Single-Cell and Single-Cycle Analysis of HIV-1 Replication

Area of Science:

• Virology
• Cell Biology
• Molecular Biology

Background:

• HIV-1 replication dynamics are poorly understood, especially in individual cells.
• Asynchrony in early replication steps complicates measuring later stages and individual cell variations.

Purpose of the Study:

• To establish microscopy-based methods for dynamic measurement of HIV-1 gene expression in individual cells.
• To quantify replication delays caused by HIV-1 gene expression and Gag matrix protein.
• To correlate restriction factor APOBEC3G dynamics with HIV-1 replication.

Main Methods:

• Microscopy-based dynamic measurement of HIV-1 reporter and antiviral gene expression.
• Analysis of HIV-1 replication cycle delays.
• Quantification of APOBEC3G removal dynamics.

Main Results:

• The interval between early and late HIV-1 gene expression is approximately 3 hours.
• The Gag matrix protein introduces a 6-12 hour delay in extracellular virion production.
• Matrix delays particle assembly until APOBEC3G is largely removed, potentially optimizing infectious virus production.

Conclusions:

• HIV-1 replication exhibits significant heterogeneity in individual cells.
• Programmed delays in virion genesis may be crucial for efficient infectious HIV-1 production.
• The late stages of HIV-1 replication, including virion assembly, are tightly regulated and occur over a short period within the cell's lifespan.