Penetration of mouse fibroblasts by the 5' phosphate of 9 β D arabinofuranosyladenine and incorporation of the nucleotide into DNA

9 β D Arabinofuranosyladenine 5' monophosphate (ara AMP) is more lethal to mouse fibroblasts (L cells) than the identical exogenous concentration of 9 β D arabinofuranosyladenine (araA). [³H, ³²P]AraAMP (0.1 mM) was taken into L cells for 4 hr in the presence of a large excess of ³²P(i). The radioactivity was subsequently found mainly in the adenine nucleotides in the acid soluble fraction and in the cell DNA. The cellular concentration of 9 β D arabinofuranosyl adenine 5' triphosphate (araATP) exceeded 2 μM. More than 90% of the ³H was associated with araA in the nucleotides. After degradation of the adenine containing triphosphates with apyrase, the adenine mononucleotides were separated by thin layer electrophoresis and chromatography. All of the ³²P and 97% of the ³H were associated with ara AMP. The small amounts of ³H and ³²P in the acid insoluble material were similar during a 4 hr incubation. The DNA fraction was degraded enzymatically to 5' mononucleotides. Both ³²P and ³H were associated predominantly with 5' dAMP. Most of the ³H was in araA, detected after dephosphorylation. Enzymatic degradation of the DNA fraction to 3' mononucleotides and fractionation revealed ³H primarily in the 3' adenine nucleotide and ³²P in each of the deoxynucleoside 3' monophosphates. After dephosphorylation of the 3' mononucleotides, 93% of the ³H was found in araA. These results suggest that small amounts of araAMP penetrated the cell as an intact nucleotide, were further phosphorylated to the triphosphate, and subsequently incorporated in internucleotide linkage into DNA.