Comparative methylation analysis of murine transgenes that undergo or escape X-chromosome inactivation

We analyzed an X-linked metallothionein–vasopressin (MTVP) fusion transgene that undergoes X-chromosome inactivation (X inactiv ation) and an X-linked transferrin (TFN) transgene that escapes X inactivation with respect to methylation in the 5′ regulatory regions. The MTVP transgene promoter region is unmethylated when the transgene is on the active X chromosome and methylated when on the inactive X chromosome. Interestingly, the MTVP transgene is not detectably transcribed from the male X chromosome, although it is unmethylated, consistent with its availability for transcription. The TFN transgene promoter region is hypomethylated on both the active and inactive X chromosomes, consistent with its expression from both chromosomes. The TFN and MTVP transgenes have been mapped to chromosomal regions D and C, respectively, by fluorescence in situ hybridization. These observations are discussed in the context of our understanding of the role of DNA methylation in the spread and maintenance of X-chromosome inactivation.