lacZ sequences prevent regulated expression of housekeeping genes.
In order to dissect the MHC class I H-2K gene regulatory sequences, we p reviously generated transgenic mice containing various H-2K/lacZ fusion genes. However contrary to transgenes where H-2K sequences were fused to other coding sequences, none of the lacZ fusion transgenes was widely ex pressed like H-2K gene. We now show that this silencing also occurs when lacZ is inserted into a larger H-2K genomic construct including promoter and other regulatory elements. Because the 5'H-2K region contains a CpG island, we suspected that the presence of lacZ coding sequences was inte rfering with the mechanism by which the H-2K promoter region is normally unmethylated and transcriptionally active. Indeed, we show that in high ( >10) copy number transgenic mice, insertion of lacZ sequences in the v icinity of the H-2K promoter results in partial or complete methylation of the H-2K CpG island. However, in low (1-3) copy number transgenic mic e no methylation was observed but the transgene was still silent, sugges ting that the silencing effect of lacZ does not only rely on abnormal CpG methylation. Intriguingly, when the H -2/lacZ construct was introduced via embryonic stem (ES) cells, regulate d transgene expression was observed in several chimaeric embryos derived from independent ES clones, but never in adult chimeras. Combined with t he fact that, despite much effort, it has been very difficult to generat e 'blue' mice, our results highlight the transcription-silencing effect of lacZ sequences when they are associated with regulatory sequences of ubiquitously expressed genes.
Cohen-Tannoudji
M
M
Unité de Biologie du Développement, CNRS URA 1960, Institut Pasteur, 25 rue du Dr. Roux, 75724, Paris, France. m-cohen@pasteur.fr
Vandormael-Pournin
S
S
Drezen
J
J
Mercier
P
P
Babinet
C
C
Morello
D
D
D020121:5' Untranslated Regions; D000818:Animals; D019677:Artificial Gene Fusion; D005786:Gene Expression Regulation; D006183:H-2 Antigens / Q000235:genetics*; D007763:Lac Operon; D051379:Mice; D008822:Mice, Transgenic; D011401:Promoter Regions, Genetic; D014162:Transfection
D020121:5' Untranslated Regions; D006183:H-2 Antigens