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Histone post-translational modifications in frontal cortex from human donors with Alzheimer's disease.

Anderson KW, Turko IV - Clin Proteomics (2015)

Bottom Line: Histone post-translational modifications (PTMs) are a key element in epigenetic regulation of gene expression and are known to be associated with the pathology of numerous diseases.Of the changes observed, notable decreases in methylation of H2B residue K108 by 25 % and H4 residue R55 by 35 % were measured and are likely associated with hydrogen bonding networks important for nucleosome stability.Beyond the structural and functional impacts of the changes we have measured, the sites of altered PTMs may be used to identify enzymes responsible for their modulation, which could be used as prospective drug targets for highly specific AD therapies.

View Article: PubMed Central - PubMed

Affiliation: Institute for Bioscience and Biotechnology Research, Rockville, MD 20850 USA ; Biomolecular Measurement Division, National Institute of Standards and Technology, Gaithersburg, MD 20899 USA ; Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742 USA.

ABSTRACT

Background: Alzheimer's disease (AD) is the sixth leading cause of death and the most costly disease in the US. Despite the enormous impact of AD, there are no treatments that delay onset or stop disease progression currently on the market. This is partly due to the complexity of the disease and the largely unknown pathogenesis of sporadic AD, which accounts for the vast majority of cases. Epigenetics has been implicated as a critical component to AD pathology and a potential "hot spot" for treatments. Histone post-translational modifications (PTMs) are a key element in epigenetic regulation of gene expression and are known to be associated with the pathology of numerous diseases. Investigation of histone PTMs can help elucidate AD pathology and identify targets for therapies.

Results: A multiple reaction monitoring mass spectrometry assay was used to measure changes in abundance of several histone PTMs in frontal cortex from human donors affected with AD (n = 6) and age-matched, normal donors (n = 6). Of the changes observed, notable decreases in methylation of H2B residue K108 by 25 % and H4 residue R55 by 35 % were measured and are likely associated with hydrogen bonding networks important for nucleosome stability. Additionally, a 91 % increase in ubiquitination of K120 on H2B was measured as well as an apparent loss in acetylation of the region near the N-terminus of H4. Our method of quantification was also determined to be precise and robust, signifying measured changes were representative of true biological differences between donors and sample groups.

Conclusion: We are the first to report changes in methylation of H2B K108, methylation of H4 R55, and ubiquitination of H2B K120 in frontal cortex from human donors with AD. These notable PTM changes may be of great importance in elucidating the epigenetic mechanism of AD as it relates to disease pathology. Beyond the structural and functional impacts of the changes we have measured, the sites of altered PTMs may be used to identify enzymes responsible for their modulation, which could be used as prospective drug targets for highly specific AD therapies.

No MeSH data available.


Related in: MedlinePlus

Location of core PTMs on nucleosome surface. H2A (blue), H2B (pink), H3 (yellow), and H4 (green) form an octamer in the nucleosome core, which is wrapped by DNA. Nucleosome crystal structure PDB ID: 2CV5 [25] was modified using PyMOL Molecular Graphics System (Version 1.7.2, Schrödinger, LLC). a Methylation sites R55 on H4 and K108 on H3 and ubiquitination site K121 on H2B are shown in red. b Hydrogen bond network of H4 R55. c Hydrogen bond network of H2B K108
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Fig3: Location of core PTMs on nucleosome surface. H2A (blue), H2B (pink), H3 (yellow), and H4 (green) form an octamer in the nucleosome core, which is wrapped by DNA. Nucleosome crystal structure PDB ID: 2CV5 [25] was modified using PyMOL Molecular Graphics System (Version 1.7.2, Schrödinger, LLC). a Methylation sites R55 on H4 and K108 on H3 and ubiquitination site K121 on H2B are shown in red. b Hydrogen bond network of H4 R55. c Hydrogen bond network of H2B K108

