1. IDENTIFICATION OF THE MOLECULAR MECHANISMS IN RETT SYNDROME AND RELATED DISORDERS (RTT-GENET) X

2. A childhood neuro-developmental disorder Seen almost exclusively in females Found in a variety of racial and ethnic groups worldwide Dr. Andreas Rett (first description-1966) Dr. Bengt Hagberg (worldwide recognition-1983) 1/10,000-15,000 females 99.5% sporadic :high rate of new mutations 1/3 – 1/4 of progressive developmental disabilities in girls ~ 10% of profound disability in females RTT variants Onset of infantile seizuresOnset of infantile seizures Congenital formCongenital form Forme frusteForme fruste Late childhood regressionLate childhood regression Preserved speech variantPreserved speech variant Rett Syndrome

3. X Exclusion mapping “Minimal Critical Region” Haplotype Analysis Paternal Maternal X chrom. Genetics of the disease –Not a genetic disease –Mitochondrial inheritance –Autosomal inheritance with sex limited expression –X-linked dominant inheritance with lethality in males

4. Search for the gene Minimal Critical Region Includes Xq28 a chromosome band rich in disease genes Candidate gene approach Systematic sequencing of all the genes (over 200) in Xq28 was Dr. Uta Francke (Stanford) and Dr. Huda Zoghbi (Baylor) Identification of the RTT gene After 14 years of search the RTT gene was finally identified in 1999 MECP2 MECP2 (Methyl CpG binding protein 2)

5. MeCP2 protein 486 amino acids and 52kD. An abundant mammalian chromosomal protein that binds to methylated CpG. Ubiquitously expressed, more abundant in brain. Can bind to single methyl-CpG pair (unlike MeCP1 which requires >10 methyl-CpGs to bind DNA) Contains four functional domains: –A methyl-CpG binding domain(MBD) –A transcriptional repression domain (TRD) –Nuclear localization signal (NLS) –C-terminal segment Potentially involved in global gene silencing Potentially involved in global gene silencing.

6. Interaction of MeCP2 protein with HDAC

7. MECP2 mutations identified Exons II III IV 316C  T R106W 502 C  T R168X 763 C  T R255X 397C  T R133C 473C  T T158M 880 C  T R294X 916C  T R306C 1 26 235 376 486 625 930 1461 808 C  T R270X Recurrent MECP2 mutations

8. Rett Syndrome is the first human disease found to be caused by defects in a protein involved in regulation of gene expression through its interaction with methylated DNA. Important!

9. Methylated promoter MeCP2 Transcriptional silencing Transcriptional noise Mutated MeCP2 ? trc A highly likely scenario on the consequences of MECP2 mutations.

10. Important issues Identification of downstream genes regulated by MECP2 MECP2 mutations and epigenetics X-inactivation Genomic imprinting Animal models Using conditional knock-out technology, mice that lack MECP2 either in all tissues or selectively in brain was generated (Bird et al., 2001, Guy et al. 2001). Cellular defects associated with MeCP2 deficiency in mouse CNS? Behavioral defects in mice? Drugs, gene therapy, stem cells with normal MECP2 can be the solution for treatment of RTT

11. A %T allele %C allele 41 (36-47) 44 (36-50) 36 (28-43) 59 (53-64) 56 (50-64) 64 (57-72) Male Cases