Formation of (dA-dT)n cruciforms in Escherichia coli cells under different environmental conditions

Abstract

We have detected cruciform formation of (dA-dT)n inserts in Escherichia coli cells by analyzing the superhelical density of isolated plasmid DNA samples and by probing intracellular DNA with chloroacetaldehyde. The plasmids we used were pUC19 containing inserts of (dA-dT)n. The cruciforms appeared after cells underwent different stresses: inhibition of protein synthesis, anaerbiosis, and osmotic shock. At the same time, all these stimuli led to an increase in superhelical density of the control pUC19 plasmid DNA. Therefore, we suggest that the increase in plasmid superhelicity in response to different environmental stimuli entails the appearance of cruciform structures. The use of the (dA-dT)n units of various lengths made it possible to estimate the superhelical density of the plasmid DNA in vivo.