ingest
cdrApp
2017-07-05T20:25:58.388Z
b743dd6f-fb31-445f-b2b9-a0aa4f8b7562
modifyDatastreamByValue
RELS-EXT
fedoraAdmin
2017-07-05T20:56:23.344Z
Setting exclusive relation
modifyDatastreamByValue
RELS-EXT
fedoraAdmin
2017-07-05T20:56:31.385Z
Setting exclusive relation
addDatastream
MD_TECHNICAL
fedoraAdmin
2017-07-05T20:56:39.292Z
Adding technical metadata derived by FITS
modifyDatastreamByValue
RELS-EXT
fedoraAdmin
2017-07-05T20:56:54.741Z
Setting exclusive relation
addDatastream
MD_FULL_TEXT
fedoraAdmin
2017-07-05T20:57:02.953Z
Adding full text metadata extracted by Apache Tika
modifyDatastreamByValue
RELS-EXT
fedoraAdmin
2017-07-05T20:57:18.730Z
Setting exclusive relation
modifyDatastreamByValue
RELS-EXT
cdrApp
2017-07-06T11:40:22.303Z
Setting exclusive relation
modifyDatastreamByValue
MD_DESCRIPTIVE
cdrApp
2018-01-25T05:28:52.465Z
modifyDatastreamByValue
MD_DESCRIPTIVE
cdrApp
2018-01-27T06:05:37.376Z
modifyDatastreamByValue
MD_DESCRIPTIVE
cdrApp
2018-03-14T02:11:57.504Z
modifyDatastreamByValue
MD_DESCRIPTIVE
cdrApp
2018-05-17T14:03:12.392Z
modifyDatastreamByValue
MD_DESCRIPTIVE
cdrApp
2018-07-11T00:41:37.556Z
modifyDatastreamByValue
MD_DESCRIPTIVE
cdrApp
2018-07-17T20:39:51.391Z
modifyDatastreamByValue
MD_DESCRIPTIVE
cdrApp
2018-08-08T20:07:23.125Z
modifyDatastreamByValue
MD_DESCRIPTIVE
cdrApp
2018-08-15T17:16:15.252Z
modifyDatastreamByValue
MD_DESCRIPTIVE
cdrApp
2018-08-16T20:18:36.390Z
modifyDatastreamByValue
MD_DESCRIPTIVE
cdrApp
2018-09-21T17:43:12.811Z
modifyDatastreamByValue
MD_DESCRIPTIVE
cdrApp
2018-09-26T20:52:43.174Z
modifyDatastreamByValue
MD_DESCRIPTIVE
cdrApp
2018-10-11T21:32:48.234Z
modifyDatastreamByValue
MD_DESCRIPTIVE
cdrApp
2019-03-20T14:59:14.741Z
Mary
Sperlazza
Author
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the brain and plays a critical role in neuron maturation, as mutations disrupting its function account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration of the chromatin architecture surrounding the mC mark. In these studies, we examine the behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the protein-protein interactions between subunits of NuRD.
Recent work suggests the primary effects of MeCP2 on gene expression in the developing mammalian brain are mediated by binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does not show high affinity or methyl-specific binding to mCA. Introduction of the Rett Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high affinity mCA site did not dramatically change binding properties, however hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic switch to redistribute MeCP2 among mCG and mCA loci.
Blocking recruitment of NuRD by MBD2 restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer. Towards understanding the recruitment of the NuRD components to the complex, this study characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2. We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and CHD4.
Spring 2017
2017
Biophysics
Biochemistry
epigenetics, methyl‑binding domain, protein‑DNA interaction, protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
University of North Carolina at Chapel Hill Graduate School
Degree granting institution
Chemistry
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
Mary
Sperlazza
Creator
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR
COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of
increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins
recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven
proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the
brain and plays a critical role in neuron maturation, as mutations disrupting its function
account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome
remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration
of the chromatin architecture surrounding the mC mark. In these studies, we examine the
behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the
protein-protein interactions between subunits of NuRD. Recent work suggests the primary
effects of MeCP2 on gene expression in the developing mammalian brain are mediated by
binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This
work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and
orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does
not show high affinity or methyl-specific binding to mCA. Introduction of the Rett
Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and
lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high
affinity mCA site did not dramatically change binding properties, however
hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We
suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic
switch to redistribute MeCP2 among mCG and mCA loci. Blocking recruitment of NuRD by MBD2
restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced
tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in
cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting
MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer.
Towards understanding the recruitment of the NuRD components to the complex, this study
characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2.
We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and
CHD4.
