title: Single point mutation in adeno-associated viral vectors -DJ capsid leads to improvement for gene delivery in vivo.
abstract: BACKGROUND
  Rational design of AAV capsids is a simple method for enhancing AAV transduction efficiency. AAV-DJ is a highly recombinogenic hybrid vector created from DNA shuffling of eight AAV serotypes, which mediates efficient gene expression both in vitro and in vivo. AAV2 and AAV8 are the closest parental vectors of AAV-DJ and it has been reported that mutations on the 137/251/503 ubiquitination or phosphorylation sites of the AAV2 or AAV8 capsid lead to dramatic enhancement of gene delivery. Here, we aimed to find out whether the same point mutations on the AAV-DJ capsid could lead to significant improvement for gene delivery both in vitro and in vivo.
  
  
  RESULTS
  We constructed three single point mutants (K137R/T251A/S503A) of AAV-DJ and the transduction efficiency of these mutants and AAV-DJ were investigated using two reporter gene systems including green fluorescent protein (GFP) and dual-luciferase (Gaussia luciferase and Firefly luciferase). Data indicated that single point mutations T251A/S503A lead to significant improvement of dual-luciferase expression in vivo after tail vein (TV) injection in mice respectively, despite limited enhancement of GFP expression in 293 T, Hela and HepG2 cells in vitro. Moreover, in vivo bioluminescence image and viral genome DNA copy number in tissue analysis showed that these mutants reserved the liver tropism characteristics, consistent with AAV-DJ.
  
  
  CONCLUSIONS
  Single point mutations on the 251/503 sites of AAV-DJ capsid can lead to a significant improvement for in vivo gene expression. These enhanced AAV vectors have great potential in gene therapy applications.
authors.affiliation: School of Biotechnology, Southern Medical University, 1023 South Shatai Road, Guangzhou, Guangdong, 510515, China.
authors.forename: Yingying
authors.initials: Y
authors.lastname: Mao
authors.affiliation: Department of Biotechnology, Beijing Institute of Radiation Medicine, Beijing, China. xjwang@bmi.ac.cn.
authors.forename: Xuejun
authors.initials: X
authors.lastname: Wang
authors.affiliation: School of Biotechnology, Southern Medical University, 1023 South Shatai Road, Guangzhou, Guangdong, 510515, China.
authors.forename: Renhe
authors.initials: R
authors.lastname: Yan
authors.affiliation: Department of Biotechnology, Beijing Institute of Radiation Medicine, Beijing, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Hu
authors.affiliation: Department of Biomedical Engineering, The Johns Hopkins University School of Medicine, Baltimore, USA.
authors.forename: Andrew
authors.initials: A
authors.lastname: Li
authors.affiliation: Department of Biotechnology, Beijing Institute of Radiation Medicine, Beijing, China. sqwang@bmi.ac.cn.
authors.forename: Shengqi
authors.initials: S
authors.lastname: Wang
authors.affiliation: School of Biotechnology, Southern Medical University, 1023 South Shatai Road, Guangzhou, Guangdong, 510515, China. hongwei1@yahoo.com.
authors.forename: Hongwei
authors.initials: H
authors.lastname: Li
mesh_terms: D000818:Animals; D036022:Capsid Proteins / Q000235:genetics*; D002460:Cell Line; D000229:Dependovirus / Q000235:genetics*; D016679:Genome, Viral / Q000235:genetics; D006367:HeLa Cells; D056945:Hep G2 Cells; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D061848:Optical Imaging; D017354:Point Mutation / Q000235:genetics*; D014018:Tissue Distribution; D014162:Transfection / Q000379:methods*
chemical_list: D036022:Capsid Proteins; D008156:Luciferases