Abstract

Methyl-CpG binding protein 2 (MeCP2) has crucial roles in transcriptional regulation and microRNA processing1,2,3,4. Mutations in the MECP2 gene are found in 90% of patients with Rett syndrome, a severe developmental disorder with autistic phenotypes5. Duplications of MECP2-containing genomic segments cause the MECP2 duplication syndrome, which shares core symptoms with autism spectrum disorders6. Although Mecp2-null mice recapitulate most developmental and behavioural defects seen in patients with Rett syndrome, it has been difficult to identify autism-like behaviours in the mouse model of MeCP2 overexpression7,8. Here we report that lentivirus-based transgenic cynomolgus monkeys (Macaca fascicularis) expressing human MeCP2 in the brain exhibit autism-like behaviours and show germline transmission of the transgene. Expression of the MECP2 transgene was confirmed by western blotting and immunostaining of brain tissues of transgenic monkeys. Genomic integration sites of the transgenes were characterized by a deep-sequencing-based method. As compared to wild-type monkeys, MECP2 transgenic monkeys exhibited a higher frequency of repetitive circular locomotion and increased stress responses, as measured by the threat-related anxiety and defensive test9. The transgenic monkeys showed less interaction with wild-type monkeys within the same group, and also a reduced interaction time when paired with other transgenic monkeys in social interaction tests. The cognitive functions of the transgenic monkeys were largely normal in the Wisconsin general test apparatus, although some showed signs of stereotypic cognitive behaviours. Notably, we succeeded in generating five F1 offspring of MECP2 transgenic monkeys by intracytoplasmic sperm injection with sperm from one F0 transgenic monkey, showing germline transmission and Mendelian segregation of several MECP2 transgenes in the F1 progeny. Moreover, F1 transgenic monkeys also showed reduced social interactions when tested in pairs, as compared to wild-type monkeys of similar age. Together, these results indicate the feasibility and reliability of using genetically engineered non-human primates to study brain disorders.

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Accessions

Primary accessions

Gene Expression Omnibus

Data deposits

The raw sequence and processed data have been submitted to the NCBI Gene Expression Ominbus (GEO) under accession number GSE57974.

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Acknowledgements

We thank M.-m. Poo for comments on the manuscript, Y.-Z. Li, Y.-Z. Lu, F. Liu and X. Zhou for maintaining monkey colony, W. Lu, X.-H. Guo and Y. F. Zhou of Fudan Children’s Hospital for assistance in mass spectrometry and electroencephalogram, D. Chen and B. Zhang of Novel Bioinformatics for RNA-seq data analysis, and C.-H. Li of Shanghai Geneskies Company for analysis of genomic integration sites. This work was supported by CAS Strategic Priority Research Program (XDB02050400), the MoST 973 Program (2011CBA00400), NSFC grants (91432111, 91232712 and 81527901), National Key Technology R&D Program of China 2014BAI03B00, Shanghai City Committee of Science and Technology Project 14140900100.

Author information

Author notes

    • Zhen Liu
    •  & Xiao Li

    These authors contributed equally to this work.

Affiliations

  1. Institute of Neuroscience, CAS Key Laboratory of Primate Neurobiology, State Key Laboratory of Neuroscience, CAS Center for Excellence in Brain Science and Intelligence Technology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 320 Yue-Yang Road, Shanghai 200031, China

    • Zhen Liu
    • , Xiao Li
    • , Jun-Tao Zhang
    • , Yi-Jun Cai
    • , Tian-Lin Cheng
    • , Cheng Cheng
    • , Yan Wang
    • , Chen-Chen Zhang
    • , Yan-Hong Nie
    • , Zhi-Fang Chen
    • , Wen-Jie Bian
    • , Bin Lu
    • , Yue-Fang Zhang
    • , Xiao-Di Zhang
    • , Xiao Sang
    • , Jia-Jia Wu
    • , Zhi-Qi Xiong
    • , Xiang Yu
    • , Neng Gong
    • , Qiang Sun
    •  & Zilong Qiu
  2. State Key Laboratory of Genetic Engineering and Ministry of Education Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center of Genetics and Development, School of Life Sciences, Fudan University, Shanghai 200438, China

    • Ling Zhang
    • , Jianqiu Xiao
    •  & Feng Zhang
  3. Department of Child Healthcare, Children’s Hospital of Fudan University, Shanghai 201102, China

    • Xiu Xu
  4. Department of Neonatology, Children’s Hospital of Fudan University, Shanghai 201102, China

    • Wen-Hao Zhou

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Contributions

Z.Q. and Q.S. conceived and supervised the project. T.-L.C. constructed the lentiviral constructs. Q.S. and Z.L. performed the cynomolgus oocytes preparation and injection. Y.-J.C., Y.W., C.-C.Z., Y.-H.N. and Z.L. contributed to monkey reproductive experiments. Y.-F.Z. performed PCR-based genotyping experiments. Z.-F.C., W.-J.B., X.-D.Z. and X.Y. performed immunohistochemistry and AccuCopy experiments. C.C., B.L., X.S. and Z.-Q.X. performed western blot experiments. X.L. and J.-J.W. performed behavioural analysis. J.-T.Z. and N.G. performed WGTA tests. W.-H.Z. and X.X. contributed to metabolic measurements and behavioural analysis. T.-L.C. and X.L. performed genomic integration sites analysis based on deep-sequencing. J.X., L.Z. and F.Z. helped with identification of genomic integration sites of transgenes. Z.Q. wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Qiang Sun or Zilong Qiu.

Extended data

Supplementary information

PDF files

  1. 1.

    Supplementary Figures

    This file contains the Western blots raw data for Figures 1d, e, f, h and 4c.

  2. 2.

    Supplementary Tables

    This file contains Supplementary Tables 1-7

Zip files

  1. 1.

    Supplementary Information

    This zipped file contains Supplementary Audio files 1-3 comprising: (1) Grunt vocal responses of monkeys examined in TAD tests; (2) Coo vocal responses of monkeys examined in TAD tests; (3) Stream vocal responses of monkeys examined in TAD tests.

Videos

  1. 1.

    Video 1:Example of normal motor behaviours

    This video shows the representative normal motor behaviours for monkeys we examined.

  2. 2.

    Video 2: Example of repetitive motor behaviours

    This video shows the typical repetitive motor behaviours for four F0 TG monkeys out of eight F0 TG monkeys we examined.

  3. 3.

    Video 3: Example of social interaction behaviours.

    This video shows the representative social interaction behaviours of monkeys, such as sitting together, in natural community.

  4. 4.

    Video 4: Social Interaction behaviours in pairing experiments

    This video shows the typical social behaviour, sitting together, in pairing experiments.

  5. 5.

    Video 5: Examples of learning behaviors of monkey in learning set of WGTA tests.

    This video shows the example of learned behaviours for W09 out of five WT monkeys we examined.

  6. 6.

    Video 6. Examples of stereotypic behaviours of MECP2 F0 TG monkey in learning set of WGTA tests

    This video shows the example of stereotypic responses of F0 TG monkeys (T09) out of seven F0 TG monkeys we examined.

About this article

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DOI

https://doi.org/10.1038/nature16533

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