DOI: 10.5061/dryad.g1jwstr2k
title: Data from: Coordination of the host Vps4-Vta1 complex and the viral core protein Ac93 facilitates entry of Autographa californica multiple nucleopolyhedrovirus budded virions

description: The endosomal sorting complex required for transport (ESCRT) is a protein
 machine mediating membrane scission. In intraluminal vesicles (ILVs)
 formation, ESCRT-0 targets cargoes and recruits ESCRT-I/-II to create
 membrane invagination, whereas ESCRT-III coordinates with the AAA ATPase
 Vps4 and its cofactor Vta1 to catalyze the membrane fission. Recently,
 ESCRT-I/-III and Vps4 were found to involve in entry of the baculovirus
 Autographa californica multiple nucleopolyhedrovirus (AcMNPV). However,
 the necessity of other ESCRT components and the interplay of viral
 proteins and ESCRTs in regulating the virus entry remain elusive. Here, we
 identified ESCRT-0 (Hse1 and Vps27), ESCRT-II (Vps22, Vps25, and Vps36),
 and Vta1 of S. frugiperda. RNAi depletion of Vta1 but not the components
 of ESCRT-0 or ESCRT-II in Sf9 cells significantly reduced budded virus
 (BV) production. Quantitative PCR together with confocal microscopy
 analyses indicated that Vta1 was required for internalization and
 endosomal trafficking of BV. In the late phase of infection, although Vps4
 and Vta1 were both distributed to the nucleus and at the plasma membrane,
 depletion of Vta1 did not affect BV release. Further analysis revealed
 that 7 of 14 BV envelope proteins (Ac75, Ac93, E25, F-like, P33, P48, and
 vUbiquitin) interacted with Vps4 and Vta1. Intriguingly, Ac93 adopted a
 similar mode as ESCRT-III proteins to interact with the MIT domains of
 Vps4 and Vta1 via its C-terminal MIM1 motif, and the interactions were
 necessary for BV internalization. Together, our studies highlight the
 coordination of Vps4-Vta1 and Ac93, and probably other BV envelope
 proteins, in facilitating entry of AcMNPV.
description: # Coordination of the host Vps4-Vta1 complex and the viral core protein
 Ac93 facilitates entry of Autographa californica multiple
 nucleopolyhedrovirus budded virions
 [https://doi.org/10.5061/dryad.g1jwstr2k](https://doi.org/10.5061/dryad.g1jwstr2k) ## Description of the data and file structure The data files contain the LacZ ((β-galactosidase) and GUS (β-glucuronidase) activities and the virus titers. The extended data file contains the supplemental figures. For measurement of LacZ and GUS activities, a brief description of the methods as below (detailed Materials and Methods see the associated paper): Sf9 cells in 12-well plates (200000 cells/well) were transfected with the dsRNA of gfp, Vps4, or Vta1 (5 g/well for each gene) using the standard CaPO4 precipitation method. At 48 h post-transfection (p.t.), the cells were transfected again with the bacmid DNA of AcMNPV-LacZGUS (4 g/well). After transfecting the viral DNA for 24 h, the cells were lysed using 1% Triton X-100 containing 2 mg/ml Chlorophenol red--D-galactopyranoside (Roche Diagnostics GmbH) or 1mM 4-Nitrophenyl -D-glucuronide (Merck) to measure the activity of -Gal or -Gluc. For measurement of the titers of wild-type virus (AcMNPV) and vAc93-HA-Cm, a brief description of the methods as below (detailed Materials and Methods see the associated paper): Sf9 cells were infected with each of the virus (MOI=5) and the virus titers were measured by a 50% tissue culture infection dose (TCID50) assay at indicated time points. ### Files and variables The data files listed as: 1. File 1: LacZ activities 2. File 2: GUS activities 3. File 3: Virus titers of vAc93-HA-Cm 4. File 4: Virus titers of wild-type AcMNPV The extended data file contains 6 figures: 1. Extended Figure S1: Amino acid sequence alignment of the homologs of ESCRT-0 components Hse1 and Vps27. 2. Extended Figure S2: Amino acid sequence of Hse1 and Vps27 of *S*. *frugiperda*. 3. Extended Figure S3: The activities of -galactosidase and -glucuronidase. 4. Extended Figure S4: Yeast-two hybrid analysis of the interaction between Vps4 or Vta1 of *S*. *frugiperda* and BV-envelope proteins of AcMNPV. 5. Extended Figure S5: Western blot analysis of the expression of Nm- and Cm-fused Vps4 and Vta1 of *S*. *frugiperda*, and BV envelope proteins of AcMNPV. 6. Extended Figure S6: One-step growth curves of wild-type (wt) AcMNPV or Ac93-HA-Cm-repaired *ac93*-knockout virus.