Intercellular trafficking of VP22-GFP fusion proteins.
The herpes simplex virus protein VP22 exhibits the unusual property of intercellular transport whereby after being synthesised in a subpopulation of cells, in which it is largely cytoplasmic, the protein is transported to adjacent cells where it accumulates mainly in the nucleus. Here we examine the transport of a fusion protein consisting of VP22 linked to the green fluorescent protein (GFP). Intercellular transport, nuclear accumulation and chromatin binding of VP22-GFP could be detected by intrinsic GFP fluorescence in fixed cells. However, while the cytoplasmic localisation of VP22-GFP could be detected in live cells actively synthesising the protein, we were unable to detect intercellular transport by intrinsic GFP fluorescence in livecells, indicating that the levels of transported protein may be below those required for live detection, or that GFP fluorescence was quenched. The use of antibody to GFP was more sensitive than intrinsic GFP fluorescence and allowed ready detection of transport and nuclear accumulation of VP22-GFP. Intercellular transport was also confirmed in coplating experiments. Consistent with previous results showing a requirement for the C-terminus of VP22 in transport of the native protein, a fusion protein consisting of GFP linked to the N-terminal 1-192 residues of VP22 failed to transport between cells. The results support the proposal that VP22 has the ability to transport cargo proteins between cells and that it has significant potential in the field of gene therapy.
Elliott
G
G
Marie Curie Research Institute, Oxted, Surrey, UK.
O'Hare
P
P
D000818:Animals; D001692:Biological Transport; D019556:COS Cells; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods*; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D015678:Viral Structural Proteins / Q000378:metabolism*
D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D015678:Viral Structural Proteins; C073413:herpes simplex virus type 1 protein VP22; D049452:Green Fluorescent Proteins