For decades, it has been recognized that transcribing along the DNA double helix by a RNA polymerase can enhance localized DNA supercoiling. This process has been elegantly explained by a """"""""twin-supercoiled- domain"""""""" model in which positive DNA supercoils are generated ahead of a translocating RNA polymerase and negative supercoils behind it. However, detailed mechanisms at the molecular level are still unavailable. For instance, how DNA topological barriers affect transcription-coupled DNA supercoiling (TCDS) has not been fully understood. The long-term goal of the proposed research is to understand the molecular mechanisms and biological functions of TCDS and DNA topological barriers. The objectives of this application are to determine what properties make certain DNA-looping and -wrapping proteins function as topological barriers and block supercoil diffusion, to study how DNA topological barriers regulate TCDS in E. coli cells, and to examine how localized, transient TCDS activates gene expression in vivo. Our central hypothesis is that TCDS by RNA polymerases is a major chromosome remodeling force in E. coli cells where the cells use nucleoprotein-based, topological barriers to confine TCDS to localized regions and, as a result, greatly influence the nearby, coupled DNA transactions. Our hypothesis has been formulated on the basis of strong preliminary data produced in our laboratory and will be tested by pursuing the following three specific aims: 1) to determine molecular mechanisms of DNA topological barriers and examine how the DNA topological barriers confine supercoils to localized regions and affect the efficiency of TCDS;2) to examine the role of DNA topological barriers in TCDS in E. coli cells;3) to investigate how transient, dynamic TCDS activates transcription in E. coli cells and how DNA topological barriers affect the efficiency of transcription activation by TCDS. Information from the proposed experiments will ultimately provide us with a better understanding of the mechanisms of TCDS, especially the roles of DNA topological barriers in TCDS and the coupled gene transcription and expression.

Public Health Relevance

Almost all DNA molecules inside cells including human cells are supercoiled. In fact, DNA supercoiling affects all DNA transactions, i.e., DNA replication, recombination, and transcription, and greatly influences genomic stability and susceptibility to cancer and certain hereditary diseases, such as fragile X syndrome, Huntington's disease, and autism. The significance of this research stems from its potential to provide a basis for better understanding of essential biological processes: gene transcription and expression.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Academic Research Enhancement Awards (AREA) (R15)
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Special Emphasis Panel (ZRG1)
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Carter, Anthony D
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Florida International University
Schools of Arts and Sciences
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Yan, Yan; Ding, Yue; Leng, Fenfei et al. (2018) Protein-mediated loops in supercoiled DNA create large topological domains. Nucleic Acids Res 46:4417-4424
Yan, Yan; Leng, Fenfei; Finzi, Laura et al. (2018) Protein-mediated looping of DNA under tension requires supercoiling. Nucleic Acids Res 46:2370-2379
Liu, Yingting; Hua, Zhi-Chun; Leng, Fenfei (2018) DNA Supercoiling Measurement in Bacteria. Methods Mol Biol 1703:63-73
Dages, Samantha; Dages, Kelley; Zhi, Xiaoduo et al. (2018) Inhibition of the gyrA promoter by transcription-coupled DNA supercoiling in Escherichia coli. Sci Rep 8:14759
Liu, Yingting; Berrido, Andrea M; Hua, Zi-Chun et al. (2017) Biochemical and biophysical properties of positively supercoiled DNA. Biophys Chem 230:68-73
Zhi, Xiaoduo; Dages, Samantha; Dages, Kelley et al. (2017) Transient and dynamic DNA supercoiling potently stimulates the leu-500 promoter in Escherichia coli. J Biol Chem 292:14566-14575
Fulcrand, Geraldine; Chapagain, Prem; Dunlap, David et al. (2016) Direct observation of a 91 bp LacI-mediated, negatively supercoiled DNA loop by atomic force microscope. FEBS Lett 590:613-8
Sun, Pengfei; Leeson, Cristian; Zhi, Xiaoduo et al. (2016) Characterization of an epoxide hydrolase from the Florida red tide dinoflagellate, Karenia brevis. Phytochemistry 122:11-21
Fulcrand, Geraldine; Dages, Samantha; Zhi, Xiaoduo et al. (2016) DNA supercoiling, a critical signal regulating the basal expression of the lac operon in Escherichia coli. Sci Rep 6:19243
Leng, Fenfei (2016) Protein-induced DNA linking number change by sequence-specific DNA binding proteins and its biological effects. Biophys Rev 8:123-133

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