Mentions: While HDAC has demonstrated potential as a prospective AD therapy, therapies targeting other histone-modifying enzymes have yet to come forward. Other PTMs, such as methylation, are modulated by enzymes with greater residue specificity than acetyltransferases and deacetylases. Often times, a methyltransferase or demethylase may only act on one site in a histone [23]. This specificity may correlate to improved efficacy and safety when targeting enzymes for specific methylation sites compared to HDAC inhibition alone. Additionally, they may be more efficacious as a combinatorial treatment. H2B methylation at K108 decreased 25 % and H4 methylation at R55 decreased 35 % in AD frontal cortex (Fig. 2b). Both of these sites have been reported to be methylated in human cells [24]. To our knowledge, H2B K108 and H4 R55 methylation have not been investigated in AD-affected human frontal cortex. Both H2B K108 and H4 R55 are located on the outermost surface of the nucleosome (Fig. 3a) [25], suggesting their accessibility to methyltransferases and demethylases may incline them to be used as switches for transcriptional activity. Monomethylation is commonly a marker of transcription activation. Since AD is characterized by a global repression in gene expression, our observed decrease in several methylation sites strongly suggests their role in reducing transcription consistent with AD. H4 R55 participates in a complex hydrogen bond network in the nucleosome (Fig. 3b) [25]. H4 residues Q27 and E52 and the I29 backbone form a hydrogen bond network that structurally thrusts N25 into a hydrogen bond with E73 on neighboring histone H3 [26]. By forming a hydrogen bonding network that increases affinity between H3 and H4, the nucleosome becomes structurally more stable. However, methylation of H4 R55 likely disrupts this bonding network, thereby lessening the H3–H4 interaction and weakening the nucleosome [26]. Our observed decrease in methylation of R55 may be a mechanism for increasing nucleosome stability and favoring denser chromatin, which decreases gene expression. Similarly, H2B K108 is central to a hydrogen bond network involving histones H2A, H2B, and H4 (Fig. 3c) [25]. H2B K108 and E105 participate in intra-chain hydrogen bonding which structurally positions H2B residues for two interactions between neighboring histones favoring nucleosome stability. First, K108 forms a hydrogen bond with E105, pulling E105 away from close proximity to E93 on histone H2A. E105 and E93 can create electrostatic repulsion, however when E105 is distanced from E93 and hydrogen bonded to K108, this electrostatic repulsion is greatly reduced and H2A and H2B are able nestle together [25]. Second, the K108 hydrogen bond with E105 twists the helix-loop-helix, shown in Fig. 3c, to position R99 on H2B directly facing T71 on H4 for a hydrogen bond. This hydrogen bond between residues on H2B and H4 strengthen the H2B–H4 stability. Methylation likely disturbs this H4–H2B–H2A inter-chain network and would favor disassembly of the nucleosome. The structural and functional effect of our observed decrease in methylation of H2B K108 is consistent with the decrease in H4 R55, suggesting methylation of histones is used to down regulate transcriptional activity and gene expression in AD pathology. Establishing specific sites of histone methylation and their modulation, as described herein, is important in characterizing AD pathology.Fig. 3


Histone post-translational modifications in frontal cortex from human donors with Alzheimer's disease.

Anderson KW, Turko IV - Clin Proteomics (2015)

Location of core PTMs on nucleosome surface. H2A (blue), H2B (pink), H3 (yellow), and H4 (green) form an octamer in the nucleosome core, which is wrapped by DNA. Nucleosome crystal structure PDB ID: 2CV5 [25] was modified using PyMOL Molecular Graphics System (Version 1.7.2, Schrödinger, LLC). a Methylation sites R55 on H4 and K108 on H3 and ubiquitination site K121 on H2B are shown in red. b Hydrogen bond network of H4 R55. c Hydrogen bond network of H2B K108
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4591557&req=5

