Background
Identification of biallelic CNPY3 mutations in patients with epileptic encephalopathy and abnormal electroencephalography findings of Cnpy3 knock-out mice have indicated that the loss of CNPY3 function causes neurological disorders such as epilepsy. However, the basic property of CNPY3 in the brain remains unclear. New method: We generated C-terminal 2xHA-tag knock-in Cnpy3 mice by i-GONAD in vivo genome editing system to investigate the expression and function of Cnpy3 in the mouse brain.
Conclusions
Taken together, we demonstrated that Cnpy32xHA knock-in mice would be useful to further elucidate the property of Cnpy3 in brain function and neurological disorders.
Methods
Conventional Cnpy3 antibodies could not allow us to investigate the distribution of Cnpy3 expression in the brain, while HA-tagging revealed the expression of CNPY3 in neuronal cells. Conclusions: Taken together, we demonstrated that Cnpy32xHA knock-in mice would be useful to further elucidate the property of Cnpy3 in brain function and neurological disorders.
Results
2xHA-tagged Cnpy3 was confirmed by immunoblot analysis using anti-HA and CNPY3 antibodies, although HA tagging caused the decreased Cnpy3 protein level. Immunohistochemical analysis of Cnpy32xHA knock-in mice showed that Cnpy3-2xHA was predominantly expressed in the neuron. In addition, Cnpy3 and Cnpy3-2xHA were both localized in the endoplasmic reticulum and synaptosome and showed age-dependent expression changes in the brain. Comparison with existing methods: Conventional Cnpy3 antibodies could not allow us to investigate the distribution of Cnpy3 expression in the brain, while HA-tagging revealed the expression of CNPY3 in neuronal cells. Conclusions: Taken together, we demonstrated that Cnpy32xHA knock-in mice would be useful to further elucidate the property of Cnpy3 in brain function and neurological disorders.