Premium
Metabolic Fate of [ 3 H]1‐β‐ d ‐Arabinofuranosyl‐5‐[( E )‐2‐bromovinyl]uracil in Herpes Simplex Virus Type 1‐Infected Cells
Author(s) -
Ashida Noriyuki,
Sakata Shinji,
Machida Haruhiko
Publication year - 1997
Publication title -
microbiology and immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.664
H-Index - 70
eISSN - 1348-0421
pISSN - 0385-5600
DOI - 10.1111/j.1348-0421.1997.tb01955.x
Subject(s) - micrococcal nuclease , dna , thymidine , biology , herpes simplex virus , microbiology and biotechnology , uracil , biochemistry , virus , virology , chromatin , nucleosome
The metabolic fate of 1‐β‐ d ‐arabinofuranosyl‐5‐[( E )‐2‐bromovinyl]uracil (BV‐araU) in herpes simplex virus type 1‐infected cells was studied using tritium‐labeled BV‐araU. [ 3 H]BV‐araU was selectively taken‐up by infected cells. Approximately 10% of the total uptake of [ 3 H]BV‐araU was recovered from the acid‐insoluble fraction at any time post‐infection. Both cellular uptake of [ 3 H]BV‐araU and its incorporation into the acid‐insoluble fraction increased with increasing incubation time through 8 hr post‐infection. Uptake of [ 3 H]BV‐araU and its incorporation into the acid‐insoluble fraction also increased proportionally to the duration of exposure to [ 3 H]BV‐araU. An alkaline sucrose gradient sedimentation analysis revealed that the radioactive DNA obtained from cells pulse‐labeled with [ 3 H]BV‐araU were small DNA fragments which remained at the top following a chasing period in isotope‐free medium, whereas that pulse‐labeled with [ 3 H]thymidine was chased to a fraction of high molecular weight DNA. Nuclease P 1 digestion reduced 99% of the [ 3 H]BV‐araU‐labeled DNA extracted from infected cells to a low molecular weight. Following digestion of [ 3 H]BV‐araU‐labeled DNA with micrococcal nuclease and spleen exonuclease, all of the radioactivity was recovered as [ 3 H]BV‐araU 3′‐monophosphate. Thus, BV‐araU strongly inhibits the elongation of viral DNA strands as demonstrated by the alkaline sucrose gradient sedimentation analysis, whereas at least a portion of the [ 3 H]BV‐araU is incorporated inside viral DNA strands in infected cells.