Fractionation and characterization of euchromatin isolated from mouse ascites sarcoma cells

Euchromatin specimen prepared by the usual method formed large clumps and had various shapes under electron microscopy. A method of separation of the euchromatin specimen into chromatin fractions having relatively homogeneous form was examined and partial characterization of these fractions was carried out. The heavy euchromatin fraction was a large network of thin fibrils (about 100 A in diameter) and various thick fibers. The intermediate euchromatin fraction consisted of relatively homogeneous networks of thick knobby fibers (about 250 A in diameter). The light euchromatin fraction had metworks of thick fibers. These chromatin fractions were quantitatively prepared from sonicated nuclei of mouse ascites sarcoma cells. Twenty-one or twenty-two bands of non-histone proteins besides histones were detected in these chromatin fractions by SDS-polyacrylamide gel electrophoresis. There were significant differences in the electrophoretic patterns of non-histone proteins among these chromatin fractions.</p

Syncytia formation of human transformed cell lines by simian sarcoma virus type I (SSV-I/SSAV-I).

Human cells derived from malignant tumors (HeLa, HEp-2 and KB) and human cells transformed by tumor viruses (KCand RSb) formed syncytia by simian sarcoma virus type I (SSV-I/SSAV-I), but human diploid or non-transformed cells (WI-38, HEL and HEC) did not.</p

Effects of some polyamines, polyanions and antitumor drugs on replicative DNA synthesis and unscheduled DNA synthesis in vitro.

The effects of various compounds on replicative DNA synthesis in permeable mouse ascites sarcoma cells and on unscheduled DNA synthesis in permeable cells or in isolated rat liver nuclei were studied. Polyamines such as spermidine, putrescine and cadaverine inhibited replicative DNA synthesis. Unscheduled DNA synthesis was inhibited by spermidine and cadaverine, but slightly stimulated by putrescine at low concentrations. Aurintricarboxylic acid, a low molecular weight polyanion, inhibited both replicative DNA synthesis and unscheduled DNA synthesis. Replicative DNA synthesis was inhibited by heparin, a high molecular weight polyanion, whereas unscheduled DNA synthesis was stimulated at low heparin concentrations. Antitumor drugs such as daunomycin, neocarzinostatin and bleomycin inhibited replicative DNA synthesis. Unscheduled DNA synthesis was inhibited by daunomycin, slightly induced by neocarzinostatin and highly induced by bleomycin. The present system was thought to be useful for studying the separate effects of various drugs on either replicative DNA synthesis or unscheduled DNA synthesis in vitro.</p

Structural Basis for Variant-Specific Neuroligin-Binding by α-Neurexin

Neurexins (Nrxs) are presynaptic membrane proteins with a single membrane-spanning domain that mediate asymmetric trans-synaptic cell adhesion by binding to their postsynaptic receptor neuroligins. α-Nrx has a large extracellular region comprised of multiple copies of laminin, neurexin, sex-hormone-binding globulin (LNS) domains and epidermal growth factor (EGF) modules, while that of β-Nrx has but a single LNS domain. It has long been known that the larger α-Nrx and the shorter β-Nrx show distinct binding behaviors toward different isoforms/variants of neuroligins, although the underlying mechanism has yet to be elucidated. Here, we describe the crystal structure of a fragment corresponding to the C-terminal one-third of the Nrx1α ectodomain, consisting of LNS5-EGF3-LNS6. The 2.3 Å-resolution structure revealed the presence of a domain configuration that was rigidified by inter-domain contacts, as opposed to the more common flexible “beads-on-a-string” arrangement. Although the neuroligin-binding site on the LNS6 domain was completely exposed, the location of the α-Nrx specific LNS5-EGF3 segment proved incompatible with the loop segment inserted in the B+ neuroligin variant, which explains the variant-specific neuroligin recognition capability observed in α-Nrx. This, combined with a low-resolution molecular envelope obtained by a single particle reconstruction performed on negatively stained full-length Nrx1α sample, allowed us to derive a structural model of the α-Nrx ectodomain. This model will help us understand not only how the large α-Nrx ectodomain is accommodated in the synaptic cleft, but also how the trans-synaptic adhesion mediated by α- and β-Nrxs could differentially affect synaptic structure and function

Autoradiographic studies on proviral DNA synthesis in enucleated chick embryo fibroblasts infected with avian sarcoma virus

After infection of RNA tumor viruses to susceptible cells, viral RNA is reversely transcribed into proviral DNA. In order to disclose the site of proviral DNA synthesis in the cells, chick embryo fibroblasts (CEF) were enucleated by centrifugation in the presence of cytochalasin B, and the enucleated CEF (cytoplasts) were infected with B77 strain of avian sarcoma virus (B77-ASV). Incorporation of (3)H-thymidine into DNA in the cytoplasts was investigated by autoradiography. Photopositive grains were observed in cytoplasts infected with B77-ASV, but not in mock-infected cytoplasts. The photopositive grains in the cytoplasts infected with B77-ASV disappeared almost completely by DNase I treatment. N-demethyl rifampicin, which is a specific inhibitor of reverse transcriptase, inhibited the appearance of photopositive grains. The B77-ASV-infected cytoplasts were ultrathinsectioned for electron microscopic autoradiography. The photopositive grains appeared in the cytoplasm without relation to mitochondria. These results indicate that the proviral DNA synthesis is initiated in the cytoplasm of B77-ASV- infected chick embryo fibroblast without the direct participation of nucleus

Spry2 is a novel therapeutic target for periodontal tissue regeneration through fibroblast growth factor receptor signaling and epidermal growth factor signaling

Sprouty2 (Spry2) inhibits the activation of the extracellular signal-regulated kinase (ERK) pathway via receptor tyrosine kinase signaling. In a recent paper published in Journal of Cellular Biochemistry, we demonstrated that transfection of a dominant-negative mutant of Spry2 enhanced fibroblast growth factor (FGF)- and epidermal growth factor (EGF)-induced ERK activation in osteoblasts. In contrast, it decreased their activation in gingival epithelial cells. Consistent with these observations, the sequestration of Spry2 increased osteoblast proliferation by FGFR and EGFR stimulation, whereas it decreased gingival epithelial cell proliferation via the ubiquitination and degradation of EGF receptors (EGFR). In addition, reduction of Spry2 activity upregulated Runx2 expression and downregulated Twist, a negative regulator of Runx2 through FGFR and EGFR signaling, resulting in enhanced osteoblastogenesis in osteoblasts. Furthermore, we also found that suppression of Spry2 upregulated cell proliferation and migration in human periodontal ligament cell lines when they were stimulated by both FGF and EGF, and led to a shift in macrophage polarization, exerted immunosuppressive and tissue-repairing effects in macrophages. These results suggest that the application of a Spry2 inhibitor may effectively resolve inflammation by periodontitis and allow periodontal ligament and alveolar bone to grow and block the ingrowth of gingival epithelial cells in bony defects, biologically mimicking the barrier effect seen in conventional GTR. This approach has potential for developing a new regeneration strategy

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アル ジンケツ カンテイ ノ モンダイ テン ヒロサキ ジケン ゴハン ゲンイン ノ サイケントウ ノタメニ ソノ1

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