Research Awards
Aharon Razin, Ph.D.
The Hebrew University of Jerusalem-Hadassah Medical School, Israel
"MeCP2 and Rett Syndrome"
1-Year Award: $50,000
Research Sponsor: Rett Syndrome Association of Illinois / Adam (Buddha) Lavey
Final Report (November 2003)
In 80% of Rett syndrome patients a mutation in the MeCP2 gene had been identified, leaving around 20% of Rett syndrome cases undiagnosed genetically. It is possible that these cases are inflicted with a mutation outside the gene, or that they carry a deletion of the entire gene. However, it cannot be ruled out that in some Rett Syndrome cases the MeCP2 gene is intact, a normal protein is produced, but one or more of its target genes are impaired. Our first attempt to examine these possibilities was to search for possible mutations in the promoter of the gene. To this end, we have analyzed samples from over 100 Rett syndrome patients from many countries around the world. We failed so far to identify any mutations in promoter sequences in these samples. However, we have identified in two Israeli Rett syndrome patients a bona fide mutation within another non coding sequence. This mutation (a deletion of one adenine residue) was found at a position which is believed to be essential for processing the gene product. We will study this mutation and try to verify its relevance to Rett Syndrome. We have already detected no MeCP2 protein in cells from this patient. If we will be convinced that this mutation is relevant to the Rett Syndrome, we will screen for this mutation in all the samples of the Rett syndrome patients that we analyzed for promoter mutations.
In parallel, we have generated a lymphoblast cell line from another Rett patient with no detectable mutation in the MeCP2 coding region. These cells have an intact MeCP2 protein and this may suggest that genes other than MeCP2 are involved in the disease. Such genes may be targets of MeCP2 activity or, alternatively, genes that code proteins that interact with MeCP2. A mutation in one of these genes could theoretically result in the Rett syndrome phenotype.
To identify MeCP2 target genes we have generated two immortalized lymphoblast cell lines originating from a Rett syndrome patient that carries a mutation in the coding sequence of the MeCP2 gene. One cell line expresses the MeCP2 protein (M10) and the other (M12), is MeCP2 deficient. We performed microarray analysis to compare the expression profile of 30,000 genes in these MeCP2 deficient and normal cell lines. This analysis revealed 90 genes with elevated expression levels in M12 compared to M10. Twelve genes with the largest change in expression level were chosen for further analysis.
The next step will be to try to restore the normal expression levels of these genes by introducing the normal MeCP2 gene into the MeCP2 deficient M12 cell line. If expression can be restored, it will prove that the observed changes are reversible and the Rett phenotype can be rescued.
Aharon Razin, Ph.D.The Hebrew University of Jerusalem-Hadassah Medical School, Israel
"MeCP2 and Rett Syndrome"
1-Year Award: $50,000
Research Sponsor: Rett Syndrome Association of Illinois / Adam (Buddha) Lavey
Final Report (November 2003)
In 80% of Rett syndrome patients a mutation in the MeCP2 gene had been identified, leaving around 20% of Rett syndrome cases undiagnosed genetically. It is possible that these cases are inflicted with a mutation outside the gene, or that they carry a deletion of the entire gene. However, it cannot be ruled out that in some Rett Syndrome cases the MeCP2 gene is intact, a normal protein is produced, but one or more of its target genes are impaired. Our first attempt to examine these possibilities was to search for possible mutations in the promoter of the gene. To this end, we have analyzed samples from over 100 Rett syndrome patients from many countries around the world. We failed so far to identify any mutations in promoter sequences in these samples. However, we have identified in two Israeli Rett syndrome patients a bona fide mutation within another non coding sequence. This mutation (a deletion of one adenine residue) was found at a position which is believed to be essential for processing the gene product. We will study this mutation and try to verify its relevance to Rett Syndrome. We have already detected no MeCP2 protein in cells from this patient. If we will be convinced that this mutation is relevant to the Rett Syndrome, we will screen for this mutation in all the samples of the Rett syndrome patients that we analyzed for promoter mutations.
In parallel, we have generated a lymphoblast cell line from another Rett patient with no detectable mutation in the MeCP2 coding region. These cells have an intact MeCP2 protein and this may suggest that genes other than MeCP2 are involved in the disease. Such genes may be targets of MeCP2 activity or, alternatively, genes that code proteins that interact with MeCP2. A mutation in one of these genes could theoretically result in the Rett syndrome phenotype.
To identify MeCP2 target genes we have generated two immortalized lymphoblast cell lines originating from a Rett syndrome patient that carries a mutation in the coding sequence of the MeCP2 gene. One cell line expresses the MeCP2 protein (M10) and the other (M12), is MeCP2 deficient. We performed microarray analysis to compare the expression profile of 30,000 genes in these MeCP2 deficient and normal cell lines. This analysis revealed 90 genes with elevated expression levels in M12 compared to M10. Twelve genes with the largest change in expression level were chosen for further analysis.
The next step will be to try to restore the normal expression levels of these genes by introducing the normal MeCP2 gene into the MeCP2 deficient M12 cell line. If expression can be restored, it will prove that the observed changes are reversible and the Rett phenotype can be rescued.