When the chromosomal locus of a gene is changed by transposition, inversion, or experimental transgenics position-dependent effects on gene expression are often observed. Position-effects are mediated by enhancers and silencers which normally regulate other genes, or by repressive heterochromatin. Flanking DNA sequence elements from the human keratin- 18 gene can insulate heterologous genes from position-effects in transgenic mice. In this proposal, an 825 nt keratin-18 sequence element will be tested for its' ability to block position-effects on a metallothionein reporter construct in transgenic mice. The 825 nt fragment will be tested for its ability to block activation of an HSV-1 thymidine kinase promoter by the SV4O enhancer in stablely transfected mammalian cells. Finally, it will be determined whether the 825 nt fragment will block chromatin-mediated repression of the URA3 gene at the left arm of S. cerevisae chromosome VII. This work is directed toward understanding how active genes avoid heterochromatin-mediated repression and how adjacent genes on the chromosome can be regulated independently, despite their close proximity. Such studies are critical for the development of safe and effective human gene therapeutic techniques.
| Willoughby, D A; Vilalta, A; Oshima, R G (2000) An Alu element from the K18 gene confers position-independent expression in transgenic mice. J Biol Chem 275:759-68 |
