lacZ sequences prevent regulated expression of housekeeping genes

In order to dissect the MHC class I H-2K gene regulatory sequences, we previously 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 like H-2K gene. We now show that this silencing also occurs when lacZ is inserted into a larger H-2K genomic construct including promotor and other regulatory elements. Because the 5' H-2K region contains a CpG island, we suspected that the presence of lacZ coding sequences was interfering 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 vicinity 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 mice no methylation was observed but 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 from independent ES clones, but never in adult chimaeras. Combined with the fact that, despite much effort, it has been very difficult to generate 'blue' mice, our results highlight the transcription-silencing effect of lacZ sequences when they are associated with regulatory sequences of ubiquitously expressed genes. (C) 2000 Elsevier Science Ireland Ltd. All rights reserved.