The Herpesvirus capsid surface protein, VP26, and the majority of the tegument proteins are dispensable for capsid transport toward the nucleus

Abstract

Upon entering a cell, alphaherpesvirus capsids are transported toward the minus ends of microtubules and ultimately deposit virus DNA within the host nucleus. The virus proteins that mediate this centripetal transport are unknown but are expected to be either viral tegument proteins, which are a group of capsid-associated proteins, or a surface component of the capsid itself. Starting with derivatives of pseudorabies virus that encode a fluorescent protein fused to a structural component of the virus, we have made a collection of 12 mutant viruses that lack either the VP26 capsid protein or an individual tegument protein. Using live-cell fluorescence microscopy, we tracked individual virus particles in axons following infection of primary sensory neurons. Quantitative analysis of the VP26-null virus indicates that this protein plays no observable role in capsid transport. Furthermore, viruses lacking tegument proteins that are nonessential for virus propagation in cell culture were also competent for axonal transport. These results indicate that a protein essential for viral propagation mediates transport of the capsid to the nucleus.

FIG. 1.

Single-step growth kinetics of VP26- and VP26-null fluorescent viruses. Virions were harvested from media (dashed lines, open symbols) and PK15 cells (solid lines, filled symbols) at the indicated times. Squares, PRV-GS962; circles, PRV-GS1205.

FIG. 2.

VP26-null capsid retrograde axonal transport. Example of virus particle transport resulting from infection of dorsal root sensory neurons with PRV-GS1205 and imaged within the first hour postinfection. A montage of eight frames from a subregion of a time-lapse recording are shown (see movie M1 in the supplemental material for the entire time-lapse recording). Each frame is a 200-ms exposure representing every fourth frame of the original recording (the montage represents a 6.4-s time window). A single VP26-null capsid (mRFP1-VP1/2) complex is shown in the montage. The frames are each 2.7 μm × 15.2 μm.

FIG. 3.

Analysis of capsid transport velocities. Histogram of retrograde transport velocities of individual virus particles resulting from infections of dorsal root sensory neurons with PRV-GS962 (a) and PRV-GS1205 (b). The smooth curve in each panel represents the best-fit Gaussian curve for each sample.