Spring 2017
2017
Biophysics
Biochemistry
epigenetics, methyl‑binding domain, protein‑DNA
interaction, protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
University of North Carolina at Chapel Hill Graduate School
Degree granting
institution
Chemistry
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
Mary
Sperlazza
Creator
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the brain and plays a critical role in neuron maturation, as mutations disrupting its function account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration of the chromatin architecture surrounding the mC mark. In these studies, we examine the behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the protein-protein interactions between subunits of NuRD. Recent work suggests the primary effects of MeCP2 on gene expression in the developing mammalian brain are mediated by binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does not show high affinity or methyl-specific binding to mCA. Introduction of the Rett Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high affinity mCA site did not dramatically change binding properties, however hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic switch to redistribute MeCP2 among mCG and mCA loci. Blocking recruitment of NuRD by MBD2 restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer. Towards understanding the recruitment of the NuRD components to the complex, this study characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2. We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and CHD4.
Spring 2017
2017
Biophysics
Biochemistry
epigenetics, methyl‑binding domain, protein‑DNA interaction, protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
University of North Carolina at Chapel Hill Graduate School
Degree granting institution
Chemistry
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
Mary
Sperlazza
Creator
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the brain and plays a critical role in neuron maturation, as mutations disrupting its function account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration of the chromatin architecture surrounding the mC mark. In these studies, we examine the behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the protein-protein interactions between subunits of NuRD. Recent work suggests the primary effects of MeCP2 on gene expression in the developing mammalian brain are mediated by binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does not show high affinity or methyl-specific binding to mCA. Introduction of the Rett Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high affinity mCA site did not dramatically change binding properties, however hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic switch to redistribute MeCP2 among mCG and mCA loci. Blocking recruitment of NuRD by MBD2 restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer. Towards understanding the recruitment of the NuRD components to the complex, this study characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2. We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and CHD4.
2017-05
2017
Biophysics
Biochemistry
epigenetics, methyl‑binding domain, protein‑DNA interaction, protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
University of North Carolina at Chapel Hill Graduate School
Degree granting institution
Chemistry
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
Mary
Sperlazza
Creator
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the brain and plays a critical role in neuron maturation, as mutations disrupting its function account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration of the chromatin architecture surrounding the mC mark. In these studies, we examine the behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the protein-protein interactions between subunits of NuRD. Recent work suggests the primary effects of MeCP2 on gene expression in the developing mammalian brain are mediated by binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does not show high affinity or methyl-specific binding to mCA. Introduction of the Rett Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high affinity mCA site did not dramatically change binding properties, however hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic switch to redistribute MeCP2 among mCG and mCA loci. Blocking recruitment of NuRD by MBD2 restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer. Towards understanding the recruitment of the NuRD components to the complex, this study characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2. We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and CHD4.
2017
Biophysics
Biochemistry
epigenetics, methyl‑binding domain, protein‑DNA interaction, protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
University of North Carolina at Chapel Hill Graduate School
Degree granting institution
Chemistry
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
2017-05
Mary
Sperlazza
Creator
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the brain and plays a critical role in neuron maturation, as mutations disrupting its function account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration of the chromatin architecture surrounding the mC mark. In these studies, we examine the behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the protein-protein interactions between subunits of NuRD. Recent work suggests the primary effects of MeCP2 on gene expression in the developing mammalian brain are mediated by binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does not show high affinity or methyl-specific binding to mCA. Introduction of the Rett Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high affinity mCA site did not dramatically change binding properties, however hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic switch to redistribute MeCP2 among mCG and mCA loci. Blocking recruitment of NuRD by MBD2 restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer. Towards understanding the recruitment of the NuRD components to the complex, this study characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2. We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and CHD4.
2017
Biophysics
Biochemistry
epigenetics, methyl‑binding domain, protein‑DNA interaction, protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
University of North Carolina at Chapel Hill Graduate School
Degree granting institution
Chemistry
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
2017-05
Mary
Sperlazza
Creator
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the brain and plays a critical role in neuron maturation, as mutations disrupting its function account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration of the chromatin architecture surrounding the mC mark. In these studies, we examine the behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the protein-protein interactions between subunits of NuRD. Recent work suggests the primary effects of MeCP2 on gene expression in the developing mammalian brain are mediated by binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does not show high affinity or methyl-specific binding to mCA. Introduction of the Rett Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high affinity mCA site did not dramatically change binding properties, however hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic switch to redistribute MeCP2 among mCG and mCA loci. Blocking recruitment of NuRD by MBD2 restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer. Towards understanding the recruitment of the NuRD components to the complex, this study characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2. We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and CHD4.
