Dynamics of histone acetylation in vivo. A function for acetylation turnover?

2002 ◽  
Vol 80 (3) ◽  
pp. 363-378 ◽  
Author(s):  
Jakob H Waterborg
Keyword(s):  
Histone Acetylation ◽  
Gene Transcription ◽  
Turnover Rates ◽  
Nucleosome Remodeling ◽  
Amino Terminal ◽  
Rapid Changes ◽  
Core Histones ◽  
Reversible Modification ◽  
Amino Terminal Domain

Histone acetylation, discovered more than 40 years ago, is a reversible modification of lysines within the amino-terminal domain of core histones. Amino-terminal histone domains contribute to the compaction of genes into repressed chromatin fibers. It is thought that their acetylation causes localized relaxation of chromatin as a necessary but not sufficient condition for processes that repackage DNA such as transcription, replication, repair, recombination, and sperm formation. While increased histone acetylation enhances gene transcription and loss of acetylation represses and silences genes, the function of the rapid continuous or repetitive acetylation and deacetylation reactions with half-lives of just a few minutes remains unknown. Thirty years of in vivo measurements of acetylation turnover and rates of change in histone modification levels have been reviewed to identify common chromatin characteristics measured by distinct protocols. It has now become possible to look across a wider spectrum of organisms than ever before and identify common features. The rapid turnover rates in transcriptionally active and competent chromatin are one such feature. While ubiquitously observed, we still do not know whether turnover itself is linked to chromatin transcription beyond its contribution to rapid changes towards hyper- or hypoacetylation of nucleosomes. However, recent experiments suggest that turnover may be linked directly to steps in gene transcription, interacting with nucleosome remodeling complexes.Key words: histone, acetylation, turnover, chromatin, transcription.

scholarly journals Assembly of amino-terminally deleted desmin in vimentin-free cells.

1990 ◽  
Vol 111 (5) ◽  
pp. 1971-1985 ◽  
Author(s):  
J M Raats ◽  
F R Pieper ◽  
W T Vree Egberts ◽  
K N Verrijp ◽  
F C Ramaekers ◽  
...  
Keyword(s):  
Amino Acids ◽  
Hela Cells ◽  
Filament Formation ◽  
Mutant Proteins ◽  
Amino Terminal ◽  
Double Label ◽  
Amino Terminal Domain ◽  
Mcf 7 ◽  
Vimentin Filaments

To study the role of the amino-terminal domain of the desmin subunit in intermediate filament (IF) formation, several deletions in the sequence encoding this domain were made. The deleted hamster desmin genes were fused to the RSV promoter. Expression of such constructs in vimentin-free MCF-7 cells as well as in vimentin-containing HeLa cells, resulted in the synthesis of mutant proteins of the expected size. Single- and double-label immunofluorescence assays of transfected cells showed that in the absence of vimentin, desmin subunits missing amino acids 4-13 are still capable of filament formation, although in addition to filaments large numbers of desmin dots are present. Mutant desmin subunits missing larger portions of their amino terminus cannot form filaments on their own. It may be concluded that the amino-terminal region comprising amino acids 7-17 contains residues indispensable for desmin filament formation in vivo. Furthermore it was shown that the endogenous vimentin IF network in HeLa cells masks the effects of mutant desmin on IF assembly. Intact and mutant desmin colocalized completely with endogenous vimentin in HeLa cells. Surprisingly, in these cells endogenous keratin also seemed to colocalize with endogenous vimentin, even if the endogenous vimentin filaments were disturbed after expression of some of the mutant desmin proteins. In MCF-7 cells some overlap between endogenous keratin and intact exogenous desmin filaments was also observed, but mutant desmin proteins did not affect the keratin IF structures. In the absence of vimentin networks (MCF-7 cells), the initiation of desmin filament formation seems to start on the preexisting keratin filaments. However, in the presence of vimentin (HeLa cells) a gradual integration of desmin in the preexisting vimentin filaments apparently takes place.

scholarly journals The amino-terminal domain of pyrrolysyl-tRNA synthetase is dispensable in vitro but required for in vivo activity

FEBS Letters ◽  
2007 ◽  
Vol 581 (17) ◽  
pp. 3197-3203 ◽  
Author(s):  
Stephanie Herring ◽  
Alexandre Ambrogelly ◽  
Sarath Gundllapalli ◽  
Patrick O'Donoghue ◽  
Carla R. Polycarpo ◽  
...  
Keyword(s):  
Trna Synthetase ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Amino Terminal Domain

scholarly journals Dynamics of Nuclear Pore Distribution in Nucleoporin Mutant Yeast Cells

