Simultaneous expression of different transgenes in neurons and glia by combining in utero electroporation with the Tol2 transposon-mediated gene transfer system.
In utero electroporation is widely used to study neuronal development and function by introducing plasmid DNA into neural progenitors during embryogenesis. This is an effective and convenient method of introducing plasmid DNA into neural precursors and is suitable for manipulating gene expression in cells of the CNS. However, the applicability of this technique is comparatively limited to neuronal research, as the plasmid DNA introduced into neural progenitors during embryogenesis is diluted by cell proliferation and is not stably maintained in glial cells generated around and after birth. To overcome this limitation, we applied the Tol2 transposon system, which integrates a transgene into the genome of the host cell, to in utero electroporation. With this system, we confirmed that the transgene was effectively maintained in the progeny of embryonic neural precursors, astrocytes and oligodendrocytes. Using the glial promoters GFAP and S100beta, targeted and stable expressions of transgenes in glia were obtained, which enabled the expression of different transgenes simultaneously in neurons and glia. Glia-targeted expression of the transgene that causes neuronal migration defect was achieved without the defect. Thus, use of the Tol2 transposon system in combination with in utero electroporation is a powerful method for studying glia-neuron interactions in vivo.
Yoshida
Ayako
A
Department of Genetics, Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Yamaguchi
Yoshifumi
Y
Nonomura
Keiko
K
Kawakami
Koichi
K
Takahashi
Yoshiko
Y
Miura
Masayuki
M
D000818:Animals; D004251:DNA Transposable Elements; D018274:Electroporation / Q000379:methods*; D005260:Female; D018507:Gene Expression Regulation, Developmental; D018014:Gene Transfer Techniques; D005904:Glial Fibrillary Acidic Protein / Q000235:genetics / Q000378:metabolism; D051379:Mice; D009414:Nerve Growth Factors / Q000235:genetics / Q000378:metabolism; D009457:Neuroglia / Q000166:cytology / Q000502:physiology*; D009474:Neurons / Q000166:cytology / Q000502:physiology*; D011247:Pregnancy; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D064568:S100 Calcium Binding Protein beta Subunit; D009418:S100 Proteins / Q000235:genetics / Q000378:metabolism; D013234:Stem Cells / Q000166:cytology / Q000502:physiology; D019076:Transgenes; D019895:Transposases / Q000378:metabolism
D004251:DNA Transposable Elements; D005904:Glial Fibrillary Acidic Protein; D009414:Nerve Growth Factors; D011993:Recombinant Fusion Proteins; D064568:S100 Calcium Binding Protein beta Subunit; D009418:S100 Proteins; D019895:Transposases
Nature. 2001 Feb 8;409(6821):714-20
11217860
EMBO J. 2003 Aug 15;22(16):4190-201
12912917
Dev Biol. 2004 Jun 1;270(1):200-13
15136150
Nature. 2004 Apr 15;428(6984):767-71
15085137
Science. 1988 Sep 9;241(4871):1342-5
3137660
Neuron. 1993 Feb;10(2):201-12
8439409
Eur J Neurosci. 2006 Oct;24(7):1897-906
17067293
J Neurochem. 1990 Oct;55(4):1180-8
2398353
Dev Dyn. 2007 May;236(5):1273-86
17377980
Dev Biol. 2008 Aug 15;320(2):456-68
18582453
Neuroscience. 2001;103(4):865-72
11301197
Dev Growth Differ. 1999 Feb;41(1):59-72
10445503
Dev Biol. 2007 May 15;305(2):625-36
17359965
Genesis. 2006 Sep;44(9):438-45
16906529
Glia. 2009 Aug 1;57(10):1115-29
19115384
J Biol Chem. 2005 Feb 11;280(6):5082-8
15557338
Proc Natl Acad Sci U S A. 2000 Oct 10;97(21):11403-8
11027340
Neurochem Int. 2004 Jul-Aug;45(2-3):191-202
15145537
J Neurosci. 2006 Jan 18;26(3):1045-56
16421324
Dev Biol. 2001 Dec 1;240(1):237-46
11784059
Cereb Cortex. 2009 Jul;19 Suppl 1:i120-5
19395528
Trends Neurosci. 2009 Jun;32(6):303-11
19398132
Cell. 2008 Nov 14;135(4):738-48
19013281
J Comp Neurol. 2003 Mar 17;457(4):404-19
12561079
Development. 2006 Oct;133(20):4085-96
16987869
Cell. 2008 Nov 14;135(4):749-62
19013282
Prog Neurobiol. 2008 Dec 11;86(4):342-67
18948166
Neuron. 2007 May 3;54(3):357-69
17481390
Dev Biol. 2007 May 15;305(2):616-24
17362912
Proc Natl Acad Sci U S A. 2008 Mar 4;105(9):3581-6
18299565
Science. 2001 Nov 2;294(5544):1071-4
11567107
J Neurosci. 1999 Dec 1;19(23):10357-71
10575033
J Neurosci Res. 2007 Dec;85(16):3567-73
17671991
Genetics. 2004 Feb;166(2):895-9
15020474
Biochem Biophys Res Commun. 1997 Jan 13;230(2):376-80
9016787
Gene. 2008 Dec 1;425(1-2):64-8
18775483