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Query: EC:3.1.31.1 (micrococcal nuclease)
 2,818 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The biochemical role of poly(ADP-ribosyl)ation on internucleosomal DNA fragmentation associated with apoptosis was investigated in HL 60 human premyelocytic leukemia cells. It was found that UV light and chemotherapeutic drugs including adriamycin, mitomycin C, and cisplatin increased poly(ADP-ribosyl)ation of nuclear proteins, particularly histone H1. A poly(ADP-ribose) polymerase inhibitor, 3-aminobenzamide, prevented both internucleosomal DNA fragmentation and histone H1 poly(ADP-ribosyl)ation in cells treated with the apoptosis inducers. When nuclear chromatin was made accessible to the exogenous nuclease in a permeabilized cell system, chromatin of UV-treated cells was more susceptible to micrococcal nuclease than the chromatin of control cells. Suppression of histone H1 poly(ADP-ribosyl)ation by 3-aminobenzamide reduced the micrococcal nuclease digestibility of internucleosomal chromatin in UV-treated cells. These results suggest that the poly(ADP-ribosyl)ation of histone H1 correlates with the internucleosomal DNA fragmentation during apoptosis mediated by DNA damaging agents. This suggestion is supported by the finding that xeroderma pigmentosum cells which are defective in introducing incision at the site of DNA damage, failed to induce DNA fragmentation as well as histone H1 poly(ADP-ribosyl)ation after UV irradiation. We propose that poly(ADP-ribosyl)ation of histone H1 protein in the early stage of apoptosis facilitates internucleosomal DNA fragmentation by increasing the susceptibility of chromatin to cellular endonuclease.
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PMID:Poly(ADP-ribosyl)ation of histone H1 correlates with internucleosomal DNA fragmentation during apoptosis. 862 64

To investigate changes in chromatin organization associated with DNA replication during the first stages of development of the sea urchin Tetrapygus niger, we compared micrococcal nuclease (MNase) digestion patterns of chromatin from zygotes harvested during the first S phase and from unfertilized eggs. We observed that the majority of DNA fragments derived from MNase digested zygote nuclei were similar to or smaller than a mononucleosome, while those derived from unfertilized egg nuclei were larger (1,500 to 410 bp). This result indicates that in zygotes, where active DNA replication is occurring, the major chromatin fraction is represented as unfolded nucleosomes. In contrast, in unfertilized eggs chromatin appears to be organized into polynucleosomes. To determine if the unfolded structure of nucleosomes observed during S phase is related to the level of poly (ADP-ribosylation) of cleavage stage (CS) histone variants, zygotes were treated with 20 mM 3-Amino Benzamide (3 ABA) during the interval between 3 and 30 min post-insemination (p.i.). This treatment with 3 ABA decreases the poly (ADP-ribosylation) of CS histone variants and inhibits the first S phase in zygotes [Imschenetzky et al. (1991): J Cell Biochem 46:234-241; Imschenetzky et al. (1993): J Cell Biochem 51:198-205]. When the MNase digested patterns of chromatin from these 3 ABA treated and control zygotes were compared, we found that the unfolded structure of the nucleosomes remains unaltered by the inhibition of the poly(ADP-ribose) synthetase with 3 ABA. This result indicates that the unfolded nucleosomal structure, particular to the chromatin of S phase zygotes, is not contemporaneous to DNA replication and is independent of the normal level of poly(ADP-ribosylation) of CS histone variants.
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PMID:Sea urchin zygote chromatin exhibit an unfolded nucleosomal array during the first S phase. 890 10

Although the cellular response to chemical-induced stress is relatively well characterized, particularly the response to DNA damage, factors that govern the outcome of the stress response (cell survival or cell death) are less clearly defined. In this context, the mitogen-activated protein kinase (MAPK) family responds to a variety of physical and chemical stresses. The activation of MAPKs, especially the extracellular-regulated protein kinase subfamily, seems to play a causal role in death of renal proximal tubular epithelial cells (LLC-PK1) induced by reactive oxygen species (ROS). In this study, we show that extracellular signal receptor-activated kinase (ERK) activation may be coupled with LLC-PK1 cell death via changes in chromatin structure, which is mediated by increases in the phosphorylation of histone H3 (a post-translational modification required for both chromosome condensation and segregation during mitosis) and premature chromatin/chromosomal condensation, leading to cell death. In support of this view, 2,3,5-tris-(glutathione-S-yl)hydroquinone (TGHQ)-induced phosphorylation of histone H3 is accompanied by increases in chromatin condensation, as observed with the use of 4,6-diamidino-2-phenylindole-fluorescent staining, and by decreases in the sensitivity of chromatin to digestion by micrococcal nuclease. Changes in chromatin structure precede cell death. TGHQ-induced histone H3 phosphorylation and chromatin condensation are inhibited by PD098059, which selectively inhibits MAPK kinase, an upstream regulator of ERKs. Moreover, histone phosphorylation is modulated by poly(ADP-)ribosylation. Thus, the inhibition of poly(ADP-ribose)polymerase with 3-aminobenzamide prevents histone H3 phosphorylation and increases cell survival, suggesting that ADP-ribosylation and histone H3 phosphorylation are coupled in this model of ROS-induced DNA damage and cell death. The coupling of histone phosphorylation with ribosylation has not been previously demonstrated.
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PMID:Histone H3 phosphorylation is coupled to poly-(ADP-ribosylation) during reactive oxygen species-induced cell death in renal proximal tubular epithelial cells. 1145 28

