scholarly journals Molecular basis of the selective methylation of histone H3.1

2014 ◽  
Vol 70 (a1) ◽  
pp. C1584-C1584 ◽  
Author(s):  
Jean-Francois Couture
Keyword(s):  
Structural Studies ◽  
Histone H4 ◽  
Set Domain ◽  
Methyltransferase Activity ◽  
Histone Proteins ◽  
Post Translational Modifications ◽  
Heterochromatin Formation ◽  
Functional Consequences ◽  
Binding Cleft ◽  
Critical Components

Histone proteins are critical components of the chromatin fiber. With the exception of histone H4, histone proteins exist as different variants with assigned specialized functions and are proposed to act as key elements for the selective deposition of histone post-translational modifications (PTMs). Among these variants, H3.3, which differs from H3.1 by only 5 residues, has been predominantly found in the proximity of genes that are highly expressed; however the functional consequences of that marking have remained enigmatic. Herein we report the crystal structure of the trithorax-related SET domain histone H3.1 K27 methyltransferase ATXR5 in complex with histone H3.1 and the product cofactor S-adenosylhomocysteine. Overall, the SET domain folds as an all β-strands structure preceded by the pre-SET domain which folds as three long α-helices. The histone H3 is maintained in an elongated conformation by residues located both in the SET and pre-SET domains of ATXR5. Interestingly, we found that a three residue loop folds back on top of the ATXR histone H3.1 binding cleft shielding the peptide from the solvent and maintaining P30 and A31 of H3.1 in a shallow hydrophobic pocket. Strikingly, substitution of A31 for a threonine, the corresponding residue in histone H3.3 in plant, severely impairs ATXR5 methyltransferase activity. Finally, biochemical and structural studies revealed that, with the exception of R26 mono-methylation, post-transcriptional modifications of residues neighboring K27 is detrimental to ATXR5 activity. Overall, our results suggest that the deposition of H3.3 serves to prevent K27 mono-methylation and heterochromatin formation during DNA replication.

scholarly journals JARID2 and AEBP2 regulate PRC2 activity in the presence of H2A ubiquitination or other histone modifications

2020 ◽  
Author(s):  
Vignesh Kasinath ◽  
Curtis Beck ◽  
Paul Sauer ◽  
Simon Poepsel ◽  
Jennifer Kosmatka ◽  
...  
Keyword(s):  
Zinc Fingers ◽  
Regulation Of Gene Expression ◽  
Inhibitory Effect ◽  
Histone H2a ◽  
Set Domain ◽  
Methyltransferase Activity ◽  
Post Translational Modifications ◽  
Nucleosomal Dna ◽  
Polycomb Repressive Complexes ◽  
Stimulation Of

ABSTRACTThe Polycomb repressive complexes PRC1 and PRC2 functionally interact to coordinate cell type identity by the epigenetic regulation of gene expression. It has been proposed that PRC2 is recruited to genomic loci via the recognition of PRC1-mediated mono-ubiquitination of histone H2A at lysine 119 (H2AK119ub1), but the mechanism of this process remains poorly understood. Here, we report the cryo-EM structure of human PRC2 with cofactors JARID2 and AEBP2 bound to a nucleosome substrate containing H2AK119ub1. We find that JARID2 and AEBP2 each interact with one of the two ubiquitin molecules in the nucleosome. A ubiquitin-interaction motif (UIM) in JARID2 is sandwiched between ubiquitin and the histone H2A-H2B acidic patch. Simultaneously, the tandem zinc-fingers of AEBP2 interact with the second ubiquitin and the histone H2A-H2B surface on the opposite side of the nucleosome. JARID2 plays a dual role in the H2AK119ub1 dependent stimulation of PRC2 through interactions with both EED via its K116 trimethylation and with the H2AK119-ubiquitin. AEBP2, on the other hand, appears to primarily serve as a scaffold contributing to the interaction between PRC2 and the H2AK119ub1 nucleosome. Our structure also provides a detailed visualization of the EZH2-nucleosome interface, revealing a segment of EZH2 (named “bridge helix”) that is stabilized as it bridges the EZH2(SET) domain, the H3 tail and the nucleosomal DNA. In addition to the role played by AEBP2 and JARID2 in PRC2 regulation by H2AK119ub1 recognition, we also observe that the presence of these cofactors partially overcomes the inhibitory effect that H3K4- and H3K36-trimethylation have on core PRC2. Together, our results reveal the central role played by cofactors JARID2 and AEBP2 in orchestrating the crosstalk between histone post-translational modifications and PRC2 methyltransferase activity.

scholarly journals Bridging the gap between structure and kinetics of human SGLT1

