Rett Syndrome is caused by mutations in X-linked MECP2, encoding methyl-CpG-binding protein 2

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Rett Syndrome is caused by mutations in X-linked
MECP2, encoding methyl-CpG-binding protein 2

• Presented By
• Vanessa Bowers

Amir, R., I.B. Van den Veyver, M. Wan, C.Q. Tran,
U. Francke, and H.Y. Zoghbi. 1999. Rett Syndrome
is caused by mutations in X-linked MECP2,
encoding methyl-CpG-binding protein 2. Nature
Genetics. 23185-188.
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What is Rett Syndrome

• A progressive neurodevelopmental disorder that
  begins at early infancy
• Caused by a mutation in the methyl-cysteine
  binding protein 2 gene (MeCP2)
• Most common cause of mental retardation in
  females
• Shows high lethality in hemizygous males (causing
  miscarriage, still birth, or early death)

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Symptoms Stages of Rett Syndrome(RTT)

• Patients with RTT appear to be born healthy and
  develop normally until about 6-18 months of age
• Several stages then follow
• After initial regression, the condition
  stabilizes and female patients usually survive
  into adulthood

Stages Age Symptoms
Stage 1 6-18 months Disinterest in play, hypotonia
Stage 2 1-3 years Rapid regression, irritability, autistic like symptoms, microcephaly
Stage 3 2-10 years Seizures, microcephaly, mental retardation, hand-wringing, hyperventilation, bruxism, ataxia, aerophagi
Stage 4 10 years Scoliosis, muscle wasting, rigidity, improved eye contact
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What is the MeCP2 gene?

• MeCP2 gene is located on the X-chromosome (mapped
  to the region Xq28)
• It is a transcriptional repressor that prevents
  unscheduled transcription throughout the body
• The MeCP2 gene encodes the MeCP2 protein which
  binds methylated cytosine residue of DNA
  dinucleotides and mediates transcriptional
  silencing through the interaction with histone
  deacetylase and the corepressor mSIN3A
• These cytosine residues are subject to
  transcriptional silencing after DNA methylation

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Why is transcriptional silencing important in the
body?

• It is a mechanism of transcriptional control
  where DNA in no longer accessible for future
  transcription
• This is important because in different stages of
  development, certain genes need to be silenced or
  turned off and others need to be turned on in
  order for proper development
• In RTT, the mutation of the MeCP2 gene doesnt
  allow for normal transcriptional silencing,
  therefore causing the disease

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What does the MeCP2 protein look like?

• MeCP2 protein is a chain of 486 amino acids
• Two functional domains
• - an 85 amino acid methyl-CpG binding domain
  (MBD)
• - a 104 amino acid transcription repression
  domain (TRD)
• The MBD is essential for the binding of the
  protein to methyl cytosine at the beginning of
  genes in the body
• The TRD recruits other repressor proteins
  (mSIN3a, histone deascetylase) to inhibit
  transcription

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Where does RTT originate?

• RTT is caused by mutations in the MeCP2 gene,
  which causes changes in the corresponding MeCP2
  protein
• Mutations occur in either of the parents
  reproductive cells sperm or egg
• Mutations are usually sporadic and not familial

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There are two scenarios for the origin of
mutations in Rett Syndrome 1) The
father has a mutation on his X chromosome or
2) The mother has a mutation on one of her
x chromosomes
Scenario 1
Scenario 2
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Rett Syndrome is Neurodevelopmental Disorder

• MeCP2 is very abundant and highly expressed in
  the brain. In the developing cerebral cortex, the
  appearance of MeCP2 correlates with neuronal
  maturation.
• With this high abundance of MeCP2 within the
  brain, these mutations of the MeCP2 protein
  affect the development of neurons and therefore
  cause many of the neurological symptoms that are
  related to Rett Syndrome

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The Experiment

• 21 sporadic and 8 familial RTT patients genomic
  DNA were screened by conformation-sensitive gel
  electrophoresis
• Sporadic patients no known family members
  expressed the disorder
• Familial patients were
• 5 pairs of full sisters
• 2 pairs of half sisters
• 1 pair of second half cousins

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Results

• Among sporadic patients 3 missense,
• 1 nonsense, and 1 frameshift mutations were
  identified among patients 6, 22, 24, 29, and 39
• In two half sisters, the same missense mutation
  was found in each, but was not present in the
  mother

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Quick Definitions

• Missense mutation - A mutation that changes a
  codon so that it codes for a different amino acid
  
• Nonsense mutation - A mutation in which one of
  the three terminator codons in the RNA (TAG, TAA,
  TGA), used to signal the end of a polypeptide,
  appears in the middle of a genetic message,
  causes premature termination of transcription,
  and releases incomplete, nonfunctional
  polypeptides from the ribosome.
• Frameshift mutation - A mutation resulting from
  an addition or subtraction that is not an exact
  multiple of 3 base pairs in a coding sequence.
  From the point of mutation onwards, codons are
  read out of phase the reading frame of the gene
  is changed, and a completely different set of
  amino acids are made into protein.

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Patient 39 a missense mutation occurred at amino
acid 133, arginine was replaced by cysteine
Patient 24 a missense mutation occurred at
amino acid 155, thymine was replaced by
cytosine Patient 6 a missense mutation
occurred at amino acid 158, cytosine was
replaced by thymine
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• Patient 22 a nonsense mutation occurred with a
  substitution of cytosine to thymine, which
  converted a CGA codon to a TGA codon

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• Patient 29 A frameshift mutation occurred from
  an insertion at codon 208 which shifted the
  reading frame and introduced a stop codon after
  27 amino acids

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Familial Patients Results

• In two half sisters with the same mother, the
  same missense mutation of cytosine replaced by
  thymine was identified
• No mutation was found in the mother (she is an
  obligate carrier)

Sister
Mother
Sister
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Discussion

• These missense mutations disrupt the structure of
  the methyl binding region (MBR) of the MeCP2
  protein
• The nonsense and frameshift mutations disrupt the
  transcription repression domain (TRD)
• These disruptions of the MBD and the TRD of the
  MeCP2 protein interferes with their functions of
  transcriptional silencing and cause Rett syndrome

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Conclusion

• Rett syndrome patients display some type of
  mutation (missense, nonsense, or frameshift).
  These mutations affect the MBD or TRD domains of
  the MeCP2 protein, which cause defects that dont
  allow for the proper functioning of the MeCP2
  gene as a transcriptional silencer. Without
  proper functioning of this gene, patients will
  develop Rett Syndrome.

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• QUESTIONS?