Fig. 1: PVCs can be reprogrammed for custom protein delivery in eukaryotic cells.
From: Programmable protein delivery with a bacterial contractile injection system
a, Schematic of the P. asymbiotica PVCpnf locus containing 16 structural and accessory genes (in blue and violet, respectively) followed by two payload genes (Pdp1 and Pnf, in red) and four putative regulatory genes (in orange). PVC illustrations are approximations and are not drawn to scale. b, Proposed mechanism of PVC-mediated protein delivery. PVCs probably recognize target cells via tail fibres (Pvc13), leading to a contraction of the sheath mechanism that drives a spike through the cellular membrane. Payload proteins are then thought to enter the cell via disassembly of the spike–tube complex. c, PVCs can be purified from E. coli. The PVCpnf locus was transformed into E. coli and intact PVC particles were isolated. The resulting PVC preparations were then run on a denaturing SDS–PAGE gel and imaged with negative-stain TEM. Scale bars, 100 nm. d, PVCs can be visualized binding to target cells. PVC particles containing epitope-tagged sheath proteins (Pvc2) were incubated with Sf9 cells and binding was visualized via immunofluorescence. Scale bar, 100 μm. e, Non-native payloads can be loaded into PVC particles. The Pdp1 packaging ___domain (PD) (Extended Data Fig. 3b,c) was fused to novel proteins and loading was determined via denaturing western blot. Pvc12 (baseplate) served as a loading control. WT, wild-type. f, Wild-type PVC particles kill cultured insect cells. PVC-mediated cytotoxicity required both the action of the putative target recognition gene (pvc13; tail fibre) and payload loader (pvc15; ATPase). Scale bar, 200 μm. g, PVCs can deliver a non-native payload (loaded as in e) into target cells. Scale bar, 200 μm. Data in f,g are mean ± s.d. with n = 3 biological replicates; one-way ANOVA with Bonferroni post hoc test. ****P < 0.0001.
