lacZ sequences prevent regulated expression of housekeeping genes

In order to dissect the MHC class I H-2K gene regulatory sequences, we prev iously 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 expressed l ike H-2K gene. We now show that this silencing also occurs when lacZ is ins erted into a larger H-2K genomic construct including promotor and other reg ulatory elements. Because the 5' H-2K region contains a CpG island, we susp ected that the presence of lacZ coding sequences was interfering with the m echanism by which the H-2K promoter region is normally unmethylated and tra nscriptionally active. Indeed, we show that in high ( > 10) copy number tra nsgenic mice, insertion of lacZ sequences in the vicinity of the H-2K promo ter results in partial or complete methylation of the H-2K CpG island. Howe ver, in low (1-3) copy number transgenic mice no methylation was observed b ut the transgene was still silent, suggesting 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, regulated transgene expression was observed in several chimaeric embryos derived fro m independent ES clones, but never in adult chimaeras. Combined with the fa ct that, despite much effort, it has been very difficult to generate 'blue' mice, our results highlight the transcription-silencing effect of lacZ seq uences when they are associated with regulatory sequences of ubiquitously e xpressed genes. (C) 2000 Elsevier Science Ireland Ltd. All rights reserved.