Sindbis Macrodomain Poly-ADP-Ribose Hydrolase Activity Is Important for Viral RNA Synthesis

Area of Science:

• Virology
• Molecular Biology
• Biochemistry

Background:

• ADP-ribosylation is a key posttranslational modification involved in cellular stress and viral infections.
• Alphaviruses possess macrodomains with ADP-ribosylhydrolase (ARH) activity, suggesting a role in modifying the host or viral ADP-ribosylome.
• The specific contribution of mono- versus poly-ADP-ribose removal by viral macrodomains to replication remained unclear.

Purpose of the Study:

• To differentiate the roles of mono- and poly-ARH activities of alphavirus macrodomains.
• To investigate the necessity of viral poly-ARH activity for efficient alphavirus replication in mammalian systems.
• To elucidate the specific function of the alphavirus macrodomain in the viral life cycle.

Main Methods:

• Engineered a Sindbis virus (SINV) mutant with attenuated poly-ARH activity.
• Compared the replication efficiency of the mutant virus to wild-type (WT) SINV in mammalian cells.
• Assessed the translation of genomic RNA for both WT and mutant viruses.

Main Results:

• Viral poly-ARH activity is specifically required for high-titer replication of alphaviruses in mammalian cells.
• The engineered SINV mutant showed reduced capacity to establish productive infection despite efficient translation of genomic RNA.
• This indicates that poly-ARH activity is critical for later stages of the viral life cycle, beyond initial RNA translation.

Conclusions:

• Alphavirus replication is critically dependent on the poly-ADP-ribosylhydrolase activity of their encoded macrodomains.
• Viral macrodomain's poly-ARH function is essential for efficient productive infection and high viral titers.
• These findings deepen the understanding of ADP-ribosyltransferase biology and have implications for antiviral drug development targeting viral macrodomains.