Chromatin nuclear

Martins, S., Eikvar, S., Fumkawa, K. and Collas, P. (2003) HA95 and LAP2 beta mediate a novel chromatin-nuclear envelope interaction implicated in initiation of DNA replication. J. Cell Biol. 160, 177-188. 

Global transcription of endogenous genes was also coupled to DNA replication. Aphidicolin-treated, 1-cell embryos assayed for BrUTP incorporation in G2 of the first cell cycle revealed a 35% decrease when compared to untreated control embryos (Aoki et al., 1997). Interestingly, the effect of aphidicolin on transcription by the male pronucleus was greater than that for the female pronucleus. While the basis for this difference is not known, it may be yet another manifestation of the differences in chromatin/nuclear structure between the two pronuclei. 

In parallel with the detailed study of glycoprotein assembly, new information has emerged about the subcellular localisation of carbohydrates. They are now recognised as components of the mitochondria, lysosomes, nuclear envelope, chromatin, nuclear matrix, ribosomes and extracellular matrix. 

The procedures reported here have been used to investigate the development of male pronuclei in cell-free systems of sea urchins and, to a lesser extent, surf clams. Pronuclear formation in vitro is a slower process than that in vivo. This property has been used advantageously to examine the steps of pronuclear formation. Each step can be easily manipulated, but the methods described here may require some adjustments for other organisms. Sea urchin and surf clam male pronuclei formed in vitro are virtually complete, with decondensed chromatin, nuclear envelopes, pores, and lamina. 

In this chapter we describe data favoring the idea that the altered DNA repair kinetics in alkylated cells treated with the poly(ADP-ribose) synthesis inhibitor ben-zatnide are the consequence of an inhibition of DNA repair at a limited number of sites in combination with an increase in the number of incisions. Both alterations could be explained by an involvement of poly(ADP-ribosyl)ation in the maintenance of chromatin structure in order to allow correct repair of the chromatin following alkylation. Preliminary studies analyzing the chromatin structure in ceDs cultivated in the presence of benzamide have indeed pointed to a participation of poly(ADP-ribosyl)ation in the organization of the chromatin, possibly in the chromatin/nuclear matrix interaction. 

The native form of chromatin in cells assumes a higher order stmcture called the 30-nm filament, which adopts a solenoidal stmcture where the 10-nm filament is arranged in a left-handed cod (Fig. 5). The negative supercoiling of the DNA is manifested by writhing the hehcal axis around the nucleosomes. Chromatin stmcture is an example of toroidal winding whereas eukaryotic chromosomes are linear, the chromatin stmctures, attached to a nuclear matrix, define separate closed-circular topological domains. 

The detachment of chromatin from the nuclear scaffold, leading to chromatin conden.sation. 

Nucleus The nucleus is separated from the cytosol by a double membrane, the nuclear envelope. The DNA is complexed with basic proteins (histones) to form chromatin fibers, the material from which chromosomes are made. A distinct RNA-rich region, the nucleolus, is the site of ribosome assembly. The nucleus is the repository of genetic information encoded in DNA and organized into chromosomes. During mitosis, the chromosomes are replicated and transmitted to the daughter cells. The genetic information of DNA is transcribed into RNA in the nucleus and passes into the cytosol where it is translated into protein by ribosomes. 

The antagonist-induced conformation of nuclear hormone receptors attracts co-repressors like Nco/SMRT (nuclear hormone receptor co-repressor/silencing mediator of retinoid and thyroid receptors) which further recruit other nuclear proteins with histone deacetylase activity. Their action leads to chromatin condensation, thus preventing the general transcription apparatus from binding to promoter regions. 

Beside coactivators so-called corepressors exist that are bound to transcription factors such as nuclear receptors and inhibit the initiation of transcription. These factors include the nuclear receptor corepressor (NCoR) and the silencing mediator of retinoic acid and thyroid hormone receptor (SMRT), which interact with nuclear receptors and serve as platforms for complexes containing histone deacetylases (HDACs). These enzymes cause the reversal of histone acetylation of histones leading to a tightening of chromatin and enhancing its inaccessibility for RNA polymerase containing complexes. 

Another morphological assay of apoptosis is done with acridine orange, a nuclear staining that reveals chromatin condensation under light and fluorescent microscope. 

The assembly of nucleosomes is mediated by one of several chromatin assembly factors facilitated by histone chaperones, proteins such as the anionic nuclear protein nucleoplasmin. As the nucleosome is assembled, histones are released from the histone chaperones. Nucleosomes appear to exhibit preference for certain regions on specific DNA molecules, but the basis for this nonrandom distribution, termed phasing, is not completely... 

Chromatin remodeling, transcription factor modification by various enzyme activities, and the communication between the nuclear receptors and the basal transcription apparatus are accomplished by protein-protein interactions with one or more of a class of coregulator molecules. The number of these coregulator molecules now exceeds 100, not counting species variations and splice variants. The first of these to be described was the CREB-binding protein, CBP. CBP, through an amino terminal domain, binds to phosphorylated serine 137 of CREB and mediates transactivation in response to cAMP. It thus is described as a coactivator. CBP and... 

Another large family of coregulator proteins remodel chromatin, modify other transcription factots, and bridge the nuclear receptors to the basal ttanscription apparatus. 

In an ideal stain, the cytoplasm of cysts and trophozoites is blue-green tinged with purple. Entamoeba coli cyst cytoplasm is often more purple than that of other species. Nuclear chromatin, chromatoid bodies, erythrocytes, and bacteria stain red or purplish red. Other ingested particles such as yeasts often stain green. Parasite eggs and larvae usually stain red. Inflammatory cells and tissue cells stain in a fashion similar to that of protozoa. Color reactions may vary from the above. 

Direct division of the chromatin bodies of E. coli has been demonstrated by Mason and Powelson in a series of remarkable phasecontrast photomicrographs. These observations were made on living bacteria. The nuclear areas in the dividing cells appeared to be as clearly defined as the areas in fixed, hydrolyzed, and stained cells. 

Figure 12.7 Colon fixed in buffered zinc formalin (Z-Fix, Anatech Ltd., Battle Creek, MI) for only 6h. Cellular detail is very prominent, nuclear chromatin is unusually well displayed and cytoplasmic staining is intensified. 

Figure 12.9 Salivary gland fixed in a glyoxal solution (Prefer, Anatech Ltd.). Membranes around mucus cells are unusually clear and nuclear chromatin is sharply defined. 

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