A novel piggyBac transposon inducible expression system identifies a role for AKT signalling in primordial germ cell migration.
In this work, we describe a single piggyBac transposon system containing both a tet-activator and a doxycycline-inducible expression cassette. We demonstrate that a gene product can be conditionally expressed from the integrated transposon and a second gene can be simultaneously targeted by a short hairpin RNA contained within the transposon, both in vivo and in mammalian and avian cell lines. We applied this system to stably modify chicken primordial germ cell (PGC) lines in vitro and induce a reporter gene at specific developmental stages after injection of the transposon-modified germ cells into chicken embryos. We used this vector to express a constitutively-active AKT molecule during PGC migration to the forming gonad. We found that PGC migration was retarded and cells could not colonise the forming gonad. Correct levels of AKT activation are thus essential for germ cell migration during early embryonic development.
The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Easter Bush Campus, Midlothian, United Kingdom.
James D
JD
Glover
Lorna
L
Taylor
Adrian
A
Sherman
Caroline
C
Zeiger-Poli
Helen M
HM
Sang
Michael J
MJ
McGrew
D053687:Adult Stem Cells / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D002465:Cell Movement; D002642:Chick Embryo; D002645:Chickens; D003001:Cloning, Molecular; D004251:DNA Transposable Elements; D053595:Embryonic Stem Cells / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D015870:Gene Expression; D005818:Genetic Engineering; D005854:Germ Cells / Q000502:physiology* / Q000637:transplantation; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D012661:Semen / Q000166:cytology; D015398:Signal Transduction; D015533:Transcriptional Activation
D004251:DNA Transposable Elements; D049452:Green Fluorescent Proteins; D051057:Proto-Oncogene Proteins c-akt