title: Bacterial delivery of TALEN proteins for human genome editing.
abstract: Transcription Activator-Like Effector Nucleases (TALENs) are a novel class of sequence-specific nucleases that have recently gained prominence for its ease of production and high efficiency in genome editing. A TALEN pair recognizes specific DNA sequences and introduce double-strand break in the target site, triggering non-homologous end joining and homologous recombination. Current methods of TALEN delivery involves introduction of foreign genetic materials, such as plasmid DNA or mRNA, through transfection. Here, we show an alternative way of TALEN delivery, bacterial type III secretion system (T3SS) mediated direct injection of the TALEN proteins into human cells. Bacterially injected TALEN was shown to efficiently target host cell nucleus where it persists for almost 12 hours. Using a pair of TALENs targeting venus gene, such injected nuclear TALENs were shown functional in introducing DNA mutation in the target site. Interestingly, S-phase cells seem to show greater sensitivity to the TALEN mediated target gene modification. Accordingly, efficiency of such genome editing can easily be manipulated by the infection dose, number of repeated infections as well as enrichment of S phase cells. This work further extends the utility of T3SS in the delivery of functional proteins into mammalian cells to alter their characters for biomedical applications.
authors.affiliation: Key Laboratory of Ministry of Education, Department of Microbiology, Nankai University, Tianjin, China.
authors.forename: Jingyue
authors.initials: J
authors.lastname: Jia
authors.affiliation: Key Laboratory of Ministry of Education, Department of Microbiology, Nankai University, Tianjin, China.
authors.forename: Yongxin
authors.initials: Y
authors.lastname: Jin
authors.affiliation: Key Laboratory of Ministry of Education, Department of Microbiology, Nankai University, Tianjin, China.
authors.forename: Ting
authors.initials: T
authors.lastname: Bian
authors.affiliation: The Key Laboratory of Regenerative Biology and The Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, Guangzhou Institute of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, China.
authors.forename: Donghai
authors.initials: D
authors.lastname: Wu
authors.affiliation: Department of Pathology, University of Florida, Gainesville, Florida, United States of America.
authors.forename: Lijun
authors.initials: L
authors.lastname: Yang
authors.affiliation: Department of Pathology, University of Florida, Gainesville, Florida, United States of America.
authors.forename: Naohiro
authors.initials: N
authors.lastname: Terada
authors.affiliation: Key Laboratory of Ministry of Education, Department of Microbiology, Nankai University, Tianjin, China.
authors.forename: Weihui
authors.initials: W
authors.lastname: Wu
authors.affiliation: Key Laboratory of Ministry of Education, Department of Microbiology, Nankai University, Tianjin, China; Department of Molecular Genetics and Microbiology, University of Florida, Gainesville, Florida, United States of America.
authors.forename: Shouguang
authors.initials: S
authors.lastname: Jin
mesh_terms: D018407:Bacterial Physiological Phenomena; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D058947:Bacterial Secretion Systems; D001483:Base Sequence; D002453:Cell Cycle / Q000235:genetics; D004720:Endonucleases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D018390:Gene Targeting; D015894:Genome, Human; D006367:HeLa Cells; D059765:Homologous Recombination; D006801:Humans; D008164:Luminescent Proteins / Q000378:metabolism; D011993:Recombinant Fusion Proteins; D014162:Transfection
chemical_list: D001426:Bacterial Proteins; D058947:Bacterial Secretion Systems; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; C054562:yellow fluorescent protein, Bacteria; D004720:Endonucleases