Discovery, characterization, and application of chromosomal integration sites in the hyperthermophilic archaeon Sulfolobus islandicus
University of Illinois at Urbana-Champaign Institute for Sustainability, Energy, and Environment
Sulfolobus islandicus, an archaeal model organism, offers unique advantages for metabolic engineering and synthetic biology applications owing to its ability to thrive under low pH and high temperature conditions. Although several genetic tools exist for this organism, the absence of well-defined chromosomal integration sites continues to limit its development as a cellular factory.
A research team at the University of Illinois Urbana-Champaign employed the CRISPR-COPIES pipeline and a multi-omics strategy that integrates genomics and epigenomics to guide the selection of genomic regions suitable for integration. Researchers then characterized these candidate sites in vivo using the endogenous Type I-A CRISPR-Cas system and a lacS reporter system.
The team identified 66 crRNAs targeting 21 intergenic regions with potential for gene integration, and functionally validated 13 crRNAs corresponding to eight sites using a β-galactosidase activity assay.
Overexpression of GrsB (glycerol dibiphytanyl glycerol tetraether [GDGT] ring synthase B) at one of these sites modulated lipid ether composition, increasing the number of cyclopentane moieties.
This work expands the genetic toolbox for non-conventional hosts, advancing the potential for robust platforms for synthetic biology and industrial biotechnology.
The study was supported by the Center for Advanced Bioenergy and Bioproducts Innovation (CABBI), a U.S. Department of Energy-funded Bioenergy Research Center. Other funding was provided by the National Science Foundation, the National Research Foundation of Korea , and the Gordon and Betty Moore Foundation.
Images available upon request.
Disclaimer: AAAS and EurekAlert! are not responsible for the accuracy of news releases posted to EurekAlert! by contributing institutions or for the use of any information through the EurekAlert system.