Noncanonical Role for the Host Vps4 AAA+ ATPase ESCRT Protein in the Formation of Tomato Bushy Stunt Virus Replicase
Figure 5
Complex membranous compartments containing both tombusvirus VRCs and the ESCRT protein Vps4p fused with MT and HA tags in yeast.
(A) General organization of the complex membranous compartments containing the tombusvirus VRCs as seen in ultrathin sections of yeast cells (BY4741) processed by immunogold labeling and stained with uranyl acetate and lead citrate. Vps4p-MT-HA was detected with anti-HA and 10 nm gold particles (black arrows) and replicating TBSV repRNA with anti-dsRNA antibodies and 5 nm gold particles (white arrows). Asterisks mark the center of spherule-containing membranous structures that are connected to form a complex compartment where Vps4p and dsRNA are clustered together. (B) Membranous structures containing spherules and VRCs are surrounded by mitochondria (mi). Note that proximal location of mitochondria might be useful for robust virus replication that is dependent on ATP generation. This arrangement of VRCs and mitochondria are frequently observed in other viral replication platforms. (C–D) Membranous structures with spherules of ∼70 nm containing both Vps4 and dsRNA. The Vps4p-MT-HA and dsRNA containing areas are connected with a complex system of membranes. Bars, 100 nm.