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Structural and functional analysis of the membrane-spanning domain of the human immunodeficiency virus type 1 Vpu
E Tiganos1, J Friborg, B Allain
1Département de Microbiologie et Immunologie, Faculté de Médecine, Université de Montréal, CP 6128, Succursale Centre-ville, Montréal, Québec, H3C 3J7, Canada.
Insights
The HIV-1 Vpu protein
Area of Science:
- Virology
- Molecular Biology
- Cell Biology
Background:
- Human immunodeficiency virus type 1 (HIV-1) Vpu is an integral membrane phosphoprotein.
- Vpu has two main functions: degrading CD4 and enhancing viral release.
- These functions are linked to distinct Vpu domains: transmembrane (TM) and cytoplasmic.
Purpose of the Study:
- To investigate the role of the Vpu transmembrane (TM) domain in viral release and CD4 degradation.
- To identify specific amino acids within the TM domain crucial for Vpu's biological activities.
Main Methods:
- Mutational analysis of the Vpu TM domain.
- Integration and localization studies in canine pancreatic microsomal membranes.
- Assessment of viral particle release and CD4 binding/degradation.
Main Results:
- Mutations in the Vpu TM domain impaired viral particle release.
- The C-terminal region of the TM domain is critical for both viral release and CD4 degradation.
- Specific TM domain alterations affected CD4 binding and subsequent degradation.
Conclusions:
- The Vpu TM domain contains key determinants for enhancing viral particle release.
- Both the TM and cytoplasmic domains of Vpu are essential for its biological functions.
- The C-terminal TM region plays a dual role in Vpu activity.
Abstract:
The human immunodeficiency virus type 1 (HIV-1) vpu gene product is a class I integral membrane phosphoprotein that is capable of oligomerization. Two distinct biological activities have been attributed to Vpu: induction of CD4 degradation in the endoplasmic reticulum and enhancement of viral particle release from the plasma membrane of infected cells. These two biological activities were shown to involve two separable structural domains: the N-terminal transmembrane (TM) domain and the C-terminal cytoplasmic domain. The TM domain mediates enhancement of viral particle release, whereas phosphorylation of the cytoplasmic domain is essential for Vpu-induced CD4 degradation. In this study, we performed a mutational analysis of the TM domain of Vpu to delineate amino acids that are important in the process of viral particle release or in Vpu-induced CD4 degradation. Substitution of conserved amino acids from the N-terminal, middle, or C-terminal parts of the native VpuTM domain generated proteins that integrated normally into canine pancreatic microsomal membranes, exhibited subcellular localization similar to those of wild-type Vpu, but partially lost their ability to enhance viral particle release, strongly suggesting that the VpuTM domain contains determinants responsible for Vpu-mediated enhancement of viral particle release. Interestingly, the C-terminal TM mutant VpuIVW, in contrast to the other mutants, also lost its ability to bind and consequently degrade the CD4 molecule, indicating that the alteration of the C-terminal part of the TM did interfere with this function of Vpu. Taken together, our study supports the notion that both structural elements of Vpu (TM and cytoplasmic) contribute to the biological activities of Vpu.