Fig3: Location of core PTMs on nucleosome surface. H2A (blue), H2B (pink), H3 (yellow), and H4 (green) form an octamer in the nucleosome core, which is wrapped by DNA. Nucleosome crystal structure PDB ID: 2CV5 [25] was modified using PyMOL Molecular Graphics System (Version 1.7.2, Schrödinger, LLC). a Methylation sites R55 on H4 and K108 on H3 and ubiquitination site K121 on H2B are shown in red. b Hydrogen bond network of H4 R55. c Hydrogen bond network of H2B K108
Mentions: While HDAC has demonstrated potential as a prospective AD therapy, therapies targeting other histone-modifying enzymes have yet to come forward. Other PTMs, such as methylation, are modulated by enzymes with greater residue specificity than acetyltransferases and deacetylases. Often times, a methyltransferase or demethylase may only act on one site in a histone [23]. This specificity may correlate to improved efficacy and safety when targeting enzymes for specific methylation sites compared to HDAC inhibition alone. Additionally, they may be more efficacious as a combinatorial treatment. H2B methylation at K108 decreased 25 % and H4 methylation at R55 decreased 35 % in AD frontal cortex (Fig. 2b). Both of these sites have been reported to be methylated in human cells [24]. To our knowledge, H2B K108 and H4 R55 methylation have not been investigated in AD-affected human frontal cortex. Both H2B K108 and H4 R55 are located on the outermost surface of the nucleosome (Fig. 3a) [25], suggesting their accessibility to methyltransferases and demethylases may incline them to be used as switches for transcriptional activity. Monomethylation is commonly a marker of transcription activation. Since AD is characterized by a global repression in gene expression, our observed decrease in several methylation sites strongly suggests their role in reducing transcription consistent with AD. H4 R55 participates in a complex hydrogen bond network in the nucleosome (Fig. 3b) [25]. H4 residues Q27 and E52 and the I29 backbone form a hydrogen bond network that structurally thrusts N25 into a hydrogen bond with E73 on neighboring histone H3 [26]. By forming a hydrogen bonding network that increases affinity between H3 and H4, the nucleosome becomes structurally more stable. However, methylation of H4 R55 likely disrupts this bonding network, thereby lessening the H3–H4 interaction and weakening the nucleosome [26]. Our observed decrease in methylation of R55 may be a mechanism for increasing nucleosome stability and favoring denser chromatin, which decreases gene expression. Similarly, H2B K108 is central to a hydrogen bond network involving histones H2A, H2B, and H4 (Fig. 3c) [25]. H2B K108 and E105 participate in intra-chain hydrogen bonding which structurally positions H2B residues for two interactions between neighboring histones favoring nucleosome stability. First, K108 forms a hydrogen bond with E105, pulling E105 away from close proximity to E93 on histone H2A. E105 and E93 can create electrostatic repulsion, however when E105 is distanced from E93 and hydrogen bonded to K108, this electrostatic repulsion is greatly reduced and H2A and H2B are able nestle together [25]. Second, the K108 hydrogen bond with E105 twists the helix-loop-helix, shown in Fig. 3c, to position R99 on H2B directly facing T71 on H4 for a hydrogen bond. This hydrogen bond between residues on H2B and H4 strengthen the H2B–H4 stability. Methylation likely disturbs this H4–H2B–H2A inter-chain network and would favor disassembly of the nucleosome. The structural and functional effect of our observed decrease in methylation of H2B K108 is consistent with the decrease in H4 R55, suggesting methylation of histones is used to down regulate transcriptional activity and gene expression in AD pathology. Establishing specific sites of histone methylation and their modulation, as described herein, is important in characterizing AD pathology.Fig. 3

Bottom Line: Histone post-translational modifications (PTMs) are a key element in epigenetic regulation of gene expression and are known to be associated with the pathology of numerous diseases.Of the changes observed, notable decreases in methylation of H2B residue K108 by 25 % and H4 residue R55 by 35 % were measured and are likely associated with hydrogen bonding networks important for nucleosome stability.Beyond the structural and functional impacts of the changes we have measured, the sites of altered PTMs may be used to identify enzymes responsible for their modulation, which could be used as prospective drug targets for highly specific AD therapies.

View Article: PubMed Central - PubMed

Affiliation: Institute for Bioscience and Biotechnology Research, Rockville, MD 20850 USA ; Biomolecular Measurement Division, National Institute of Standards and Technology, Gaithersburg, MD 20899 USA ; Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742 USA.

ABSTRACT

Background: Alzheimer's disease (AD) is the sixth leading cause of death and the most costly disease in the US. Despite the enormous impact of AD, there are no treatments that delay onset or stop disease progression currently on the market. This is partly due to the complexity of the disease and the largely unknown pathogenesis of sporadic AD, which accounts for the vast majority of cases. Epigenetics has been implicated as a critical component to AD pathology and a potential "hot spot" for treatments. Histone post-translational modifications (PTMs) are a key element in epigenetic regulation of gene expression and are known to be associated with the pathology of numerous diseases. Investigation of histone PTMs can help elucidate AD pathology and identify targets for therapies.

Results: A multiple reaction monitoring mass spectrometry assay was used to measure changes in abundance of several histone PTMs in frontal cortex from human donors affected with AD (n = 6) and age-matched, normal donors (n = 6). Of the changes observed, notable decreases in methylation of H2B residue K108 by 25 % and H4 residue R55 by 35 % were measured and are likely associated with hydrogen bonding networks important for nucleosome stability. Additionally, a 91 % increase in ubiquitination of K120 on H2B was measured as well as an apparent loss in acetylation of the region near the N-terminus of H4. Our method of quantification was also determined to be precise and robust, signifying measured changes were representative of true biological differences between donors and sample groups.

Conclusion: We are the first to report changes in methylation of H2B K108, methylation of H4 R55, and ubiquitination of H2B K120 in frontal cortex from human donors with AD. These notable PTM changes may be of great importance in elucidating the epigenetic mechanism of AD as it relates to disease pathology. Beyond the structural and functional impacts of the changes we have measured, the sites of altered PTMs may be used to identify enzymes responsible for their modulation, which could be used as prospective drug targets for highly specific AD therapies.

No MeSH data available.


Related in: MedlinePlus