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Transcriptionally driven cruciform formation in vivo
A Dayn1, S Malkhosyan, S M Mirkin
1Department of Genetics, University of Illinois, Chicago 60612.
Nucleic Acids Research
|November 25, 1992
Summary
Transcriptionally driven negative supercoiling in E.coli induces d(A-T)n cruciform formation. This DNA structure transition is linked to promoter activity and transcription induction by IPTG.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- DNA can adopt non-canonical structures like cruciforms.
- These structures are often associated with specific DNA sequences, such as d(A-T)n repeats.
- The in vivo formation of such structures is influenced by cellular processes like transcription.
Purpose of the Study:
- To investigate the formation of d(A-T)n cruciforms in E. coli cells.
- To determine the role of transcription and promoter activity in cruciform formation.
- To understand the mechanism by which transcription influences DNA structure in vivo.
Main Methods:
- Utilized E. coli as a model system.
- Engineered plasmids containing d(A-T)16 sequences adjacent to trc promoters.
- Employed chloroacetaldehyde probing to detect modified DNA bases, indicating cruciform formation.
- Analyzed DNA base modifications to confirm intracellular cruciform structures.
Main Results:
- Transcription induction by IPTG triggered d(A-T)16 cruciform formation when located upstream or between divergent trc promoters.
- Cruciforms formed even without IPTG when d(A-T)16 was situated between two divergent promoters.
- Increased promoter activity, indicated by promoter opening, correlated with enhanced cruciform formation.
Conclusions:
- Transcriptionally induced negative supercoiling is a key driver for in vivo cruciform formation.
- Promoter activity directly influences the propensity of d(A-T)n sequences to form cruciforms.
- The cellular transcription machinery plays a significant role in shaping DNA topology and structure.