Cell death induced by the herpes simplex virus-1 thymidine kinase delivered by human immunodeficiency virus-1-based virus-like particles.
HIV-1 Nef incorporates into virions at low levels, likely about 10 molecules per viral particle. Here, we describe a Nef mutant (Nef7) apparently showing more than 100-fold higher efficiency of virion incorporation. Interestingly, Nef7 can act as a cargo molecule for protein delivery into the cells, as its virion incorporation appeared conserved even upon C-terminal fusion with proteins of up to 30 kDa. This was demonstrated first by assessing the intracellular fluorescence of cells challenged with lentivirus-based virus-like particles (VLPs) pseudotyped with the vesicular stomatitis virus envelope glycoprotein (VSV-G) and incorporating Nef7 fused with the green fluorescent protein. Furthermore, the biologic activity of products delivered by Nef7-based VLPs was demonstrated by tagging Nef7 with the herpes simplex virus-1 thymidine kinase (HSV-1 TK). In fact, we observed that both cell lines and primary human macrophages challenged with (VSV-G) Nef7/TK VLPs died after 5 to 7 days of treatment with ganciclovir (GCV). In sum, our findings support the notion that Nef7-based VLPs can be considered platforms for original systems of protein delivery. In particular, the here- described Nef7/TK VLPs represent a first applicative example opening the way toward new HSV-1 TK/GCV-based cell suicide therapies circumventing cell gene engineering procedures.
Peretti
Silvia
S
Department of Infectious, Parasitic, and Immune-Mediated Diseases, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Rome, Italy.
Schiavoni
Ilaria
I
Pugliese
Katherina
K
Federico
Maurizio
M
D015153:Blotting, Western; D002460:Cell Line; D003902:Detergents / Q000494:pharmacology; D015774:Ganciclovir / Q000494:pharmacology; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods; D049452:Green Fluorescent Proteins / Q000378:metabolism; D015497:HIV-1 / Q000235:genetics*; D018259:Herpesvirus 1, Human / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D016086:Lentivirus / Q000235:genetics; D008264:Macrophages / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism; D021962:Membrane Microdomains; D008957:Models, Genetic; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D012190:Retroviridae / Q000235:genetics; D013937:Thymidine Kinase / Q000235:genetics*; D013997:Time Factors; D014759:Viral Envelope Proteins / Q000378:metabolism
D003902:Detergents; C028391:G protein, vesicular stomatitis virus; D008562:Membrane Glycoproteins; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; D049452:Green Fluorescent Proteins; D013937:Thymidine Kinase; D015774:Ganciclovir