2012 ◽  
Vol 302 (9) ◽  
pp. C1293-C1305 ◽  
Author(s):  
Monica Sala-Rabanal ◽  
Bruce A. Hirayama ◽  
Donald D. F. Loo ◽  
Vincent Chaptal ◽  
Jeff Abramson ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Binding Site ◽  
Sugar Transport ◽  
Gene Families ◽  
Structural Studies ◽  
Sugar Binding ◽  
Substituted Cysteine Accessibility ◽  
Biochemical Assays ◽  
Functional Consequences ◽  
Binding Residues

The Na+-glucose cotransporter hSGLT1 is a member of a class of membrane proteins that harness Na+ electrochemical gradients to drive uphill solute transport. Although hSGLT1 belongs to one gene family (SLC5), recent structural studies of bacterial Na+ cotransporters have shown that Na+ transporters in different gene families have the same structural fold. We have constructed homology models of hSGLT1 in two conformations, the inward-facing occluded (based on vSGLT) and the outward open conformations (based on Mhp1), mutated in turn each of the conserved gates and ligand binding residues, expressed the SGLT1 mutants in Xenopus oocytes, and determined the functional consequences using biophysical and biochemical assays. The results establish that mutating the ligand binding residues produces profound changes in the ligand affinity (the half-saturation concentration, K0.5); e.g., mutating sugar binding residues increases the glucose K0.5 by up to three orders of magnitude. Mutation of the external gate residues increases the Na+ to sugar transport stoichiometry, demonstrating that these residues are critical for efficient cotransport. The changes in phlorizin inhibition constant ( Ki) are proportional to the changes in sugar K0.5, except in the case of F101C, where phlorizin Ki increases by orders of magnitude without a change in glucose K0.5. We conclude that glucose and phlorizin occupy the same binding site and that F101 is involved in binding to the phloretin group of the inhibitor. Substituted-cysteine accessibility methods show that the cysteine residues at the position of the gates and sugar binding site are largely accessible only to external hydrophilic methanethiosulfonate reagents in the presence of external Na+, demonstrating that the external sugar (and phlorizin) binding vestibule is opened by the presence of external Na+ and closes after the binding of sugar and phlorizin. Overall, the present results provide a bridge between kinetics and structural studies of cotransporters.

scholarly journals SUV39 SET domains mediate crosstalk of heterochromatic histone marks

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Alessandro Stirpe ◽  
Nora Guidotti ◽  
Sarah J Northall ◽  
Sinan Kilic ◽  
Alexandre Hainard ◽  
...  
Keyword(s):  
Structure Function ◽  
Fission Yeast ◽  
Catalytic Domain ◽  
Constitutive Heterochromatin ◽  
Histone H3 ◽  
H3k9 Methylation ◽  
Set Domain ◽  
Heterochromatin Formation ◽  
Heterochromatin Silencing ◽  
Genomic Regions

The SUV39 class of methyltransferase enzymes deposits histone H3 lysine 9 di- and trimethylation (H3K9me2/3), the hallmark of constitutive heterochromatin. How these enzymes are regulated to mark specific genomic regions as heterochromatic is poorly understood. Clr4 is the sole H3K9me2/3 methyltransferase in the fission yeast Schizosaccharomyces pombe, and recent evidence suggests that ubiquitination of lysine 14 on histone H3 (H3K14ub) plays a key role in H3K9 methylation. However, the molecular mechanism of this regulation and its role in heterochromatin formation remain to be determined. Our structure-function approach shows that the H3K14ub substrate binds specifically and tightly to the catalytic domain of Clr4, and thereby stimulates the enzyme by over 250-fold. Mutations that disrupt this mechanism lead to a loss of H3K9me2/3 and abolish heterochromatin silencing similar to clr4 deletion. Comparison with mammalian SET domain proteins suggests that the Clr4 SET domain harbors a conserved sensor for H3K14ub, which mediates licensing of heterochromatin formation.

scholarly journals Human-specific microglial Siglec-11 transcript variant has the potential to affect polysialic acid-mediated brain functions at a distance

Glycobiology ◽  
2020 ◽  
Author(s):  
Masaya Hane ◽  
Dillon Y Chen ◽  
Ajit Varki
Keyword(s):  
Polysialic Acid ◽  
Innate Immune ◽  
Set Domain ◽  
Tissue Macrophage ◽  
Brain Functions ◽  
Functional Consequences ◽  
Molecular Patterns ◽  
The Brain ◽  
Better Than ◽  
Human Specific

