Importance of basic residues in the nucleocapsid sequence for retrovirus Gag assembly and complementation rescue

Area of Science:

• Virology
• Molecular Biology
• Protein Structure

Background:

• Gag proteins of Rous sarcoma virus (RSV) and human immunodeficiency virus (HIV) contain interaction (I) domains within nucleocapsid (NC) sequences, overlapping zinc finger motifs.
• These I domains are crucial for viral particle density and proper viral assembly.
• The number and properties of I domains vary across different retroviruses, influencing viral particle formation.

Purpose of the Study:

• To characterize sequence features and properties of I domains in viral Gag proteins.
• To investigate the role of zinc finger motifs versus basic residue clusters in particle density.
• To determine how I domain number affects Gag protein rescue into viral particles.

Main Methods:

• Analysis of Gag proteins with varying numbers of zinc finger motifs using chimeric constructs (RSV, MLV, HFV).
• Mutational analysis of MLV NC sequence and inspection of HFV Gag sequences.
• Complementation rescue experiments with RSV-MLV chimeras to assess Gag protein rescue efficiency.

Main Results:

• Chimeric proteins with MLV (one zinc finger) had one I domain, while those with HFV (no zinc fingers) had at least two.
• Clusters of basic residues, not zinc finger motifs, are essential for proper particle density.
• A string of basic residues can substitute for RSV I domains.
• The NC sequence of MLV, not I domain number, dictates differences in Gag protein rescue efficiency.
• RSV-MLV chimeras could be rescued when budding blocks occurred post-membrane transport.

Conclusions:

• Basic residue clusters in NC sequences are critical for viral particle density, independent of zinc finger motifs.
• MLV Gag proteins initiate interactions later in synthesis compared to RSV and HIV, affecting rescue efficiency.