1997 ◽  
Vol 136 (4) ◽  
pp. 747-759 ◽  
Author(s):  
Naïma Belgareh ◽  
Valérie Doye
Keyword(s):  
Fusion Proteins ◽  
Fluorescent Protein ◽  
Pore Distribution ◽  
Yeast Cells ◽  
Nuclear Pore ◽  
Amino Terminal ◽  
Dividing Cells ◽  
Green Fluorescent ◽  
Amino Terminal Domain

To follow the dynamics of nuclear pore distribution in living yeast cells, we have generated fusion proteins between the green fluorescent protein (GFP) and the yeast nucleoporins Nup49p and Nup133p. In nup133− dividing cells that display a constitutive nuclear pore clustering, in vivo analysis of GFP-Nup49p localization revealed changes in the distribution of nuclear pore complex (NPC) clusters. Furthermore, upon induction of Nup133p expression in a GAL-nup133 strain, a progressive fragmentation of the NPC aggregates was observed that in turn led to a wild-type nuclear pore distribution. To try to uncouple Nup133p- induced NPC redistribution from successive nuclear divisions and nuclear pore biogenesis, we devised an assay based on the formation of heterokaryons between nup133− mutants and cells either expressing or overexpressing Nup133p. Under these conditions, the use of GFP-Nup133p and GFP-Nup49p fusion proteins revealed that Nup133p can be rapidly targeted to the clustered nuclear pores, where its amino-terminal domain is required to promote the redistribution of preexisting NPCs.

scholarly journals The CBP Bromodomain and Nucleosome Targeting Are Required for Zta-Directed Nucleosome Acetylation and Transcription Activation

2003 ◽  
Vol 23 (8) ◽  
pp. 2633-2644 ◽  
Author(s):  
Zhong Deng ◽  
Chi-Ju Chen ◽  
Michaela Chamberlin ◽  
Fang Lu ◽  
Gerd A. Blobel ◽  
...  
Keyword(s):  
Complex Formation ◽  
Histone Acetylation ◽  
Protein Sequencing ◽  
Creb Binding Protein ◽  
Barr Virus ◽  
Amino Terminal ◽  
Viral Promoters ◽  
Epstein Barr

ABSTRACT The Epstein-Barr virus (EBV)-encoded lytic activator Zta is a bZIP protein that can stimulate nucleosomal histone acetyltransferase (HAT) activity of the CREB binding protein (CBP) in vitro. We now show that deletion of the CBP bromo- and C/H3 domains eliminates stimulation of nucleosomal HAT activity in vitro and transcriptional coactivation by Zta in transfected cells. In contrast, acetylation of free histones was not affected by the addition of Zta or by deletions in the bromo or C/H3 domain of CBP. Zta stimulated acetylation of oligonucleosomes assembled on supercoiled DNA and dinucleosomes assembled on linear DNA, but Zta-stimulated acetylation was significantly reduced for mononucleosomes. Western blotting and amino-terminal protein sequencing indicated that all lysine residues in the H3 and H4 amino-terminal tails were acetylated by CBP and enhanced by the addition of Zta. Histone acetylation was also dependent upon the Zta basic DNA binding domain, which could not be substituted with the homologous basic region of c-Fos, indicating specificity in the bZIP domain nucleosome binding function. Finally, we show that Zta and CBP colocalize to viral immediate-early promoters in vivo and that overexpression of Zta leads to a robust increase in H3 and H4 acetylation at various regions of the EBV genome in vivo. Furthermore, deletion of the CBP bromodomain reduced stable CBP-Zta complex formation and histone acetylation at Zta-responsive viral promoters in vivo. These results suggest that activator- and bromodomain-dependent targeting to oligonucleosomal chromatin is required for stable promoter-bound complex formation and transcription activity.

Kinetic analysis of histone acetylation turnover and Trichostatin A induced hyper- and hypoacetylation in alfalfa

2002 ◽  
Vol 80 (3) ◽  
pp. 279-293 ◽  
Author(s):  
Jakob H Waterborg ◽  
Tamás Kapros
Keyword(s):  
Gene Expression ◽  
Steady State ◽  
Histone Acetylation ◽  
Histone Deacetylases ◽  
Trichostatin A ◽  
Similar Rate ◽  
Lysine Methylation ◽  
Histone H2a ◽  
Turnover Rates