Abstract CD33-related Siglecs are often found on innate immune cells and modulate their reactivity by recognition of sialic acid-based “self-associated molecular patterns” and signaling via intracellular tyrosine-based cytosolic motifs. Previous studies have shown that Siglec-11 specifically binds to the brain-enriched polysialic acid (polySia/PSA) and that its microglial expression in the brain is unique to humans. Furthermore, human microglial Siglec-11 exists as an alternate splice form missing the exon encoding the last (fifth) Ig-like C2-set domain of the extracellular portion of the protein, but little is known about the functional consequences of this variation. Here, we report that the recombinant soluble human microglial form of Siglec-11 (hSiglec-11(4D)-Fc) binds endogenous and immobilized polySia better than the tissue macrophage form (hSiglec-11(5D)-Fc) or the chimpanzee form (cSiglec-11(5D)-Fc). The Siglec-11 protein is also prone to aggregation, potentially influencing its ligand-binding ability. Additionally, Siglec-11 protein can be secreted in both intact and proteolytically cleaved forms. The microglial splice variant has reduced proteolytic release and enhanced incorporation into exosomes, a process that appears to be regulated by palmitoylation of cysteines in the cytosolic tail. Taken together, these data demonstrate that human brain specific microglial hSiglec-11(4D) has different molecular properties and can be released on exosomes and/or as proteolytic products, with the potential to affect polySia-mediated brain functions at a distance.

scholarly journals Global Replication-Independent Histone H4 Exchange in Budding Yeast

2006 ◽  
Vol 5 (10) ◽  
pp. 1780-1787 ◽  
Author(s):  
Jeffrey Linger ◽  
Jessica K. Tyler
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Dna Replication ◽  
Budding Yeast ◽  
Histone H3 ◽  
Eukaryotic Genome ◽  
Histone H4 ◽  
Histone Proteins ◽  
Dynamic Exchange ◽  
Histone Exchange

ABSTRACT The eukaryotic genome is packaged together with histone proteins into chromatin following DNA replication. Recent studies have shown that histones can also be assembled into chromatin independently of DNA replication and that this dynamic exchange of histones may be biased toward sites undergoing transcription. Here we show that epitope-tagged histone H4 can be incorporated into nucleosomes throughout the budding yeast (Saccharomyces cerevisiae) genome regardless of the phase of the cell cycle, the transcriptional status, or silencing of the region. Direct comparisons reveal that the amount of histone incorporation that occurs in G1-arrested cells is similar to that occurring in cells undergoing DNA replication. Additionally, we show that this histone incorporation is not dependent on the histone H3/H4 chaperones CAF-1, Asf1, and Hir1 individually. This study demonstrates that DNA replication and transcription are not necessary prerequisites for histone exchange in budding yeast, indicating that chromatin is more dynamic than previously thought.

scholarly journals The MMSET protein is a histone methyltransferase with characteristics of a transcriptional corepressor

Blood ◽  
2008 ◽  
Vol 111 (6) ◽  
pp. 3145-3154 ◽  
Author(s):  
Jotin Marango ◽  
Manabu Shimoyama ◽  
Hitomi Nishio ◽  
Julia A. Meyer ◽  
Dong-Joon Min ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Malignant Cell ◽  
Myeloma Cell ◽  
Set Domain ◽  
Methyltransferase Activity ◽  
Myeloma Cell Line ◽  
Chromatin Regulators ◽  
Igh Locus ◽  
Reporter Assays ◽  
Made In

Abstract MMSET, identified by its fusion to the IgH locus in t(4;14)-associated multiple myeloma, possesses domains found within chromatin regulators, including the SET domain. MMSET protein is overexpressed and highly associated with chromatin in myeloma cell lines carrying t(4;14). MMSET possesses methyltransferase activity for core histone H3 lysine 4 and histone 4 lysine 20, whereas MMSET made in cells only modified H4. Segments of MMSET fused to the Gal4 DNA binding domain repressed transcription of a chromatin-embedded Gal4 reporter gene. MMSET-mediated repression was associated with increased H4K20 methylation gene and loss of histone acetylation. Consistent with this repressive activity, MMSET could form a complex with HDAC1 and HDAC2, mSin3a, and the histone demethylase LSD1, suggesting that it is a component of corepressor complexes. Furthermore, MMSET coexpression enhances HDAC1- and HDAC2-mediated repression in transcriptional reporter assays. Finally, shRNA-mediated knockdown of MMSET compromised viability of a myeloma cell line, suggesting a biologic role for the protein in malignant cell growth. Collectively, these data suggest that, by acting directly as a modifier of chromatin as well as through binding of other chromatin-modifying enzymes, MMSET influences gene expression and potentially acts as a pathogenic agent in multiple myeloma.

scholarly journals Structural and Functional Consequences Induced by Post-Translational Modifications in α-Defensins

2011 ◽  
Vol 2011 ◽  
pp. 1-7 ◽  
Author(s):  
Enrico Balducci ◽  
Alessio Bonucci ◽  
Monica Picchianti ◽  
Rebecca Pogni ◽  
Eleonora Talluri
Keyword(s):  
Antimicrobial Peptide ◽  
Marked Change ◽  
Antibacterial Activities ◽  
Post Translational Modification ◽  
Post Translational Modifications ◽  
Adp Ribosylation ◽  
Functional Consequences ◽  
Adp Ribosyltransferase ◽  
Guanidino Group ◽  
Ribose Moiety

