scholarly journals The Molecular Basis for Histone H4- and H2A-Specific Amino-Terminal Acetylation by NatD

Structure ◽  
2015 ◽  
Vol 23 (2) ◽  
pp. 332-341 ◽  
Author(s):  
Robert S. Magin ◽  
Glen P. Liszczak ◽  
Ronen Marmorstein
Keyword(s):  
Molecular Basis ◽  
Histone H4 ◽  
Amino Terminal

Islands of acetylated histone H4 in polytene chromosomes and their relationship to chromatin packaging and transcriptional activity

1990 ◽  
Vol 96 (2) ◽  
pp. 335-346
Author(s):  
B.M. Turner ◽  
L. Franchi ◽  
H. Wallace
Keyword(s):  
Polytene Chromosomes ◽  
Active Regions ◽  
Immunogold Electron Microscopy ◽  
Double Labelling ◽  
Histone H4 ◽  
Nucleosome Core ◽  
Amino Terminal ◽  
Experimental Systems ◽  
Lysine Residues ◽  
And Function

The four histones of the nucleosome core particle are all subject to enzyme-catalysed, post-translational acetylation at defined lysine residues in their amino-terminal domains. Much circumstantial evidence suggests a role for this process in modifying chromatin structure and function, but detailed mechanisms have not been defined. To facilitate studies on the functional significance of histone acetylation, we have prepared antibodies specific for the acetylated isoforms of histone H4. Because of the extreme evolutionary conservation of H4, these antisera can be applied to a wide variety of organisms and experimental systems. In the present study we have used polytene chromosomes from the salivary glands of larvae of the midge Chironomus to examine the distribution of acetylated H4 in interphase chromatin. By indirect immunofluorescence, antisera to acetylated H4 labeled the four Chironomus chromosomes with reproducible patterns of sharply defined, fluorescent bands. An antiserum to non-acetylated H4 gave a completely different, more-diffuse labelling pattern. Thus, there are defined regions, or islands, in the interphase genome that are enriched in acetylated H4. Double-labelling experiments with two antisera specific for H4 molecules acetylated at different sites, showed that each antiserum gave the same banding pattern. Immunolabelling patterns were not dependent on the pattern of phase-dense bands characteristic of these chromosomes; strongly labelled regions could correspond to phase-dense bands (i.e. condensed chromatin), to interbands or, frequently, to band-interband junctions. Immunogold electron microscopy confirmed the immunofluorescence results and showed further that regions of relatively high labelling could be either transcriptionally active or quiescent, as judged by the presence or absence of ribonucleoprotein particles. Two rapidly transcribed genes on chromosome 4 of Chironomus form characteristic ‘puffs’, the Balbiani rings BRb and BRc. The antiserum to non-acetylated H4 gave diffuse labelling throughout these puffs, demonstrating the continued presence of this histone in these transcriptionally active regions. Antisera to acetylated H4 strongly labelled the boundaries of BRb and BRc, and revealed clearly defined islands of increased H4 acetylation just within the expanded chromatin of the puffs. Labelling within the central region of each puff was much less intense. A similar pattern was observed in puffs on other chromosomes. Thus, increased H4 acetylation is not found throughout actively transcribed chromatin but occurs only at defined sites, possibly in the non-transcribed flanking regions. H4 acetylation is clearly not required for the passage of RNA polymerase through the nucleosome and we speculate that its role may be to facilitate the binding to DNA of polymerases and other proteins prior to the onset of transcription and possibly replication.

scholarly journals Initiating protease with modular domains interacts with β-glucan recognition protein to trigger innate immune response in insects

2015 ◽  
Vol 112 (45) ◽  
pp. 13856-13861 ◽  
Author(s):  
Daisuke Takahashi ◽  
Brandon L. Garcia ◽  
Michael R. Kanost
Keyword(s):  
Immune Responses ◽  
Molecular Basis ◽  
Molecular Mechanisms ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Dependent Manner ◽  
Calcium Dependent ◽  
Amino Terminal ◽  
Density Lipoprotein Receptor ◽  
Recognition Protein

