Base alterations in yeast induced by alkylating agents with differing Swain-Scott substrate constants

Area of Science:

• Molecular Biology
• Genetics
• Chemical Mutagenesis

Background:

• Alkylating agents are known mutagens that can induce DNA base alterations.
• Understanding the specific mutations induced by different agents is crucial for assessing genotoxicity and DNA repair mechanisms.

Purpose of the Study:

• To determine the base alterations induced by four distinct alkylating agents at the URA3 locus in Saccharomyces cerevisiae.
• To correlate the observed mutational spectra with the chemical properties (Swain-Scott substrate constants) of the alkylating agents.

Main Methods:

• Yeast cells (Saccharomyces cerevisiae) were treated with methyl methanesulfonate (MMS), ethyl methanesulfonate (EMS), N-nitroso-N-methylurea (MNU), and N-nitroso-N-ethylurea (ENU).
• Mutants resistant to 5-fluoroorotic acid were selected.
• DNA sequencing using the dideoxy chain termination method was performed on 125 forward mutations.

Main Results:

• Mutational spectra differed significantly based on the alkylating agent's Swain-Scott substrate constant (s value).
• Five hotspots for G.C----A.T transitions were identified, with variations among the agents.
• MMS induced the widest array of mutations, while agents with lower s values showed restricted mutation types, primarily G.C----A.T transitions and A.T----T.A transversions.

Conclusions:

• The type and spectrum of DNA mutations are influenced by the chemical properties of alkylating agents.
• Observed mutations are consistent with proposed mechanisms involving O6-alkylation of guanine, alkylation of thymine, and N1-alkylation of adenosine.