Biochemical and biological properties of 5-bromotubercidin: differential effects on cellular DNA-directed and viral RNA-directed RNA synthesis.
We have studied the biochemical and biological properties of 5-bromotubercidin (4-amino-5-bromo-7-beta-d-ribofuranosyl-pyrrolo [2,3-d]pyrimidine) (BrTu), a synthetic analogue of the highly cytotoxic pyrrolo[2,3-d]pyrimidine ribonucleoside antibiotic tubercidin (Tu) that interferes with numerous cellular processes, and has been shown to possess biological specificity and selectivity. Thus, BrTu entered the mammalian cell nucleotide pool by phosphorylation, was incorporated into RNA in an unmodified form and, as a consequence, reversibly inhibited (15 microM) mammalian cell growth and the synthesis of high-molecular-weight cellular RNA species (i.e., mRNA and rRNA). However, BrTu (300 microM) did not inhibit picornavirus RNA synthesis or multiplication, and thus discriminated between virus RNA-dependent and all forms of DNA-dependent RNA synthesis whether of cellular or viral origin; because of this BrTu should prove valuable as a metabolic probe for studying the cell-virus relationship. Furthermore, BrTu is a substrate for adenosine kinase (K(m)=24 microM), and is also its potent inhibitor (K(i)=0.93 microM); thus, low concentrations of BrTu (1.5 microM), which did not inhibit cell growth, blocked phosphorylation and the cellular uptake of other, highly cytotoxic pyrrolo-pyrimidine nucleoside analogues (e.g., tubercidin). This block in cellular uptake and incorporation of toxic analogues was associated with the protective effect of BrTu against cell killing by the analogues, providing a mechanism by which BrTu and these analogues can, as we reported elsewhere [J. Virol.1999, 73, 6444], be used for the selective inactivation of replicating picornaviruses.
Brdar
Branko
B
Department of Molecular Biology, Rudjer Boskovic Institute, Bijenicka 54, 10000 Zagreb, Croatia. brdar@rudjer.irb.hr
Reich
Edward
E
D000143:Acids; D000248:Adenosine Kinase / Q000037:antagonists & inhibitors / Q000378:metabolism; D000818:Animals; D000998:Antiviral Agents / Q000737:chemistry / Q000494:pharmacology; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D004247:DNA / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008099:Liver / Q000187:drug effects / Q000201:enzymology; D051379:Mice; D015394:Molecular Structure; D047428:Protein Kinase Inhibitors / Q000737:chemistry / Q000494:pharmacology; D012367:RNA, Viral / Q000096:biosynthesis*; D011817:Rabbits; D012995:Solubility; D013329:Structure-Activity Relationship; D014372:Tubercidin / Q000031:analogs & derivatives* / Q000737:chemistry / Q000494:pharmacology
D000143:Acids; D000998:Antiviral Agents; D047428:Protein Kinase Inhibitors; D012367:RNA, Viral; C000323:bromotubercidin; D004247:DNA; D000248:Adenosine Kinase; D014372:Tubercidin