2017
Biophysics
Biochemistry
epigenetics, methyl‑binding domain, protein‑DNA interaction, protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
University of North Carolina at Chapel Hill Graduate School
Degree granting institution
Chemistry
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
2017-05
Mary
Sperlazza
Creator
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the brain and plays a critical role in neuron maturation, as mutations disrupting its function account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration of the chromatin architecture surrounding the mC mark. In these studies, we examine the behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the protein-protein interactions between subunits of NuRD. Recent work suggests the primary effects of MeCP2 on gene expression in the developing mammalian brain are mediated by binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does not show high affinity or methyl-specific binding to mCA. Introduction of the Rett Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high affinity mCA site did not dramatically change binding properties, however hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic switch to redistribute MeCP2 among mCG and mCA loci. Blocking recruitment of NuRD by MBD2 restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer. Towards understanding the recruitment of the NuRD components to the complex, this study characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2. We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and CHD4.
2017
Biophysics
Biochemistry
epigenetics, methyl‑binding domain, protein‑DNA interaction, protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
University of North Carolina at Chapel Hill Graduate School
Degree granting institution
Chemistry
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
2017-05
Mary
Sperlazza
Creator
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the brain and plays a critical role in neuron maturation, as mutations disrupting its function account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration of the chromatin architecture surrounding the mC mark. In these studies, we examine the behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the protein-protein interactions between subunits of NuRD. Recent work suggests the primary effects of MeCP2 on gene expression in the developing mammalian brain are mediated by binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does not show high affinity or methyl-specific binding to mCA. Introduction of the Rett Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high affinity mCA site did not dramatically change binding properties, however hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic switch to redistribute MeCP2 among mCG and mCA loci. Blocking recruitment of NuRD by MBD2 restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer. Towards understanding the recruitment of the NuRD components to the complex, this study characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2. We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and CHD4.
2017
Biophysics
Biochemistry
epigenetics, methyl‑binding domain, protein‑DNA interaction, protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
Chemistry
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
2017-05
University of North Carolina at Chapel Hill
Degree granting institution
Mary
Sperlazza
Creator
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the brain and plays a critical role in neuron maturation, as mutations disrupting its function account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration of the chromatin architecture surrounding the mC mark. In these studies, we examine the behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the protein-protein interactions between subunits of NuRD. Recent work suggests the primary effects of MeCP2 on gene expression in the developing mammalian brain are mediated by binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does not show high affinity or methyl-specific binding to mCA. Introduction of the Rett Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high affinity mCA site did not dramatically change binding properties, however hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic switch to redistribute MeCP2 among mCG and mCA loci. Blocking recruitment of NuRD by MBD2 restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer. Towards understanding the recruitment of the NuRD components to the complex, this study characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2. We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and CHD4.
2017
Biophysics
Biochemistry
epigenetics, methyl‑binding domain, protein‑DNA interaction, protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
University of North Carolina at Chapel Hill Graduate School
Degree granting institution
Chemistry
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
2017-05
Mary
Sperlazza
Creator
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the brain and plays a critical role in neuron maturation, as mutations disrupting its function account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration of the chromatin architecture surrounding the mC mark. In these studies, we examine the behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the protein-protein interactions between subunits of NuRD. Recent work suggests the primary effects of MeCP2 on gene expression in the developing mammalian brain are mediated by binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does not show high affinity or methyl-specific binding to mCA. Introduction of the Rett Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high affinity mCA site did not dramatically change binding properties, however hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic switch to redistribute MeCP2 among mCG and mCA loci. Blocking recruitment of NuRD by MBD2 restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer. Towards understanding the recruitment of the NuRD components to the complex, this study characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2. We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and CHD4.
2017
Biophysics
Biochemistry
epigenetics, methyl‑binding domain, protein‑DNA interaction, protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
University of North Carolina at Chapel Hill Graduate School
Degree granting institution
Chemistry
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
2017-05
Mary
Sperlazza
Creator
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the brain and plays a critical role in neuron maturation, as mutations disrupting its function account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration of the chromatin architecture surrounding the mC mark. In these studies, we examine the behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the protein-protein interactions between subunits of NuRD. Recent work suggests the primary effects of MeCP2 on gene expression in the developing mammalian brain are mediated by binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does not show high affinity or methyl-specific binding to mCA. Introduction of the Rett Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high affinity mCA site did not dramatically change binding properties, however hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic switch to redistribute MeCP2 among mCG and mCA loci. Blocking recruitment of NuRD by MBD2 restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer. Towards understanding the recruitment of the NuRD components to the complex, this study characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2. We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and CHD4.
