MAMMALIAN-CELL PROCESSING OF A UNIQUE URACIL RESIDUE IN SIMIAN VIRUS-40 DNA

The processing of a unique uracil in DNA has been studied In mammalian cells. A synthetic oligodeoxyrlbonucleotide carrying a potential Bgl II restriction site, where one base has been substituted with a uracil, was inserted in the early intron of SV4O genome. Various heteroduplexes were constructed In such a manner that the restitution of an active BgI II restriction site corresponds in each case to the specific substitution of the uracll by one of the four bases normally present in the DNA. DNA cuts by this restriction enzyme in one or several constructed heteroduplexes Immediately determine the type of base pair substitution produced at the site of the U residue. When the uracil is Inserted opposite a purine it is fully repaired ; when facing a guanine it is replaced by a cytosine and opposite an adenlne it is replaced by a thymine. These resuits Indicate the error-free repair of uracil when it appears In the cell with the usual mechanisms such as cytosine deamination or Incorporation of dUTP in place of dTTP during replication. When the uracil is inserted opposite a pyrimidine no error free repair at all is detected for U:C or U:T mismatches. it appears, moreover, that in approximately 18% of the cases U:T mismatch leads to a C:G base pairing. In the majority of the U:pyrimidine mismatches, mutations occur in the vicinity of the uracil, including base substitutions and frameshifts by addition of one or several bases. © 1990 Oxford University Press.