Deoxyribonucleic Acid Replication and Cell Division in Escherichia coli at 33 C

At 33 C (60-min generation time) the time required to replicate the chromosome is C = 60 min. The time between the end of a round of replication and cell division is D = 20 min, as at 37 C. Nalidixic acid and a temperature shift in a dnaB mutant give identical results for the determination of the end of a round of replication.     

Replication of a specific terminal chromosome segment in Escherichia coli which is required for cell division

In the absence of protein synthesis, DNA replication does not proceed to the very end of the chromosome. A terminal segment remains unreplicated. This segment can be selectively labelled. Its size is about 0·5% of the chromosome and its replication is required for subsequent cell division.     

Resonance Raman scattering by bacterial spores

Abstract The laser Raman spectra of lyophilized spores of 5 species (6 strains) of Bacillus were examined. Under the experimental conditions employed, only Bacillus megaterium species showed 3 intense Raman bands at 1515, 1157 and 1007 cm⁻¹. Spores of other species, despite their high content of dipicolinic acid, did not show distinct Raman signals.
The strong, scattering bands at 1515 and 1157 cm⁻¹ in the spectra of spores of B. megaterium may be attributable to conjugated double-bond systems, probably of membrane-associated carotenoids. Their high intensities are due to resonance enhancement.     

Homer proteins control neuronal differentiation through IP(3) receptor signaling

Neurons expand, sustain or prune their dendritic trees during ontogenesis [Cline, H.T. (2001). Dendritic arbor development and synaptogenesis. Curr. Opin. Neurobiol. 11, 118-126; Wong, W.T. and Wong, R.O.L. (2000) Rapid dendritic movements during synapse formation and rearrangement. Curr. Opin. Neurobiol. 10, 118-124] which critically depends on neuronal activity [Wong, W.T., Faulkner-Jones, B.E., Sanes, J.R. and Wong, R.O.L. (2000) Rapid dendritic remodeling in the developing retina: dependence on neurotransmission and reciprocal regulation by Rac and Rho. J. Neurosci. 20, 5024-5036; Li, Z., Van Aelst, L. and Cline, H.T. (2000) Rho GTPases regulate distinct aspects of dendritic arbor growth in Xenopus central neurons in vivo. Nat. Neurosci. 3, 217-225; Wong, W.T. and Wong, R.O.L. (2001) Changing specificity of neurotransmitter regulation of rapid dendritic remodeling during synaptogenesis. Nat. Neurosci. 4, 351-352.] and sub-cellular Ca(2+) signals [Lohmann, C., Myhr, K.L. and Wong, R.O. (2002) Transmitter-evoked local calcium release stabilizes developing dendrites, Nature 418, 177-181.]. The role of synaptic clustering proteins connecting both processes is unclear. Here, we show that expression levels of Vesl-1/Homer 1 isoforms critically control properties of Ca(2+) release from intracellular stores and dendritic morphology of CNS neurons. Vesl-1L/Homer 1c, an isoform with a functional WH1 and coiled-coil domain, but not isoforms missing these features were capable of potentiating intracellular calcium signaling activity indicating that such regulatory interactions function as a general paradigm in cellular differentiation and are subject to changes in expression levels of Vesl/Homer isoforms.     

Involvement of SUMO modification in MBD1- and MCAF1-mediated heterochromatin formation

Small ubiquitin-related modifiers, SUMO-2/3 and SUMO-1, are involved in gene regulation and nuclear structures. However, little is known about the roles of SUMO, in heterochromatin formation of mammalian cells. Here we demonstrate that SUMOs directly interact with human MCAF1, which forms complexes with either the methyl-CpG-binding protein MBD1 or SETDB1, which trimethylates histone H3 at lysine 9 (H3-K9) in the presence of MCAF1. Modification of MBD1 with either SUMO-2/3 or SUMO-1 facilitated the interaction between MBD1 and MCAF1, suggesting that SUMOylation links the methylation of DNA and histones. In a cultured human cell line, SUMOs were localized in MBD1- and MCAF1-containing heterochromatin regions that were enriched in trimethyl-H3-K9 and the heterochromatin proteins HP1beta and HP1gamma. Specific knockdown of either SUMO-2/3 or SUMO-1 induced dissociation of MCAF1, trimethyl-H3-K9, and the HP1 proteins from the MBD1-containing heterochromatin foci, suggesting a requirement for SUMOs for heterochromatin assembly. These findings provide insights into the roles of SUMOylation in the regulation of heterochromatin formation and gene silencing.     

