Data on morphology, large-scale chromatin configuration and the occurrence of proteins and rRNA in nucleolus-like bodies of fully-grown mouse oocytes in different fixatives

Here we provide data on accessibility of nucleolus-like bodies (NLBs) of fully-grown (GV) mouse oocytes to fluorescence in situ hybridization (FISH) probes and anti-nucleolar antibodies as well as on oocyte general morphology and large scale chromatin configuration, which relate to the research article “High-resolution microscopy of active ribosomal genes and key members of the rRNA processing machinery inside nucleolus-like bodies of fully-grown mouse oocytes” (Shishova et al., 2015 [1]). Experimental factors include: a cross-linking reagent formaldehyde and two denaturing fixatives, such as 70% ethanol and a mixture of absolute methanol and glacial acetic acid (3:1, v/v)

New insights into the induction of the heat shock proteins in baculovirus infected insect cells

AbstractEight members of the HSP/HSC70 family were identified in Spodoptera frugiperda Sf9 cells infected with Autographa californica multiple nucleopolyhedrovirus (AcMNPV) by 2D electrophoresis followed by mass spectrometry (MALDI/TOF) and a Mascot search. The family includes five HSP70s induced by AcMNPV-infection and three constitutive cognate HSC70s that remained abundant in infected cells. Confocal microscopy revealed dynamic changes in subcellular localization of HSP/HSC70s in the course of infection. At the early stages (4 to 10 hpi), a fraction of HSPs is localized in distinct speckles in cytoplasm. The speckles contained ubiquitinylated proteins suggesting that they may be aggresomes where proteins targeted by ubiquitin are sequestered or processed for proteolysis. S. frugiperda HSP90 was identified in the 2D gels by Western blotting. Its amount was unchanged during infection. A selective inhibitor of HSP90, 17-AAG, decreased the rate of viral DNA synthesis in infected cells suggesting a supportive role of HSP90 in virus replication

Reassembly of functional nucleoli following in situ unraveling by low-ionic-strength treatment of cultured mammalian cells.

In order to determine the most persistent components of the nucleolus that might serve as "core" nucleolar elements, we studied the reactivity of nucleoli in living mammalian cells subjected to hypotonic buffer saline followed by the incubation of the cells in an isotonic medium. To document as precisely as possible the fine structural changes which occurred, the cells were examined by video-enhanced optical microscopy, fluorescence confocal laser scanning microscopy, and electron microscopy combined with cytochemistry. Light microscopic autoradiography was used to demonstrate the transcriptional characteristics of the reassembled nucleoli. It was shown that all the major compartments of the intact nucleolus could be substantially affected by reduction of the osmolarity of the environmental media. The dynamic events of the nucleolar unraveling in low-salt buffers occurred in the following order: dispersion of the nucleolar pars granulosa, disassociation of the fibrillar complexes into discrete fibrillar centers (FCs) and the dense fibrillar component (DFC), and the almost complete unraveling of the DFC and FCs. At the terminal stages of nucleolar dispersion, the nuclear interior was mainly composed of a loose filamentous meshwork, and none of the typically discerned nucleolar constituents was recognized. Nevertheless, when hypotonically treated cells were returned to isotonic conditions, the nucleolar bodies rapidly began to reassemble. Within 1-2 h of cell incubation under isotonicity, the nucleoli not only became clearly visible, but also reconstituted to their initial size, shape, and position within the nucleus. The ultrastructure and functional activity of the reassembled nucleoli were also found to be fully comparable to those of the untreated controls. These data indicate that the architectural composition of the interphase nucleolus is strictly controlled by the cell. As far as could be determined, none of the usual substructures of the intact nucleolus that could be substituted by complete reassembly of the nucleolar bodies in normotonic conditions, including FCs and the DFC, remained clearly preserved in the terminal stage of nucleolar unraveling. We concluded that the integrity of the nucleolus was mainly preserved by the nuclear or nucleolar matrix system rather than by any other nucleolar structural domains

Genes Responsible for H2S Production and Metabolism Are Involved in Learning and Memory in Drosophila melanogaster

The gasotransmitter hydrogen sulfide (H2S) produced by the transsulfuration pathway (TSP) is an important biological mediator, involved in many physiological and pathological processes in multiple higher organisms, including humans. Cystathionine-β-synthase (CBS) and cystathionine-γ-lyase (CSE) enzymes play a central role in H2S production and metabolism. Here, we investigated the role of H2S in learning and memory processes by exploring several Drosophila melanogaster strains with single and double deletions of CBS and CSE developed by the CRISPR/Cas9 technique. We monitored the learning and memory parameters of these strains using the mating rejection courtship paradigm and demonstrated that the deletion of the CBS gene, which is expressed predominantly in the central nervous system, and double deletions completely block short- and long-term memory formation in fruit flies. On the other hand, the flies with CSE deletion preserve short- and long-term memory but fail to exhibit long-term memory retention. Transcriptome profiling of the heads of the males from the strains with deletions in Gene Ontology terms revealed a strong down-regulation of many genes involved in learning and memory, reproductive behavior, cognition, and the oxidation–reduction process in all strains with CBS deletion, indicating an important role of the hydrogen sulfide production in these vital processes

Wild type and mutant antenna and leg in flies in normal conditions and following irradiation.

<p>Simultaneous combination of mutations at both <i>ss</i> and <i>CG5017</i> loci increases sensitivity to even low doses of X-ray radiation (1 and 10 R), which is manifested as an increase in the mutant phenotype (photographs within the red frame).</p

Wild type and mutant leg and antenna phenotypes.

<p>Micrographs show the normal morphology of the tarsal structures of the wild type <i>Canton S</i>, <i>ss^a40ahm</i> and <i>milkah-1</i> flies. Antennal structures of the <i>milkah-1</i> mutant flies do not show any difference from the wild type. The distal segment of the antenna – the arista - of the <i>ss^a40ahm</i> flies presents a certain thickening of the proximal end. The arista of the <i>ss^a40ahmmilkah-1</i> hybrid flies suffered homeotic transformation into an unsegmented tarsus. The tarsal structures of the <i>ss^a40ahmmilkah-1</i> hybrids show altered segment fusion.</p