HNP-1 is an antimicrobial peptide that undergoes proteolytic cleavage to become a mature peptide. This process represents the mechanism commonly used by the cells to obtain a fully active antimicrobial peptide. In addition, it has been recently described that HNP-1 is recognized as substrate by the arginine-specific ADP-ribosyltransferase-1. Arginine-specific mono-ADP-ribosylation is an enzyme-catalyzed post-translational modification in which NAD+ serves as donor of the ADP-ribose moiety, which is transferred to the guanidino group of arginines in target proteins. While the arginine carries one positive charge, the ADP-ribose is negatively charged at the phosphate moieties at physiological pH. Therefore, the attachment of one or more ADP-ribose units results in a marked change of cationicity. ADP-ribosylation of HNP-1 drastically reduces its cytotoxic and antibacterial activities. While the chemotactic activity of HNP-1 remains unaltered, its ability to induce interleukin-8 production is enhanced. The arginine 14 of HNP-1 modified by the ADP-ribose is in some cases processed into ornithine, perhaps representing a different modality in the regulation of HNP-1 activities.

Dynamics of plant histone modifications in response to DNA damage

2012 ◽  
Vol 445 (3) ◽  
pp. 393-401 ◽  
Author(s):  
Georgina E. Drury ◽  
Adam A. Dowle ◽  
David A. Ashford ◽  
Wanda M. Waterworth ◽  
Jerry Thomas ◽  
...  
Keyword(s):  
Dna Damage ◽  
Histone Acetylation ◽  
Dna Damage Response ◽  
Genotoxic Stress ◽  
Immunoblot Analysis ◽  
X Rays ◽  
Histone Proteins ◽  
Post Translational Modifications ◽  
Damage Response ◽  
Dna Damage Signalling

DNA damage detection and repair take place in the context of chromatin, and histone proteins play important roles in these events. Post-translational modifications of histone proteins are involved in repair and DNA damage signalling processes in response to genotoxic stresses. In particular, acetylation of histones H3 and H4 plays an important role in the mammalian and yeast DNA damage response and survival under genotoxic stress. However, the role of post-translational modifications to histones during the plant DNA damage response is currently poorly understood. Several different acetylated H3 and H4 N-terminal peptides following X-ray treatment were identified using MS analysis of purified histones, revealing previously unseen patterns of histone acetylation in Arabidopsis. Immunoblot analysis revealed an increase in the relative abundance of the H3 acetylated N-terminus, and a global decrease in hyperacetylation of H4 in response to DNA damage induced by X-rays. Conversely, mutants in the key DNA damage signalling factor ATM (ATAXIA TELANGIECTASIA MUTATED) display increased histone acetylation upon irradiation, linking the DNA damage response with dynamic changes in histone modification in plants.

scholarly journals Post-Translational Modifications of FXR; Implications for Cholestasis and Obesity-Related Disorders

2021 ◽  
Vol 12 ◽  
Author(s):  
Monique D. Appelman ◽  
Suzanne W. van der Veen ◽  
Saskia W. C. van Mil
Keyword(s):  
Bile Acids ◽  
Posttranslational Modifications ◽  
Metabolic Disorders ◽  
Target Genes ◽  
Farnesoid X Receptor ◽  
Dietary Fats ◽  
Transcriptional Level ◽  
Alcoholic Fatty Liver ◽  
Post Translational Modifications ◽  
Functional Consequences

The Farnesoid X receptor (FXR) is a nuclear receptor which is activated by bile acids. Bile acids function in solubilization of dietary fats and vitamins in the intestine. In addition, bile acids have been increasingly recognized to act as signaling molecules involved in energy metabolism pathways, amongst others via activating FXR. Upon activation by bile acids, FXR controls the expression of many genes involved in bile acid, lipid, glucose and amino acid metabolism. An inability to properly use and store energy substrates may predispose to metabolic disorders, such as obesity, diabetes, cholestasis and non-alcoholic fatty liver disease. These diseases arise through a complex interplay between genetics, environment and nutrition. Due to its function in metabolism, FXR is an attractive treatment target for these disorders. The regulation of FXR expression and activity occurs both at the transcriptional and at the post-transcriptional level. It has been shown that FXR can be phosphorylated, SUMOylated and acetylated, amongst other modifications, and that these modifications have functional consequences for DNA and ligand binding, heterodimerization and subcellular localization of FXR. In addition, these post-translational modifications may selectively increase or decrease transcription of certain target genes. In this review, we provide an overview of the posttranslational modifications of FXR and discuss their potential involvement in cholestatic and metabolic disorders.

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