Gag Sequence in HIV: Insights into Viral Architecture

HIV relies on a precise sequence of molecular events to assemble and mature into infectious particles. Central to this process is the Gag polyprotein, which orchestrates viral particle formation and structural integrity. Understanding its function provides crucial insights into how HIV replicates and spreads within host cells.

Research into the Gag sequence has revealed its role in viral architecture, including interactions with viral and host components. These findings have significant implications for antiviral strategies targeting HIV assembly and maturation.

Genome Location And Organization

The Gag sequence in HIV is encoded within the unspliced full-length RNA, serving as both the genomic template for new virions and the mRNA for Gag and Gag-Pol polyprotein synthesis. Positioned near the 5′ end of the HIV-1 genome, the gag gene spans approximately 1.5 kilobases and is one of the first regions transcribed following viral integration. This ensures early Gag expression, facilitating the formation of structural components necessary for virion assembly.

Transcription of the gag gene is driven by the HIV-1 long terminal repeat (LTR), a regulatory region containing promoter and enhancer elements responsive to host transcription factors. The LTR interacts with cellular proteins such as NF-κB and Sp1, which are upregulated in activated T cells, leading to increased Gag production in environments conducive to viral replication. Once transcribed, the gag mRNA is exported from the nucleus without splicing, a process mediated by the HIV-1 Rev protein, which binds to the Rev response element (RRE) within the viral RNA. This allows the full-length gag transcript to evade nuclear retention mechanisms that typically degrade unspliced mRNAs, ensuring efficient translation.

The gag gene encodes a single open reading frame translated into the Gag polyprotein, which is later cleaved into functional components by the viral protease. The polyprotein is synthesized on free ribosomes and trafficked to the inner leaflet of the plasma membrane, where it interacts with phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) to initiate virion assembly. This membrane association is mediated by the myristoylation of Gag’s N-terminal region, enhancing its affinity for lipid bilayers and promoting multimerization.

Key Proteins Encoded

The Gag polyprotein is processed into four major structural proteins—p17, p24, p7, and p6—each playing a distinct role in HIV particle formation and function.

p17

Also known as the matrix protein, p17 is derived from the N-terminal region of the Gag polyprotein and is essential for membrane targeting and virion stability. It contains a myristoylation signal that facilitates its association with the plasma membrane, anchoring the assembling virion.

Beyond membrane binding, p17 contributes to the structural integrity of the viral core by forming a shell beneath the lipid bilayer. It also contains a nuclear localization signal (NLS), assisting in the transport of the pre-integration complex into the nucleus of non-dividing cells. Mutations in p17 can disrupt virion assembly, leading to defective particles with reduced infectivity. Additionally, p17 interacts with host factors such as PI(4,5)P2, enhancing membrane binding and promoting Gag multimerization.

p24

The capsid protein p24 forms the conical core of the HIV virion, encapsulating the viral RNA and associated enzymes. It assembles into hexameric and pentameric units, creating a stable yet dynamic lattice that protects the viral genome while allowing controlled disassembly during infection.

p24 plays a key role in uncoating after viral entry. The timing of uncoating is crucial—premature disassembly can lead to genome degradation, while delayed uncoating can hinder reverse transcription. Host factors such as cyclophilin A (CypA) interact with p24 to modulate capsid stability. Small-molecule inhibitors like lenacapavir target p24 to interfere with capsid assembly and disassembly, highlighting its importance as a therapeutic target.

p7

The nucleocapsid protein p7 is responsible for packaging the viral RNA genome. It contains two highly conserved zinc finger motifs that bind to specific sequences within the HIV RNA, ensuring selective genome incorporation. These zinc fingers stabilize the RNA-protein complex, preventing degradation and facilitating proper genome positioning within the capsid.

Beyond RNA binding, p7 assists in reverse transcription by chaperoning the viral RNA during the early stages of infection. It helps anneal the tRNA primer to the viral genome, a necessary step for initiating DNA synthesis. Mutations in p7’s zinc finger domains impair RNA packaging and reduce viral infectivity. Additionally, p7 interacts with host proteins involved in RNA processing, further influencing genome encapsidation.

p6

The p6 protein, located at the C-terminal end of the Gag polyprotein, is involved in the final stages of virion release. It contains late (L) domains that recruit components of the host endosomal sorting complexes required for transport (ESCRT) machinery, facilitating budding and detachment of new virions. The primary L domains in p6 include the PTAP and YPXnL motifs, which interact with ESCRT-associated proteins such as Tsg101 and ALIX.

p6 also plays a role in incorporating the HIV-1 accessory protein Vpr into virions. Vpr is packaged through interactions with p6, influencing various aspects of the viral life cycle, including nuclear import of the pre-integration complex. Mutations in p6 can lead to defects in virion release, resulting in particles that remain tethered to the host cell membrane.

Role In Viral Assembly And Maturation

The Gag polyprotein orchestrates HIV assembly, driving the formation of new virions through coordinated molecular interactions. Once synthesized, Gag molecules traffic to the plasma membrane, where they accumulate at sites enriched in PI(4,5)P2. This lipid interaction is necessary for membrane binding and facilitates the conformational changes required for Gag multimerization. As Gag units associate, they form a lattice-like structure, shaping the emerging virion and ensuring proper spatial organization of viral components.

As assembly progresses, the recruitment of additional viral elements—including the genomic RNA and associated enzymes—ensures each particle possesses the necessary replication machinery. The nucleocapsid domain of Gag selectively packages the viral genome, binding specific RNA sequences to exclude non-viral nucleic acids. Proper genome incorporation is critical, as defective virions cannot initiate infection. Interactions between Gag and host factors such as Tsg101 and ALIX facilitate recruitment of ESCRT machinery, which mediates the final stages of budding. Without this, virions remain attached to the host cell membrane and are non-infectious.

Once budding is complete, the virus undergoes maturation, a process driven by Gag cleavage via the viral protease. This step converts the initially immature, spherical virion into its mature, infectious form. Proteolytic processing generates the distinct structural proteins—matrix, capsid, nucleocapsid, and p6—each adopting its final configuration. The capsid protein p24 undergoes a dramatic rearrangement, forming the characteristic conical core that houses the viral RNA. This transition is tightly regulated; premature or incomplete cleavage results in aberrant, non-infectious virions.

Importance Of Subtype Variations

HIV exhibits extensive genetic diversity, with multiple subtypes and circulating recombinant forms (CRFs) influencing viral replication and pathogenesis. Variations within the gag gene impact viral particle assembly, structural stability, and maturation dynamics. Subtype-specific differences in Gag processing efficiency have been observed, with some subtypes displaying altered cleavage kinetics due to variations in protease recognition sites. These differences affect infectivity, as improper processing can lead to defective virions with reduced replication capacity.

Structural adaptations in the Gag polyprotein among subtypes also influence viral fitness. Subtype C, the most prevalent globally, particularly in sub-Saharan Africa, has distinct Gag sequences that enhance replication in CD4+ T cells. Studies suggest subtype C Gag has a stronger association with host membrane lipids, potentially increasing transmission efficiency. In contrast, subtype B, predominant in North America and Europe, demonstrates different Gag-Pol cleavage patterns, potentially influencing maturation and drug susceptibility.