SIR2-like proteins have been implicated in a wide range of cellular events including chromosome silencing, chromosome segregation, DNA recombination and the determination of life span. We report here the molecular and functional characterization of a SIR2-related protein from the protozoan parasite Trypanosoma brucei, which we termed TbSIR2RP1. This protein is a chromosome-associated NAD-dependent enzyme which, in contrast to other known proteins of this family, catalyses both ADP-ribosylation and deacetylation of histones, particulary H2A and H2B. Under- or overexpression of TbSIR2RP1 decreased or increased, respectively, cellular resistance to DNA damage. Treatment of trypanosomal nuclei with a DNA alkylating agent resulted in a significant increase in the level of histone ADP-ribosylation and a concomitant increase in chromatin sensitivity to micrococcal nuclease. Both of these responses correlated with the level of TbSIR2RP1 expression. We propose that histone modification by TbSIR2RP1 is involved in DNA repair.
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PMID:A chromosomal SIR2 homologue with both histone NAD-dependent ADP-ribosyltransferase and deacetylase activities is involved in DNA repair in Trypanosoma brucei. 1459 82

We have evaluated the influence of protein poly(ADP-ribosyl)ation in the relaxation of chromatin by exposing a rat liver polynucleosomal extract to micrococcal nuclease (MNase) digestion. The kinetic susceptibility of polynucleosomes to endonuclease digestion was determined as a function of the time of incubation as well as endonuclease concentration. To validate our assay, we also ran control experiments with protein-free calf thymus DNA as the opposite of polynucleosomal DNA. Rat liver chromatin was also incubated in the absence or presence of exogenously added 200 microM betaNAD(+), the poly(ADP-ribosyl)ation substrate, before MNase digestion. For incubations in the presence of betaNAD(+), the synthesis of polynucleosomal poly(ADP-ribose) was stopped with 1 mM benzamide. After addition of MNase, endonuclease digestion was blocked with EDTA to chelate the Mg(2+) ions needed for enzymatic activation, and the samples were subjected to electrophoresis through 1.5% agarose gels. As expected, a faster degradation of chromatin into oligonucleosomal DNA ladders was observed upon protein poly(ADP-ribosyl)ation when the chromatin extract was preincubated with 200 microM betaNAD(+). Thus, our results are consistent with the conclusion that the covalent poly(ADP-ribosyl)ation of polynucleosomal proteins favors a more "relaxed" or "open" structure, which renders chromatin more susceptible to MNase digestion.
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PMID:Polynucleosomal synthesis of poly(ADP-ribose) causes chromatin unfolding as determined by micrococcal nuclease digestion. 1565 42

Much is known about the distal DNA damage repair response. In particular, many of the enzymes and auxiliary proteins that participate in DNA repair have been characterized. In addition, knowledge of signaling pathways activated in response to DNA damage is increasing. In contrast, comparatively less is known of DNA damage-sensing molecules or of the specific alterations to chromatin structure recognized by such DNA damage sensors. Thus, precisely how chromatin structure is altered in response to DNA damage and how such alterations regulate DNA repair processes remain important unanswered questions. In vertebrates, phosphorylation of the histone variant H2A.X occurs rapidly after double-strand break formation, extends over megabase chromatin domains, and is required for stable accumulation of repair proteins at damage foci. We have shown that reactive oxygen species (ROS)-induced DNA single-strand breaks induce the incorporation of 32P specifically into histone H3. ADP-Ribosylation of histones may stimulate local chromatin relaxation to facilitate the repair process, and, indeed, histone ribosylation preceded DNA damage-induced histone H3 phosphorylation. However, H3 phosphorylation occurred concomitant with overall chromatin condensation, as revealed by decreased sensitivity of chromatin to digestion by micrococcal nuclease and by DAPI staining of nuclei. Inhibitors of the ERK and p38MAPK pathways and inhibition of poly(ADP-ribose) polymerase all reduced ROS-induced H3 phosphorylation, chromatin condensation, and cell death. Precisely how changes in the post-translational modification of histone H3 regulate the survival response remains unclear. Attempts to determine the precise site of histone H3 phosphorylation, putative histone H3 kinases, and histone H3 interacting proteins are underway.
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PMID:Ros-induced histone modifications and their role in cell survival and cell death. 1714


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