Dynamic histone acetylation is a characteristic of chromatin transcription. The first estimates for the rate of acetylation turnover of plants are reported, measured in alfalfa cells by pulse, pulse-chase, and steady-state acetylation labeling. Acetylation turnover half-lives of about 0.5 h were observed by all methods used for histones H3, H4, and H2B. This is consistent with the rate at which changes in gene expression occur in plants. Treatment with histone deacetylase inhibitor Trichostatin A (TSA) induced hyperacetylation at a similar rate. Replacement histone variant H3.2, preferentially localized in highly acetylated chromatin, displayed faster acetyl turnover. Histone H2A with a low level of acetylation was not subject to rapid turnover or hyperacetylation. Patterns of acetate labeling revealed fundamental differences between histone H3 versus histones H4 and H2B. In H3, acetylation of all molecules, limited by lysine methylation, had similar rates, independent of the level of lysine acetylation. Acetylation of histones H4 and H2B was seen in only a fraction of all molecules and involved multiacetylation. Acetylation turnover rates increased from mono- to penta- and hexaacetylated forms, respectively. TSA was an effective inhibitor of alfalfa histone deacetylases in vivo and caused a doubling in steady-state acetylation levels by 4–6 h after addition. However, hyperacetylation was transient due to loss of TSA inhibition. TSA-induced overexpression of cellular deacetylase activity produced hypoacetylation by 18 h treatment with enhanced acetate turnover labeling of alfalfa histones. Thus, application of TSA to change gene expression in vivo in plants may have unexpected consequences.

Recombinant Polypeptides Derived from the Fibrin Binding Domain of Fibronectin Are Potential Agents for the Imaging of Blood Clots

1997 ◽  
Vol 77 (04) ◽  
pp. 796-803 ◽  
Author(s):  
N Ezov ◽  
A Nimrod ◽  
B Parizada ◽  
M M Werber ◽  
A Goldlust ◽  
...  
Keyword(s):  
Thrombus Formation ◽  
Recombinant Polypeptide ◽  
Covalent Linkage ◽  
Steel Coil ◽  
Amino Terminal ◽  
Imaging Tool ◽  
Recombinant Polypeptides ◽  
Amino Terminal Domain

SummaryThrombus formation in the circulation is accompanied by covalent linkage of fibronectin (FN) through transglutamination of glutamine no. 3 in the fibrin binding amino terminal domain (FBD) of FN. We have exploited this phenomenon for thrombus detection by the employment of radioactively-labelled recombinant polypeptide molecules derived from the 5-finger FBD of human FN. Three recombinant FBD polypeptides, 12 kDa (“2 fingers”), 18.5 kDa (“3 fingers”) and 31 kDa FBD (“5 fingers”), were prepared and compared to native FN-derived 31 kDa-FBD with respect to their ability to attach to fibrin clots in vitro and in vivo. The accessibility of Gln-3 in these molecules was demonstrated by the incorporation of stoichiometric amounts of 14C- putrescine in the presence of plasma transglutaminase. Competitive binding experiments to fibrin have indicated that, although the binding affinities of the FBD molecules are lower than that of FN, substantial covalent linkage was obtained in the presence of transglutaminase, and even in the presence of excess FN or heparin. The biological clearance rates of radioactively labelled FBD molecules in rats and rabbits were much higher than those of FN and fibrinogen, thus indicating their potential advantage for use as a diagnostic imaging tool. Of the three molecules, the 12 kDa FBD exhibited the highest rate of clearance. The potential of the 12 kDa and 31 kDa FBDs as imaging agents was examined in a stainless steel coil-induced thrombus model in rats and in a jugular vein thrombus model in rabbits, using either [125I] or [111ln]-labelled materials. At 24 h, clot-to-blood ratios ranged between 10 and 22 for [125I]-12 kDa FBD and 40 and 60 for [luIn]-12 kDa FBD. In the rat model, heparin did not inhibit the uptake of FBD. Taken together, the results indicate that recombinant 12 kDa FBD is a good candidate for the diagnosis of venous thrombosis.

scholarly journals 7B2 facilitates the maturation of proPC2 in neuroendocrine cells and is required for the expression of enzymatic activity.

1995 ◽  
Vol 129 (6) ◽  
pp. 1641-1650 ◽  
Author(s):  
X Zhu ◽  
I Lindberg
Keyword(s):  
Enzymatic Activity ◽  
Secretory Protein ◽  
Neuroendocrine Cells ◽  
Parent Cell ◽  
Intermediate Lobe ◽  
Amino Terminal ◽  
Highly Correlated ◽  
Carboxy Terminal ◽  
Amino Terminal Domain

