Abstract
VP22, a structural protein of herpes simplex virus, exhibits unusual trafficking properties which we proposed might be exploited in gene and protein delivery applications. To pursue the use of the protein itself for cargo delivery into cells, we developed an expression system for the C-terminal half of VP22, residues 159-301 (VP22.C1), and purified the protein in high yields. Addition of short oligonucleotides (ODNs) induced the assembly of novel particles, which were regular spheres with a size range of 0.3 to 1.0 microm in diameter, incorporating both protein and ODN. Following the particles in living cells using fluorescently tagged ODNs, we show that they enter efficiently within 2-4 h, and reside stably in the cell cytoplasm for up to several days. Remarkably, however, light activation induced particle disruption and release of the protein and ODN to the nucleus and cytoplasm within seconds, a process that we have captured by time lapse microscopy. In addition to delivering antisense ODNs, ribozymes, and RNA/DNA hybrids, the VP22.C1 protein could also be modified to include peptides or proteins. These particles have the potential for delivery of a wide range of therapeutic agents in gene therapy and vaccine development.
Highlights
We observed that when extracts containing VP22 were applied to the medium of tissue culture cells, the protein could subsequently be detected within cells, accumulating in the
We previously reported upon the unusual trafficking properties of the HSV protein VP22 including the observation that when soluble cell-free extracts containing VP22 were applied to the medium of tissue culture cells, the protein could subsequently be detected within cells, accumulating in the nuclei [4]
Based upon these observations we proposed that VP22 protein itself could be exploited for the delivery of different types of cargoes
Summary
We observed that when extracts containing VP22 were applied to the medium of tissue culture cells, the protein could subsequently be detected within cells, accumulating in the. Initially the F-ODN was present in cytoplasmic particles as described above, while in the same cells 2 min later pronounced diffuse distribution together with nuclear accumulation could be observed.
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