Induction of mutation in mouse FM3A cells by N4-aminocytidine-mediated replicational errors.

Abstract

To explore the potential use of a nucleoside analog, N4-aminocytidine, in studies of cellular biology, the mechanism of mutation induced by this compound in mouse FM3A cells in culture was studied. On treatment of cells in suspension with N4-aminocytidine, the mutation to ouabain resistance was induced. The major DNA-replicating enzyme in mammalian cells, DNA polymerase a, was used to investigate whether the possible cellular metabolite of N4-aminocytidine, N4-aminodeoxycytidine 5'-triphosphate (dCa`TP), can be incorporated into the DNA during replication. Using [3H]dCIUTP in an in vitro DNA-synthesizing system, we were able to show that this nucleotide analog can be incorporated into newly formed DNA and that it can serve as a substitute for either dCTP or dTTP. dCarTP in the absence of dCTP maintained the activated calf thymus DNA-directed polymerization of deoxynucleoside triphosphates as efficiently as in its presence. Even in the presence of dCTP, dCarTP was incorporated into the polynucleotide. When dCaITP was used as a single substrate in the poly(dA)-oligo(dT)-directed polymerase reaction, it was incorporated into the polynucleotide fraction. The extent of incorporation was 4% of that of dTTP incorporation when dTTP was used as a single substrate. Even in the presence of dTTP, dCaNTP incorporation was observed. A copolymer containing N4-aminocytosine residues was shown to incorporate guanine residues opposite the N4-aminocytosines. However, we were unable to observe adenine incorporation opposite N4-aminocytosine in templates. These cell-free experiments show that an AT-to-GC transition can take place in the presence of dCarTP during DNA synthesis, strongly suggesting that the mutation induced in the FM3A cells by N4-aminocytidine is due to replicational errors.

Template properties of mutagenic cytosine analogues in reverse transcription

“…It was reported that the triphosphates of N 4 -hydroxycytidine (ho 4 C), N 4 -methoxycytidine (mo 4 C) and N 4 -methylcytidine (m 4 C) exhibited ambiguous properties in polymerization (22–24), and N 4 -aminodeoxycytidine triphosphate can be incorporated in place of both dCTP and dTTP (25,26). We have found that 4-semicarbazidocytidine triphosphate, a N 4 -aminocytidine derivative, can be incorporated as UTP by phage RNA polymerase (27) (Figure 1).…”

Intramolecular tautomerization and the conformational variability of some classical mutagens – cytosine derivatives: quantum chemical study

“…DCyt і 4 meCyt, фе но ме но логічні ефек ти му та ген ної дії яких на й кра ще з-поміж спо лук ана логічної будо ви пред став ле но в літе ра турі [6][7][8][9][10][11][12]. Пред мет вив чен ня -фізи ко-хімічна при ро да про цесів внутрішньо мо ле ку ляр ної та у то ме ри зації зазначених му та генів та їхні кон фор маційні влас ти вості, включа ю чи струк тур ну не жорсткість.…”

N,N-Dimethyl-5-methoxymethyl-2'-deoxycytidine

“…The cis relationship between the N 4 substituent and N 3 appears to be a common feature among N4-substituted 2'-deoxycytidine analogues (Takahashi et al, 1988;Jia et al, 1990b;Audette et al, 1997;Audette et al, 1998).…”