Subcellular compartmentalization of adeno-associated virus type 2 assembly

Area of Science:

• Molecular Biology
• Virology
• Cell Biology

Background:

• Adeno-associated virus type 2 (AAV-2) is a small DNA virus that requires a helper adenovirus for efficient replication.
• Understanding the intracellular trafficking and assembly of viral components is crucial for deciphering viral life cycles and developing antiviral strategies.

Purpose of the Study:

• To investigate the intracellular localization and dynamics of AAV-2 Rep proteins, capsid (VP) proteins, and viral DNA during adenovirus coinfection.
• To elucidate the role of Rep proteins in the nuclear redistribution of AAV capsids and capsid assembly processes.

Main Methods:

• Immunofluorescence microscopy to visualize protein localization.
• In situ hybridization to detect viral DNA.
• Analysis of viral component distribution during different stages of AAV-2/adenovirus coinfection.

Main Results:

• Rep proteins initially form punctate nuclear foci, then distribute homogeneously in the nucleoplasm, and finally cluster at the nuclear periphery.
• AAV-2 DNA colocalizes with Rep proteins throughout infection.
• Capsid proteins (VP1, VP2, VP3) transiently accumulate in the nucleolus, suggesting its role in initial capsid assembly.
• Rep proteins mediate the nuclear redistribution of capsids from the nucleolus to the nucleoplasm.
• Capsid formation is dependent on capsid protein concentration.

Conclusions:

• Nucleolar components are involved in the initiation of AAV-2 capsid assembly.
• DNA packaging occurs in the nucleoplasm.
• AAV-2 Rep proteins play a critical role in the nuclear trafficking and redistribution of viral capsids during coinfection.