Intranuclear trafficking of episomal DNA is transcription-dependent.
Our aim is to characterize the poorly understood mechanisms that influence episomal transgene expression within the nucleus. We found that plasmid DNA micro-injected directly into a nucleus moves into a speckled pattern and occupies less nuclear volume than bovine serum albumin (BSA) or other inert molecules after 4 hours. In addition, plasmids that contain eukaryotic regulatory sequences and actively transcribe transgenes condense into a few select areas of the nucleoplasm and occupy less nuclear volume than bacterial vectors. This suggests that episomal DNA moves in a sequence and transcription-dependent manner. We have also found that plasmids traffic to specific subnuclear domains depending on their sequence. Our experiments show that plasmids with polymerase II regulatory elements will target to nuclear spliceosome regions, while plasmids with the polymerase I promoter often traffic into nucleoli. Further elucidation of intranuclear plasmid trafficking behavior may lead to a better understanding of gene expression, and thereby help to improve basic laboratory techniques and clinical gene therapies.
1Division of Pulmonary and Critical Care Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, USA.
Joshua Z
JZ
Gasiorowski
David A
DA
Dean
D001483:Base Sequence; D001692:Biological Transport; D004247:DNA / Q000378:metabolism*; D017931:DNA Primers; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006801:Humans; D017403:In Situ Hybridization; D010957:Plasmids / Q000378:metabolism*; D014158:Transcription, Genetic
D017931:DNA Primers; D004247:DNA