2017
Biophysics
Biochemistry
epigenetics; methyl‑binding domain; protein‑DNA interaction; protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
University of North Carolina at Chapel Hill Graduate School
Degree granting institution
Chemistry
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
2017-05
Mary
Sperlazza
Creator
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the brain and plays a critical role in neuron maturation, as mutations disrupting its function account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration of the chromatin architecture surrounding the mC mark. In these studies, we examine the behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the protein-protein interactions between subunits of NuRD. Recent work suggests the primary effects of MeCP2 on gene expression in the developing mammalian brain are mediated by binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does not show high affinity or methyl-specific binding to mCA. Introduction of the Rett Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high affinity mCA site did not dramatically change binding properties, however hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic switch to redistribute MeCP2 among mCG and mCA loci. Blocking recruitment of NuRD by MBD2 restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer. Towards understanding the recruitment of the NuRD components to the complex, this study characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2. We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and CHD4.
2017
Biophysics
Biochemistry
epigenetics, methyl‑binding domain, protein‑DNA interaction, protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
University of North Carolina at Chapel Hill Graduate School
Degree granting institution
Chemistry
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
2017-05
Mary
Sperlazza
Creator
Department of Chemistry
College of Arts and Sciences
STRUCTURE AND FUNCTION STUDIES OF METHYL-CPG BINDING DOMAIN PROTEINS AND THEIR COMPLEXES
DNA methylation is an epigenetic mechanism of transcriptional silencing of increasing interest for treating human disease. Methyl-CpG binding domain (MBD) proteins recognize 5 methylcytosines (mC) primarily in symmetrical CpG (mCG) dinucleotides. Seven proteins comprise the MBD family, MBD1-6 and MeCP2. MeCP2 is primarily expressed in the brain and plays a critical role in neuron maturation, as mutations disrupting its function account for up to 80% of Rett Syndrome cases. MBD2/3 associates with the nucleosome remodeling and deacetylase complex (NuRD) and modulates gene expression through alteration of the chromatin architecture surrounding the mC mark. In these studies, we examine the behavior of the MBDs of MeCP2 and MBD2, in addition to further characterizing the protein-protein interactions between subunits of NuRD. Recent work suggests the primary effects of MeCP2 on gene expression in the developing mammalian brain are mediated by binding asymmetrically methylated and hydroxymethylated CpA (h/mCA) dinucleotides. This work establishes that the MeCP2 MBD binds mCA with high affinity in a strand specific and orientation dependent manner. This preference is specific to MeCP2, as the MBD2 MBD does not show high affinity or methyl-specific binding to mCA. Introduction of the Rett Syndrome-associated mutations T158M, R106W and P101S destabilized the MeCP2 MBD and lessened recognition of mCG and mCA equally. Finally, hydroxymethylation of a high affinity mCA site did not dramatically change binding properties, however hemi-hydroxylation of the same cytosine in mCG significantly decreased affinity. We suggest a model for MeCP2 recognition of mCA and for hydroxymethylation as an epigenetic switch to redistribute MeCP2 among mCG and mCA loci. Blocking recruitment of NuRD by MBD2 restores expression of developmentally silenced fetal hemoglobin and aberrantly silenced tumor suppressor genes. Additionally, knockdown of the NuRD helicase, CHD4, results in cancer cells growth arrest and increased sensitivity to DNA damage. Therefore, targeting MBD2-NuRD presents a promising avenue for treating β-hemoglobinopathies and cancer. Towards understanding the recruitment of the NuRD components to the complex, this study characterizes the GATA-like zinc-finger domains of the NuRD components GATAD2A and MTA2. We propose a model of NuRD in which MTA2 binds DNA and GATAD2A serves to bridge MBD2 and CHD4.
2017
Biophysics
Biochemistry
epigenetics; methyl‑binding domain; protein‑DNA interaction; protein‑protein interaction
eng
Doctor of Philosophy
Dissertation
University of North Carolina at Chapel Hill Graduate School
Degree granting institution
David
Williams
Thesis advisor
Dorothy
Erie
Thesis advisor
Eric
Brustad
Thesis advisor
Marcey
Waters
Thesis advisor
Brian
Kuhlman
Thesis advisor
text
2017-05
Sperlazza_unc_0153D_16792.pdf
uuid:5f42e130-07d5-4f57-b2e9-6cfc267a3a98
proquest
2019-07-05T00:00:00
2017-04-05T23:47:40Z
application/pdf
4402003
yes