The autoactivation of an initiating serine protease upon binding of pattern recognition proteins to pathogen surfaces is a crucial step in eliciting insect immune responses such as the activation of Toll and prophenoloxidase pathways. However, the molecular mechanisms responsible for autoactivation of the initiating protease remains poorly understood. Here, we investigated the molecular basis for the autoactivation of hemolymph protease 14 (HP14), an initiating protease in hemolymph of Manduca sexta, upon the binding of β-1,3-glucan by its recognition protein, βGRP2. Biochemical analysis using HP14 zymogen (proHP14), βGRP2, and the recombinant proteins as truncated forms showed that the amino-terminal modular low-density lipoprotein receptor class A (LA) domains within HP14 are required for proHP14 autoactivation that is stimulated by its interaction with βGRP2. Consistent with this result, recombinant LA domains inhibit the activation of proHP14 and prophenoloxidase, likely by competing with the interaction between βGRP2 and LA domains within proHP14. Using surface plasmon resonance, we demonstrated that immobilized LA domains directly interact with βGRP2 in a calcium-dependent manner and that high-affinity interaction requires the C-terminal glucanase-like domain of βGRP2. Importantly, the affinity of LA domains for βGRP2 increases nearly 100-fold in the presence of β-1,3-glucan. Taken together, these results present the first experimental evidence to our knowledge that LA domains of an insect modular protease and glucanase-like domains of a βGRP mediate their interaction, and that this binding is essential for the protease autoactivation. Thus, our study provides important insight into the molecular basis underlying the initiation of protease cascade in insect immune responses.

scholarly journals Molecular Basis of Secretin Docking to Its Intact Receptor Using Multiple Photolabile Probes Distributed throughout the Pharmacophore

2011 ◽  
Vol 286 (27) ◽  
pp. 23888-23899 ◽  
Author(s):  
Maoqing Dong ◽  
Polo C.-H. Lam ◽  
Delia I. Pinon ◽  
Keiko Hosohata ◽  
Andrew Orry ◽  
...  
Keyword(s):  
G Protein ◽  
Molecular Basis ◽  
Peptide Mapping ◽  
G Protein Coupled Receptors ◽  
Peptide Ligands ◽  
Amino Terminus ◽  
Amino Terminal ◽  
Binding Cleft ◽  
Mutant Receptors ◽  
G Protein Coupled

The molecular basis of ligand binding and activation of family B G protein-coupled receptors is not yet clear due to the lack of insight into the structure of intact receptors. Although NMR and crystal structures of amino-terminal domains of several family members support consistency in general structural motifs that include a peptide-binding cleft, there are variations in the details of docking of the carboxyl terminus of peptide ligands within this cleft, and there is no information about siting of the amino terminus of these peptides. There are also no empirical data to orient the receptor amino terminus relative to the core helical bundle domain. Here, we prepared a series of five new probes, incorporating photolabile moieties into positions 2, 15, 20, 24, and 25 of full agonist secretin analogues. Each bound specifically to the receptor and covalently labeled single distinct receptor residues. Peptide mapping of labeled wild-type and mutant receptors identified that the position 15, 20, and 25 probes labeled residues within the distal amino terminus of the receptor, whereas the position 24 probe labeled the amino terminus adjacent to TM1. Of note, the position 2 probe labeled a residue within the first extracellular loop of the receptor, a region not previously labeled, providing an important new constraint for docking the amino-terminal region of secretin to its receptor core. These additional experimentally derived constraints help to refine our understanding of the structure of the secretin-intact receptor complex and provide new insights into understanding the molecular mechanism for activation of family B G protein-coupled receptors.

scholarly journals Two Distinct Mechanisms of Chromatin Interaction by the Isw2 Chromatin Remodeling Complex In Vivo

