Nuclear scaffold/matrix attached region modules linked to a transcription unit are sufficient for replication and maintenance of a mammalian episome.
The activation of mammalian origins of replication depends so far on ill understood epigenetic events, such as binding of transcription factors, chromatin structure, and nuclear localization. Understanding these mechanisms is not only a scientific challenge but also represents a prerequisite for the rational design of nonviral episomal vectors for mammalian cells. In this paper, we demonstrate that a tetramer of a 155-bp minimal nuclear scaffold/matrix attached region DNA module linked to an upstream transcription unit is sufficient for replication and mitotic stability of a mammalian episome in the absence of selection. Fluorescence in situ hybridization analyses, crosslinking with cis-diammineplatinum(II)-dichloride and chromatin immunoprecipitation demonstrate that this vector associates with the nuclear matrix or scaffold in vivo by means of specific interaction of the nuclear scaffold/matrix attached region with the nuclear matrix protein SAF-A. Results presented in this paper define the minimal requirements of an episomal vector for mammalian cells on the molecular level.
Institute of Cell Biology, Witten/Herdecke University, 58448 Witten, Germany.
Andreas C W
AC
Jenke
Isa M
IM
Stehle
Frank
F
Herrmann
Tobias
T
Eisenberger
Armin
A
Baiker
Jürgen
J
Bode
Frank O
FO
Fackelmayer
Hans J
HJ
Lipps
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D004261:DNA Replication / Q000502:physiology*; D004926:Escherichia coli; D008322:Mammals; D008938:Mitosis / Q000502:physiology; D015530:Nuclear Matrix / Q000235:genetics*; D034521:Nuclear Matrix-Associated Proteins / Q000502:physiology; D010957:Plasmids / Q000235:genetics; D014158:Transcription, Genetic / Q000502:physiology*; D019076:Transgenes / Q000235:genetics
D034521:Nuclear Matrix-Associated Proteins