Cell to Cell Contact Is Not Required for Bystander Cell Killing by Escherichia coli Purine Nucleoside Phosphorylase

• Published in: The Journal of biological chemistry 1998-01, Vol.273 (4), p.2322-2328
• Main Authors: Hughes, Brian W., King, Scott A., Allan, Paula W., Parker, William B., Sorscher, Eric J.
• Format: Article
• Language: English
• Subjects: Affinity Labels; Animals; Cell Communication; Cell Death; Cell Division; Culture Media; Escherichia coli - enzymology; Humans; Mice; Prodrugs - metabolism; Prodrugs - pharmacology; Purine Nucleosides - metabolism; Purine Nucleosides - pharmacology; Purine-Nucleoside Phosphorylase - metabolism; Purine-Nucleoside Phosphorylase - pharmacology; Purines - metabolism; Purines - pharmacology; Thioinosine - analogs & derivatives; Thioinosine - pharmacology; Tumor Cells, Cultured
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.273.4.2322

Expression of Escherichia coli purine nucleoside phosphorylase (PNP) activates prodrugs and kills entire populations of mammalian cells, even when as few as 1% of the cells express this gene. This phenomenon of bystander killing has been previously investigated for herpes simplex virus-thymidine kinase (HSV-TK) and has been shown to require cell to cell contact. Using silicon rings to separate E. coli PNP expressing cells from non-expressing cells sharing the same medium, we demonstrate that bystander cell killing by E. coli PNP does not require cell-cell contact. Initially, cells expressing E. coli PNP convert the non-toxic prodrug, 6-methylpurine-2′-deoxyriboside (MeP-dR) to the highly toxic membrane permeable toxin, 6-methylpurine (MeP). As the expressing cells die, E. coli PNP is released into the culture medium, retains activity, and continues precursor conversion extracellularly (as determined by reverse phase high performance liquid chromatography of both prodrug and toxin). Bystander killing can also be observed in the absence of extracellular E. coli PNP by removing the MeP-dR prior to death of the expressing cells. In this case, 100% of cultured cells die when as few as 3% of the cells of a population express E. coli PNP. Blocking nucleoside transport with nitrobenzylthioinosine reduces MeP-dR mediated cell killing but not MeP cell killing. These mechanisms differ fundamentally from those previously reported for the HSV-TK gene.