Molecular basis for the interaction between rabies virus phosphoprotein P and the dynein light chain LC8: dissociation of dynein-binding properties and transcriptional functionality of P

Summary auto-generated

This study investigated how rabies virus phosphoprotein P interacts with dynein light chain LC8, a cellular protein involved in motor transport along microtubules. Using co-immunoprecipitation and yeast two-hybrid screening, the researchers mapped the LC8-binding site to amino acids 139–151 of P protein. Site-directed mutagenesis revealed that residues D143 and Q147 are critical for LC8 binding. A structural model based on the known LC8 crystal structure suggests P binds LC8 similarly to other cellular LC8-binding partners. Importantly, the P–LC8 interaction is not required for viral transcription, as demonstrated using a minigenome assay with a double mutant P(D143A–Q147A) that cannot bind LC8 but retains normal transcriptional activity. The authors propose that this interaction may facilitate viral axonal transport via the dynein motor complex, potentially explaining how rabies virus spreads from infection sites to the central nervous system.

Key findings

The LC8-binding site on rabies virus phosphoprotein P is located at amino acids 139–151, with D143 and Q147 being essential residues for interaction
The P–LC8 interaction occurs through a mechanism similar to other cellular LC8-binding partners, following a conserved binding motif
LC8 binding is not required for viral transcription, as P mutants unable to bind LC8 retain full transcriptional activity in minigenome assays
The interaction may facilitate viral transport along microtubules via the dynein motor complex, potentially explaining axonal migration of rabies virus to the central nervous system
Other viral proteins from rotavirus, poliovirus, and herpesviruses contain similar LC8-binding motifs, suggesting this interaction may be a common mechanism for viral motor transport

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Abstract

The lyssavirus phosphoprotein P is a co-factor of the viral RNA polymerase and plays a central role in virus transcription and replication. It has been shown previously that P interacts with the dynein light chain LC8, which is involved in minus end-directed movement of organelles along microtubules. Co-immunoprecipitation experiments and the two-hybrid system were used to map the LC8-binding site to the sequence ¹³⁹RSSEDKSTQTTGR¹⁵¹. Site-directed mutagenesis of residues D¹⁴³ and Q¹⁴⁷ to an A residue abolished binding to LC8. The PLC8 association is not required for virus transcription, since the double mutant was not affected in its transcription ability in a minigenome assay. Based on the crystal structure of LC8 bound to a peptide from neuronal nitric oxide synthase, a model for the complex between the peptide spanning residues 140150 of P and LC8 is proposed. This model suggests that P binds LC8 in a manner similar to other LC8 cellular partners.