Hum Mol Genet. 2006 Apr 13; [Epub ahead of print]
Identification of cis-regulatory elements for MECP2 expression
Liu J, Francke U.
Departments of Genetics, Stanford University School of Medicine, Stanford, CA, USA.
Abstract
Rett syndrome (RTT) is an X-linked dominant disabling neurodevelopmental disorder caused by loss of function mutations in the MECP2 gene, located at Xq28, which encodes a multifunctional protein. MECP2 expression is regulated in a developmental stage and cell-type specific manner. The need for tightly controlled MeCP2 levels in brain is strongly suggested by neurologically abnormal phenotypes of mouse models with mild overexpression, and by mental retardation in human males with MECP2 duplication. We set out to identify long-range cis-regulatory sequences that differentially regulate MECP2 transcription, and when mutated, may contribute to the pathogenesis of RTT, autism or X-linked mental retardation. By inter-species sequence comparisons we detected 27 highly conserved non-coding DNA sequences within a 210 kb region covering MECP2. We functionally confirmed four enhancer and two silencer elements by performing luciferase reporter assays in four different human cell lines. The transcription factor binding capability of the identified regulatory elements was tested by gel shift assays. To locate the human MECP2 core promoter, we dissected the promoter region by reporter assays with deletion constructs. We then used chromosome conformation capture methods to document long-range interactions of three enhancers and two silencers with the MECP2 promoter. Acting over distances of up to 130 kb, these elements may influence chromatin configurations and regulate MECP2 transcription. Our study has defined the "MECP2 functional expression module" and identified enhancer and silencer elements that are likely to be responsible for the tissue-specific, developmental stage-specific or splice-variant specific control of MeCP2 protein expression.
Lay Summary
Genes are not simply "on" (expressed) or "off" (not expressed) but rather can be "on" or "off" in varying degrees. The level of expression is a highly regulated process. For example, we know that there is a need for tight control of MeCP2 levels in the brain: too little leads to Rett Syndrome but too much also leads to neurological dysfunction. In this paper, the authors set out to identify those sections of DNA surrounding the MECP2 gene that either enhance or silence the expression of the gene. These are known as "cis-regulatory transcriptional elements," and regulate gene expression at the right time, in the right cells, and at the correct level. The authors use various experimental techniques to identify a host of those elements involved in regulation of MECP2 expression. These are very important findings, as they represent a key step in understanding the mechanisms of how, where, and when the MECP2 gene is expressed. Indeed, this needs to be fully investigated before gene therapy-type studies could be designed.