Earticle

초록

영어

G protein-coupled receptor kinase 5 (GRK5) is one of the seven GRK family members whose primary function is to desensitize G protein-coupled receptors (GPCRs). In recently, GRK5 deficiency has been linked to early Alzheimer disease (AD), but the mechanism by which GRK5 deficiency may accelerate to AD pathogenesis remains elusive. The GRK5 mRNA is expressed widely in brain and peripheral tissues, with highest expression evident heart, lung, and placenta. In cellular model systems, GRK5 can phosphorylate several neuronal GPCRs including ß2-adrenergic, M2-muscarinic, secretin, angiotensin AT1, and thyroid stimulating hormone receptors. Transcription activator-like effector nucleases (TALENs) are programmable nucleases that join FokI endonucleases with the modular DNA-binding domain of TALEs and highly effective in inducing mutations at specific genome loci. TALEN-mediated mutagenesis in zygotes is a potential alternative to conventional gene targeting in mice. In the presented study, we report the generation of mice with genetic knockout of the GRK gene using TALENs. We designed TALEN vectors for exon 1, 3 and 5 of mouse GRK5 gene and tested their ability to alter the each surrogate vector in 293T cells. We prepared of mRNAs for the linearized TALEN using the mMessage mMachine T7 Ultra kit. mRNAs (4ng/μl) was injected into cytoplasm of 180 one-cell embryos. After incubation for 24 hours, the selected two-cell embryos transferred into the oviduct of seven pseudopregnant C57BL/6 mice. We confirmed the genotype of Fo mice by sequencing and T7E1 assay. We found 6 mutant mice lines (11%) from 53 newborns. We also mated 3 Fo GRK5 mutant lines with wild type mice and confirmed the genotype of the F1 progenies. All the mutations observed in Fo mice were transmitted through the germline but not all progenies (8/3, 13/4, 7/4). Taken together, TALEN-mediated mutagenesis might accelerate the creation of genetically engineered mouse models and elucidate the mechanism of AD pathogenesis using GRK5 knock out mice.

키워드

저자

• Tsevelmaa Nanjidsuren [ 1Department of Animal Biotechnology Graduate School of Future Fusion Technology, Hankyong National University ]
• Bo-Woong Sim [ National Primate Research Center ]
• Min-Su Kim [ Department of Animal Biotechnology, Graduate School of Future Fusion Technology, Hankyong National University ]
• Chae-Won Park [ Department of Animal Biotechnology, Graduate School of Future Fusion Technology, Hankyong National University ]
• Eun-Bi Seo [ Department of Animal Biotechnology, Graduate School of Future Fusion Technology, Hankyong National University ]
• Sun-Ok Kim [ Department of Animal Biotechnology, Graduate School of Future Fusion Technology, Hankyong National University ]
• Kyu-Tae Chang [ National Primate Research Center ]
• Kwan-Sik Min [ Department of Animal Biotechnology, Graduate School of Future Fusion Technology, Hankyong National University ]

참고문헌

발행기관

간행물

표지보기 관심저널 등록

• 간행물명

  Reproductive & Developmental Biology(Supplement)

• 간기

  연간

• 수록기간

  2001~2017

• 십진분류

  KDC 527 DDC 636