Introduction: HLA-C is selectively incorporated into HIV-1 envelope. We reported that viruses produced from HLA-C silenced cells are less infectious and demonstrated a specific association between HLA-C and gp120. Our purpose is to unravel the HLA-C/gp120 association to reveal new targets for the development of neutralizing antibodies and new anti-HIV compounds. Methods: Fluorescent-tagged HLA-C and gp120 were prepared and used for bimolecular fluorescence complementation (BiFC) analysis to study their association into cellular compartments. To identify gp120 domains involved in the association with HLA-C, different gp120 deletion mutants are being constructed and tested. Results: A BiFC complementation signal between HLA-C and gp120 is detectable in the endoplasmic reticulum (ER), Golgi apparatus, late endosomes and on the cell membrane. Confocal microscopy does not show a co-localization signal between gp120 and β2-microglobulin (β2m). Discussion: The co-localization analysis shows that HLA-C, most likely as an open conformer, interacts with gp120 early in the ER and that they remain associated up to the cell membrane, travelling together trough the Golgi apparatus and late endosomes. No gp120 and β2m co-localization is evident, suggesting a competition between them for HLA-C binding.

Association analysis between HIV-1 Env and HLAC using bimolecular fluorescence complementation

ZIGLIO, Serena;ASTONE, Dalila;RACCHIOLLI, Pierpaolo;ZIPETO, Donato
2011

Abstract

Introduction: HLA-C is selectively incorporated into HIV-1 envelope. We reported that viruses produced from HLA-C silenced cells are less infectious and demonstrated a specific association between HLA-C and gp120. Our purpose is to unravel the HLA-C/gp120 association to reveal new targets for the development of neutralizing antibodies and new anti-HIV compounds. Methods: Fluorescent-tagged HLA-C and gp120 were prepared and used for bimolecular fluorescence complementation (BiFC) analysis to study their association into cellular compartments. To identify gp120 domains involved in the association with HLA-C, different gp120 deletion mutants are being constructed and tested. Results: A BiFC complementation signal between HLA-C and gp120 is detectable in the endoplasmic reticulum (ER), Golgi apparatus, late endosomes and on the cell membrane. Confocal microscopy does not show a co-localization signal between gp120 and β2-microglobulin (β2m). Discussion: The co-localization analysis shows that HLA-C, most likely as an open conformer, interacts with gp120 early in the ER and that they remain associated up to the cell membrane, travelling together trough the Golgi apparatus and late endosomes. No gp120 and β2m co-localization is evident, suggesting a competition between them for HLA-C binding.
HIV-1 HLA-C BiFC
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11562/389072
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