The prohormone convertase PC2, which is thought to mediate the proteolytic conversion of many peptide hormones, has recently been shown to interact with the neuroendocrine-specific polypeptide 7B2 in Xenopus intermediate lobe (Braks, J. A. M., and G. J. M. Martens. Cell. 78:263. 1994). In the present work we have stably transfected neuroendocrine cell lines with rat 7B2 constructs and found that overexpression of 27 kD 7B2 greatly facilitates the kinetics of maturation of proPC2, both in AtT-20/PC2 cells and in Rin5f cells. The half-life of conversion of proPC2 was reduced from 2.7 to 1.7 h in AtT-20/PC2 cells stably transfected with 27 kD 7B2 cDNA. The previously proposed "chaperone" domain was not sufficient for this facilitation event; however, a construct corresponding to the 21-kD 7B2 protein (which represents the naturally occurring maturation product) functioned well. A 7B2 construct in which maturation of 27 kD 7B2 to its 21-kD form was blocked was unable to facilitate maturation of proPC2. To correlate effects on PC2 maturation with the actual generation of PC2 enzymatic activity, a similar transfection of 21 kD 7B2 was performed using CHO cells previously amplified for the expression of proPC2. Enzymatic activity cleaving the fluorogenic substrate Cbz-Arg-Ser-Lys-Arg-AMC was highly correlated with the expression of immunoreactive 21 kD 7B2 in the conditioned medium; medium obtained from the parent cell line was completely inactive. Enzymatic activity was identified as PC2 on the basis of inhibition by the carboxy-terminal peptide of 7B2, which has previously been shown to represent a potent and specific PC2 inhibitor. Taken together, our in vivo results indicate that the interesting secretory protein 7B2 is a bifunctional molecule with an amino-terminal domain involved in proPC2 transport as well as activation.

Epigenetic effects of dietary butyrate on hepatic histone acetylation and enzymes of biotransformation in chicken

2013 ◽  
Vol 61 (4) ◽  
pp. 477-490 ◽  
Author(s):  
Gábor Mátis ◽  
Zsuzsanna Neogrády ◽  
György Csikó ◽  
Péter Gálfi ◽  
Hedvig Fébel ◽  
...  
Keyword(s):  
Histone Acetylation ◽  
Broiler Chickens ◽  
Cell Function ◽  
Enzyme Assays ◽  
Histone H2a ◽  
Epigenetic Effects ◽  
Acetylation State ◽  
Core Histones ◽  
Microsomal Cytochrome P450

The aim of the study was to investigate the in vivo epigenetic influences of dietary butyrate supplementation on the acetylation state of core histones and the activity of drug-metabolising microsomal cytochrome P450 (CYP) enzymes in the liver of broiler chickens in the starter period. One-day-old Ross 308 broilers were fed a starter diet without or with sodium butyrate (1.5 g/kg feed) for 21 days. After slaughtering, nucleus and microsome fractions were isolated from the exsanguinated liver by multi-step differential centrifugation. Histone acetylation level was detected from hepatocyte nuclei by Western blotting, while microsomal CYP activity was examined by specific enzyme assays. Hyperacetylation of hepatic histone H2A at lysine 5 was observed after butyrate supplementation, providing modifications in the epigenetic regulation of cell function. No significant changes could be found in the acetylation state of the other core histones at the acetylation sites examined. Furthermore, butyrate did not cause any changes in the drugmetabolising activity of hepatic microsomal CYP2H and CYP3A37 enzymes, which are mainly involved in the biotransformation of most xenobiotics in chicken. These data indicate that supplementation of the diet with butyrate probably does not have any pharmacokinetic interactions with simultaneously applied xenobiotics.

scholarly journals Regulation of the renal Na+-Cl− cotransporter by phosphorylation and ubiquitylation

2012 ◽  
Vol 303 (12) ◽  
pp. F1573-F1583 ◽  
Author(s):  
Gerardo Gamba
Keyword(s):  
Current Knowledge ◽  
Protein Activity ◽  
Molecular Physiology ◽  
In Vivo Models ◽  
Major Mechanism ◽  
Amino Terminal ◽  
Arterial Blood ◽  
Major Interest ◽  
Amino Terminal Domain

The activity of the renal thiazide-sensitive NaCl cotransporter (NCC) in the distal convoluted tubule plays a key role in defining arterial blood pressure levels. Increased or decreased activity of the NCC is associated with arterial hypertension or hypotension, respectively. Thus it is of major interest to understand the activity of NCC using in vivo models. Phosphorylation of certain residues of the amino-terminal domain of NCC has been shown to be associated with its activation. The development of phospho-specific antibodies against these sites provides a powerful tool that is helping to increase our understanding of the molecular physiology of NCC. Additionally, NCC expression in the plasma membrane is modulated by ubiquitylation, which represents another major mechanism for regulating protein activity. This work presents a review of our current knowledge of the regulation of NCC activity by phosphorylation and ubiquitylation.

Sign in / Sign up

Export Citation Format

Share Document