2005 ◽  
Vol 25 (21) ◽  
pp. 9165-9174 ◽  
Author(s):  
Thomas G. Fazzio ◽  
Marnie E. Gelbart ◽  
Toshio Tsukiyama
Keyword(s):  
Chromatin Remodeling ◽  
Dna Binding Proteins ◽  
Chromatin Interaction ◽  
Chromatin Binding ◽  
Histone H4 ◽  
Amino Terminal ◽  
Histone Octamer ◽  
Chromatin Remodeling Complex ◽  
Specific Association

ABSTRACT We have previously shown that Saccharomyces cerevisiae Isw2 complex slides nucleosomes to remodel chromatin in vivo. Our data suggested a model in which Isw2 complex binds the histone octamer and DNA separately to generate the force necessary for nucleosome movement. Here we find that the histone H4 “basic patch” is the only portion of any amino-terminal histone tail required for both target-specific association of Isw2 complex with chromatin and chromatin remodeling in vivo, whereas it is dispensable for basal levels of chromatin binding. Similarly, we find that nonremodeled chromatin structure and integrity of Isw2 complex are required only for target-specific association of Isw2 with chromatin. These data demonstrate fundamental differences between the target-specific and basal modes of chromatin binding by Isw2 complex in vivo and suggest that only the former involves contributions from DNA, histone H4, and sequence-specific DNA binding proteins. We propose a model for target recognition and chromatin remodeling by Isw2 complex in vivo.

scholarly journals Molecular Basis for Strain Variation in the Saccharomyces cerevisiae Adhesin Flo11p

mSphere ◽  
2016 ◽  
Vol 1 (4) ◽  
Author(s):  
Subit Barua ◽  
Li Li ◽  
Peter N. Lipke ◽  
Anne M. Dranginis
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Surface ◽  
Molecular Basis ◽  
Phenotypic Variability ◽  
Surface Hydrophobicity ◽  
Strain Variation ◽  
Content Type ◽  
Amino Terminal ◽  
Important Means ◽  
Domain Peptide

ABSTRACT As a nonmotile organism, Saccharomyces cerevisiae employs the cell surface flocculin Flo11/Muc1 as an important means of adapting to environmental change. However, there is a great deal of strain variation in the expression of Flo11-dependent phenotypes, including flocculation. In this study, we investigated the molecular basis of this strain-specific phenotypic variability. Our data indicate that strain-specific differences in the level of flocculation result from significant sequence differences in the FLO11 alleles and do not depend on quantitative differences in FLO11 expression or on surface hydrophobicity. We further have shown that beads coated with amino-terminal domain peptide bind preferentially to homologous cells. These data show that variability in the structure of the Flo11 adhesion domain may thus be an important determinant of membership in microbial communities and hence may drive selection and evolution. FLO11 encodes a yeast cell wall flocculin that mediates a variety of adhesive phenotypes in Saccharomyces cerevisiae. Flo11p is implicated in many developmental processes, including flocculation, formation of pseudohyphae, agar invasion, and formation of microbial mats and biofilms. However, Flo11p mediates different processes in different yeast strains. To investigate the mechanisms by which FLO11 determines these differences in colony morphology, flocculation, and invasion, we studied gene structure, function, and expression levels. Nonflocculent Saccharomyces cerevisiae Σ1278b cells exhibited significantly higher FLO11 mRNA expression, especially in the stationary phase, than highly flocculent S. cerevisiae var. diastaticus. The two strains varied in cell surface hydrophobicity, and Flo11p contributed significantly to surface hydrophobicity in S. cerevisiae var. diastaticus but not in strain Σ1278b. Sequencing of the FLO11 gene in S. cerevisiae var. diastaticus revealed strain-specific differences, including a 15-amino-acid insertion in the adhesion domain. Flo11p adhesion domains from strain Σ1278b and S. cerevisiae var. diastaticus were expressed and used to coat magnetic beads. The adhesion domain from each strain bound preferentially to homologous cells, and the preferences were independent of the cells in which the adhesion domains were produced. These results are consistent with the idea that strain-specific variations in the amino acid sequences in the adhesion domains cause different Flo11p flocculation activities. The results also imply that strain-specific differences in expression levels, posttranslational modifications, and allelic differences outside the adhesion domains have little effect on flocculation. IMPORTANCE As a nonmotile organism, Saccharomyces cerevisiae employs the cell surface flocculin Flo11/Muc1 as an important means of adapting to environmental change. However, there is a great deal of strain variation in the expression of Flo11-dependent phenotypes, including flocculation. In this study, we investigated the molecular basis of this strain-specific phenotypic variability. Our data indicate that strain-specific differences in the level of flocculation result from significant sequence differences in the FLO11 alleles and do not depend on quantitative differences in FLO11 expression or on surface hydrophobicity. We further have shown that beads coated with amino-terminal domain peptide bind preferentially to homologous cells. These data show that variability in the structure of the Flo11 adhesion domain may thus be an important determinant of membership in microbial communities and hence may drive selection and evolution.

