Differences in determinants required for complex formation and transactivation in related VP16 proteins

Area of Science:

• Virology
• Molecular Biology
• Gene Regulation

Background:

• Herpes simplex virus type 1 (HSV-1) protein VP16-H is crucial for viral gene expression, with distinct N-terminal and C-terminal domains.
• VP16-H recruits host proteins Oct-1 and HCF via its N-terminus and activates transcription via its C-terminus.
• Equine herpesvirus 1 (EHV-1) protein VP16-E shares N-terminal homology with VP16-H but lacks most of the acidic C-terminal domain.

Purpose of the Study:

• To investigate the mechanism of VP16-E's potent activation of immediate-early (IE) gene transcription in both EHV-1 and HSV-1.
• To determine if VP16-E follows the established two-domain model of VP16-H function.
• To identify key functional determinants within VP16-E responsible for its transactivation activity.

Main Methods:

• Comparative analysis of VP16-H and VP16-E protein structures and functions.
• Construction and testing of chimeric proteins involving domains of VP16-H and VP16-E.
• Assays to evaluate DNA binding complex formation with Oct-1 and HCF.
• Transactivation studies using GAL4 DNA-binding domain fusions.

Main Results:

• VP16-E is a potent activator of IE gene transcription, contradicting predictions based on its domain structure.
• The N-terminal domain of VP16-E does not sufficiently assemble into the Oct-1/HCF complex.
• A C-terminal region, dispensable in VP16-H, is essential for VP16-E activation.
• Chimeric proteins failed to fully recapitulate VP16-E's potency, suggesting complex interactions.

Conclusions:

• VP16-E does not adhere to the simple two-domain model of VP16-H.
• Activation by VP16-E likely involves intricate interactions between its N- and C-terminal regions.
• Efficient transactivation may depend on the specific mode of recruitment through complex formation with host factors.