PAK2), the resulting deregulation of cofilin and inhibition of host cell actin remodeling, and targeting of Lck kinase to the trans-golgi-network (TGN) were affected as well. Of particular interest, VGF integrity was essential for Nef-mediated enhancement of HIV virion infectivity and HIV replication in peripheral blood lymphocytes. For targeting of Lck kinase to the TGN and viral infectivity, especially the phenylalanine of the triplet was essential. At the molecular level, the VGF motif was required for the physical interaction of the adjacent proline-rich motif with Hck.

Conclusion

Based on these findings, we propose that this highly conserved three amino acid VGF motif together with the acidic cluster and the proline-rich motif form a previously unrecognized amphipathic surface on Nef. This surface appears to be essential for the majority of Nef functions and thus represents a prime target for the pharmacological inhibition of Nef.

【 授权许可】

   
2012 Meuwissen et al.; licensee BioMed Central Ltd.

【 图 表 】

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