scholarly journals Insect-specific viruses regulate vector competence in Aedes aegypti mosquitoes via expression of histone H4

2021 ◽  
Author(s):  
Roenick P. Olmo ◽  
Yaovi Mathias H. Todjro ◽  
Eric R. G. R. Aguiar ◽  
Joao Paulo P. de Almeida ◽  
Juliana N. Armache ◽  
...  
Keyword(s):  
Aedes Aegypti ◽  
Aedes Albopictus ◽  
Molecular Basis ◽  
Vector Competence ◽  
Spatiotemporal Analysis ◽  
Host Factor ◽  
Mosquito Vectors ◽  
Histone H4 ◽  
African Cities ◽  
Aedes Mosquitoes

Aedes aegypti and Aedes albopictus are major mosquito vectors for arthropod-borne viruses (arboviruses) such as dengue (DENV) and Zika (ZIKV) viruses. Mosquitoes also carry insect-specific viruses (ISVs) that may affect the transmission of arboviruses. Here, we analyzed the global virome in urban Aedes mosquitoes and observed that two insect-specific viruses, Phasi Charoen-like virus (PCLV) and Humaita Tubiacanga virus (HTV), were the most prevalent in A. aegypti worldwide except for African cities, where transmission of arboviruses is low. Spatiotemporal analysis revealed that presence of HTV and PCLV led to a 200% increase in the chances of having DENV in wild mosquitoes. In the laboratory, we showed that HTV and PCLV prevented downregulation of histone H4, a previously unrecognized proviral host factor, and rendered mosquitoes more susceptible to DENV and ZIKV. Altogether, our data reveals a molecular basis for the regulation of A. aegypti vector competence by highly prevalent ISVs that may impact how we analyze the risk of arbovirus outbreaks.

scholarly journals Novel bisubstrate inhibitors for protein N-terminal acetyltransferase D

2021 ◽  
Author(s):  
Youchao Deng ◽  
Sunbin Deng ◽  
Yi-Hsun Ho ◽  
Sarah M. Gardner ◽  
Zhi Huang ◽  
...  
Keyword(s):  
High Resolution ◽  
Crystal Structures ◽  
Molecular Basis ◽  
Histone H4 ◽  
Peptide Substrates ◽  
Inhibitor Development ◽  
N Terminus ◽  
Acetyl Transfer ◽  
Bisubstrate Inhibitors ◽  
Important Enzyme

ABSTRACTProtein N-terminal acetyltransferase D (NatD, NAA40, Nat4) that specifically acetylates the N-terminus of histone H4 and H2A has been implicated in various diseases, but no inhibitor has been reported for this important enzyme. Based on the acetyl transfer mechanism of NatD, we designed and prepared a series of highly potent NatD bisubstrate inhibitors by covalently linking coenzyme A to different peptide substrates via an acetyl or propionyl spacer. The most potent bisubstrate inhibitor displayed a Ki of 170 ± 16 pM. We also demonstrated that these inhibitors are highly specific towards NatD, displaying 10,000-fold selectivity over other closely-related acetyltransferases. High resolution crystal structures of NatD bound to two of these inhibitors revealed the molecular basis for their selectivity and inhibition mechanisms, providing a rational path for future inhibitor development.

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