Induction of antitumor activities in spleen cells from mice and rats activated with lipopolysaccharide immobilized on beads

Summary Lipopolysaccharides (LPS) were coupled to polystyrene beads in order to apply the LPS without toxicity. The antitumor activity of the LPS-immobilizing beads was studied in experiments in vitro and in vivo. In vitro studies showed that spleen cells from C3H/HeN mice stimulated by beads immobilizing LPS fromEscherichia coli produced cytolytic activity as strong as that of lymphokine-activated killer (LAK) cells. Spleen cells from Sprague-Dawley rats stimulated by beads immobilizing LPS fromSalmonella minnesota produced cytolytic activity stronger than that of LAK cells. However, spleen cells stimulated by beads immobilizing each component of the LPS separately could not induce cytolysis. Contact stimulation, even for a brief period, sufficed for cytolytic activity, and was enhanced by culture for 48–72 h. Through in vivo studies, the suppression of tumor growth and a prolongation of the survival time were observed in tumorbearing mice injected with spleen cells activated by beads immobilizing LPS fromE.coli, and in mice injected with LAK cells. The effect of the activated spleen cells was stronger than that of the LAK cells. In rats bearing metastatic tumors, spleen cells activated by beads immobilizing LPS fromS.minnesota suppressed lung metastases more strongly than did LAK cells. These findings indicate that LPS immobilized by beads induced killer cells more strongly than interleukin-2. Ex vivo immunomodulation with LPS-immobilizing beads can be applied usefully as an anticancer treatment.     

Interleukin-10 inhibits both production of cytokines and expression of cytokine receptors in microglia

Microglia, macrophage-like cells in the CNS, are multifunctional cells; they play an important role in removal of dead cells or their remnants by phagocytosis in the CNS degeneration and are one of important cells in the CNS cytokine network to produce and respond to a variety of cytokines. The functions of microglia are regulated by inhibitory cytokines. We have reported the expression of interleukin (IL)-10, one of the inhibitory cytokines, and its receptor in mouse microglia; therefore, IL-10 may affect microglial functions. In this study, we investigated the effects of IL-10 on purified microglia in culture. IL-10 inhibited lipopolysaccharide-induced IL-1beta and tumor necrosis factor-alpha production, lysosomal enzyme activity, and superoxide anion production in a dose-dependent manner, but did not affect granulocyte/ macrophage colony-stimulating factor-dependent proliferation of microglia. IL-10 also decreased the expression of both IL-6 receptor and lipopolysaccharide-induced IL-2 receptor but not IL-4 receptor on microglia as measured by flow cytometric analysis with an indirect immunofluorescence technique. IL-10 also decreased mRNA expression of IL-2 and IL-6 cytokine receptors. These results suggest that IL-10 is a unique and potent inhibitory factor in the CNS cytokine network involved in decreasing the expression of cytokine receptors as well as cytokine production by microglia.     

Inhibition of P-selectin-mediated cell adhesion by a sulfated derivative of sialic acid

P-selectin, a carbohydrate-binding cell adhesion molecule expressed on activated endothelial cells and platelets, plays a key role in the recruitment of leukocytes to inflammatory and hemorrhagic sites. It simultaneously recognizes a sialic acid-containing carbohydrate chain and the sulfated tyrosine residues of a specific counter-receptor expressed on the leukocyte surface. We examined the inhibitory effects of a synthetic sulfated derivative of sialic acid (NMSO3) on P-selectin-mediated cell adhesion and found the following: (1) P-selectin/IgG chimera bound to immobilized NMSO3. (2) The binding of P-selectin/IgG chimera to purified P-selectin glycoprotein ligand-1 was inhibited by soluble NMSO3. (3) The adhesion of HL60 cells to P-selectin-expressing CHO cells was inhibited by NMSO3. (4) NMSO3 inhibited P-selectin-induced tumor necrosis factor-alpha production in monocytes and activated platelet-induced generation of reactive oxygen species in neutrophils. In conclusion, NMSO3 acts as a specific inhibitor for P-selectin-mediated cell adhesion and for adhesion-dependent leukocyte activation.