Supranucleosomal Organization of Chromatin Fibers in Nuclei ofDrosophilaS2 Cells

Abstract

Earlier, the interphase chromatin structures could not be visualized due to the stickiness of the nuclear material. We have reduced stickiness by the reversal of permeabilization allowing the isolation and microscopic imaging of interphase chromatin structures. By using a high resolution of synchronization, collecting 36 elutriation fractions, we show that major intermediates of chromatin condensation include: (a) decondensed veillike chromatin at the unset of the S phase (2.0-2.2 C-value), (b) polarization of veiled chromatin (2.2-2.6 C), (c) fibrous chromatin (2.6-3.0 C), chromatin bodies (3.0-3.3 C), early precondensed chromosomes (3.3-3.6). The compaction of Drosophila chromosomes did not reach that of the mammalian cells in the final stage of condensation (3.6-4.0 C). Drosophila chromosomes consist of smaller units called rodlets. Results demonstrate that nucleosomal chromatin ("beads on string") does not form a solenoid structure; rather, the topological arrangement consists of meandering and plectonemic loops.

Cell-cycle regulation of non-enzymatic functions of the Drosophila methyltransferase PR-Set7

“…To this end, PR-Set7 protein and histone H4K20me1 levels were analyzed through out the cell cycle in Drosophila S2 cells. Logarithmically growing S2 cells were separated according to cell size by centrifugal elutriation, allowing the isolation of cells in different cell-cycle phases with minimal physiological perturbation ( 22 ). The measure of DNA content by flow cytometry confirmed the differential cell-cycle distribution in each elutriated fraction (Figure 1A , left panels).…”

“…1.5 × 10 9 S2 cells were grown in complete Schneider's Drosophila medium at a density of 3.5 × 10 6 cells/ml, collected, washed in ice-cold PBS, resuspended in 200 ml of PBS and injected in the sample injection loop of a centrifugal elutriator composed of a large chamber (40 ml) with a counterbalance at the opposite side in a JE-5.0 rotor in an Avanti J-26XP centrifuge (Beckman Coulter) ( 22 ). After 30 min centrifugation at 3500 rpm, necessary for the equilibration of cells in the chamber, fractions were collected by decreasing the rotor speed every 30 sec by 100 rpm until the rotor speed reached 2200 rpm.…”

Chromatin Fiber Structure and Plectonemic Model of Chromosome Condensation inDrosophilaCells

“…We have addressed this problem by restoring the impermeability of the nuclear membrane by the reversal of permeabilization and securing its function, but allowing its reopening any time during the interphase in mammalian and in Drosophila cells (Banfalvi, 2006;Banfalvi et al, 2007). It has not escaped notice that the nucleosomal organization of chromatin serves as an experimental evidence for a possible mechanism of chromosome condensation at least in Drosophila cells.…”

Cytotoxic Endonucleases: New Targets for Prostate Cancer Chemotherapy

“…Studies of endonucleases associated with prostate cancer are very limited. Usually neoplastic transformation is associated with the decrease of endonuclease expression and activity in various cancers, thus making them "immortal" (Banfalvi et al, 2007;Basnakian et al, 1991;Wang et al, 2008). The most profound decrease of endonuclease activity was observed in malignant invasive prostate and breast cancer cells Wang et al, 2008).…”