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Direct observation of dynamic mechanical regulation of DNA condensation by environmental stimuli.

ABSTRACT: Gene delivery is a promising way to treat hereditary diseases and cancer; however, there is little understanding of DNA:carrier complex mechanical properties, which may be critical for the protection and release of nucleic acids. We applied optical tweezers to directly measure single-molecule mechanical properties of DNA condensed using 19-mer poly-L-lysine (PLL) or branched histidine-lysine (HK) peptides. Force-extension profiles indicate that both carriers condense DNA actively, showing force plateaus during stretching and relaxation cycles. As the environment such as carrier concentration, pH, and the presence of zinc ions changes, DNA:HK complexes showed dynamically regulated mechanical properties at multiple force levels. The fundamental knowledge from this study can be applied to design a mechanically tailored complex which may enhance transfection efficiency by controlling the stability of the complex temporally and spatially.

SUBMITTER: Lee A

PROVIDER: S-EPMC5517097 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature

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Direct observation of dynamic mechanical regulation of DNA condensation by environmental stimuli.

Lee Amy A Karcz Adam A Akman Ryan R Zheng Tai T Kwon Sara S Chou Szu-Ting ST Sucayan Sarah S Tricoli Lucas J LJ Hustedt Jason M JM Leng Qixin Q Kahn Jason D JD Mixson A James AJ Seog Joonil J

Angewandte Chemie (International ed. in English) 20140821 40

Gene delivery is a promising way to treat hereditary diseases and cancer; however, there is little understanding of DNA:carrier complex mechanical properties, which may be critical for the protection and release of nucleic acids. We applied optical tweezers to directly measure single-molecule mechanical properties of DNA condensed using 19-mer poly-L-lysine (PLL) or branched histidine-lysine (HK) peptides. Force-extension profiles indicate that both carriers condense DNA actively, showing force ...[more]

PMID: 25146232

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Project description:Nucleoid remodeling facilitated by DNA supercoiling results in changes in nucleoid configuration and involves nucleoid-associated proteins (NAPs) and structural maintenance of chromosomes (SMC) proteins among others. Changes in nucleoid configurations regulated by NAPs are synchronized with cellular adaptation and influence the simultaneous expression of several genes. HU, a ubiquitous bacterial histone-like protein, is among the most conserved and abundant NAPs in eubacteria. In Escherichia coli, HU forms dimers by HUα self-association (HUαα) or by HUα-HUβ interactions (HUαβ). HUα is mostly expressed during lag and early exponential growth phase and HUβ is expressed only during the later exponential and stationary phase, pointing to distinct HUαα/DNA and HUαβ/DNA packaging of the nucleoid in regulating expression patterns during growth and stasis. Mutations or the deletion of HU transform the E. coli nucleoid to a different form and alter overall transcription program; thus, HU interactions with DNA directly affect global gene regulation. Recently, analysis of high-resolution contact maps of the E. coli nucleoid revealed an important role of HU to promote long-range DNA-DNA contacts within the nucleoid. Yet, the molecular connections between HU-DNA interactions and changes in nucleoid architecture that regulate gene expression globally remain unknown. Here, we explored the higher-order E. coli nucleoid organization by soft x-ray tomography (SXT) and revealed an effect of HU surface charges in overall nucleoid organization and rearrangements. We also studied global transcription by next-generation RNA sequencing (RNA-Seq) and found a link between nucleoid rearrangement and changes in global transcription. To determine the functional relationships of observed nucleoid rearrangement and HU-DNA interactions, we also characterized the overall organization of HU nucleoprotein complexes in solution by small angle x-ray scattering (SAXS). We found that HUαα-mediated DNA networks are different at different ionic strengths and pHs. By means of macromolecular crystallography, we additionally elucidate HUαα dependent molecular switches that modulate DNA networking. This integrative structural study explains how HUαα regulates dynamic transformations of the nucleoid by DNA bridging to control nucleoid rearrangement and global gene regulation.

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Direct observation of dynamic mechanical regulation of DNA condensation by environmental stimuli.

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Direct observation of dynamic mechanical regulation of DNA condensation by environmental stimuli.

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