title: Engineering Translational Activators with CRISPR-Cas System.
abstract: RNA parts often serve as critical components in genetic engineering. Here we report a design of translational activators which is composed of an RNA endoribonuclease (Csy4) and two exchangeable RNA modules. Csy4, a member of Cas endoribonuclease, cleaves at a specific recognition site; this cleavage releases a cis-repressive RNA module (crRNA) from the masked ribosome binding site (RBS), which subsequently allows the downstream translation initiation. Unlike small RNA as a translational activator, the endoribonuclease-based activator is able to efficiently unfold the perfect RBS-crRNA pairing. As an exchangeable module, the crRNA-RBS duplex was forwardly and reversely engineered to modulate the dynamic range of translational activity. We further showed that Csy4 and its recognition site, together as a module, can also be replaced by orthogonal endoribonuclease-recognition site homologues. These modularly structured, high-performance translational activators would endow the programming of gene expression in the translation level with higher feasibility.
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences ; Beijing, 100101, China.
authors.forename: Pei
authors.initials: P
authors.lastname: Du
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences ; Beijing, 100101, China.|College of Life Science, University of Science and Technology of China , Hefei 230027, China.
authors.forename: Chensi
authors.initials: C
authors.lastname: Miao
authors.affiliation: College of Life Sciences, Fujian Agriculture and Forestry University , Fuzhou 350002, China.
authors.forename: Qiuli
authors.initials: Q
authors.lastname: Lou
authors.affiliation: Department of Pharmacology, UNC-CH School of Medicine , Chapel Hill, North Carolina 27599, United States.|CAS-MPG Partner Instituter for Computational Biology, Shanghai 200031, China.
authors.forename: Zefeng
authors.initials: Z
authors.lastname: Wang
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences ; Beijing, 100101, China.
authors.forename: Chunbo
authors.initials: C
authors.lastname: Lou
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D064113:CRISPR-Cas Systems / Q000235:genetics*; D004722:Endoribonucleases / Q000378:metabolism; D017930:Genes, Reporter; D005818:Genetic Engineering / Q000379:methods*; D008969:Molecular Sequence Data; D012313:RNA / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism*
chemical_list: D015534:Trans-Activators; D012313:RNA; D004722:Endoribonucleases
keywords: CRISPR endoribonuclease; Csy4; RBS calculator; modularization; orthogonal parts; translational activator