Glial cell type-specific gene expression in the mouse cerebrum using the piggyBac system and in utero electroporation

Toshihide Hamabe-Horiike,Keiko Ogawa-Ochiai,Masataka Sakashita,Kanji Kawasaki,Hiroshi Kawasaki,Tomokazu Yoshizaki,Yohei Shinmyo,Chihiro Ishizu,Shin-ichi Harada

doi:10.1038/s41598-021-84210-z

Toshihide Hamabe-Horiike, Keiko Ogawa-Ochiai + Show 7 more

Open Access

https://doi.org/10.1038/s41598-021-84210-z

Copy DOI

Abstract

Glial cells such as astrocytes and oligodendrocytes play crucial roles in the central nervous system. To investigate the molecular mechanisms underlying the development and the biological functions of glial cells, simple and rapid techniques for glial cell-specific genetic manipulation in the mouse cerebrum would be valuable. Here we uncovered that the Gfa2 promoter is suitable for selective gene expression in astrocytes when used with the piggyBac system and in utero electroporation. In contrast, the Blbp promoter, which has been used to induce astrocyte-specific gene expression in transgenic mice, did not result in astrocyte-specific gene expression. We also identified the Plp1 and Mbp promoters could be used with the piggyBac system and in utero electroporation to induce selective gene expression in oligodendrocytes. Furthermore, using our technique, neuron-astrocyte or neuron-oligodendrocyte interactions can be visualized by labeling neurons, astrocytes and oligodendrocytes differentially. Our study provides a fundamental basis for specific transgene expression in astrocytes and/or oligodendrocytes in the mouse cerebrum.

Highlights

Glial cells such as astrocytes and oligodendrocytes play crucial roles in the central nervous system
In utero electroporation using the piggyBac system led to transgene expression in both neurons and glial cells in the mouse cerebrum. (A) Schematics of pPB-CAG-EGFP, pCAG-PBase and pCAG-mCherry plasmids. pPB-CAG-EGFP drives EGFP expression under the CAG promoter. pCAG-PBase has a piggyBac transposase, which recognizes the terminal region (TR) in piggyBac donor plasmids. (B) Experimental procedure. piggyBac plasmids plus pCAG-mCherry were co-transfected into the mouse cerebrum using in utero electroporation at E15.5, and coronal sections were prepared at postnatal day 30 (P30). (C, D) Coronal sections of the electroporated cerebrum
We identified a glial fibrillary acidic protein (GFAP) promoter which led to astrocyte-specific transgene expression, and a Plp[1] promoter and an myelin basic protein (Mbp) promoter which resulted in oligodendrocyte-specific transgene expression

Summary

Introduction

Our study provides a fundamental basis for specific transgene expression in astrocytes and/or oligodendrocytes in the mouse cerebrum. Glial cells such as astrocytes and oligodendrocytes play crucial roles in the central nervous system. In utero electroporation using the piggyBac system led to transgene expression in both neurons and glial cells in the mouse cerebrum. Here we searched for promoters which are specific to astrocytes or oligodendrocyte in the mouse cerebral cortex when used with the piggyBac system and in utero electroporation. The Blbp promoter, which has been used to induce astrocyte-specific gene expression in transgenic mice, did not result in astrocyte-specific gene expression when used with the piggyBac system and in utero electroporation. Our findings provide a fundamental basis for specific transgene expression in astrocytes and/or oligodendrocytes in the mouse cerebral cortex

Methods

Results

Conclusion