Induction of cell autolysis of Bacillus subtilis with lysophosphatidylcholine

Lysophosphatidylcholine (LPC) was found to cause autolysis of Bacillus subtilis 168 cells growing logarithmically at concentrations higher than 20 m, by inducing the activity of autolytic enzymes. The lytic activity depended upon the carbon-chain length of the acyl moiety in the LPC molecule, being most effective with palmitoyl LPC. Lysophosphatidylethanolamine also caused cell lysis but to a lesser extent, whereas lysophosphatidylglycerol did not. LPC stimulated cell autolysis in TRIS-KCl buffer and potassium phosphate buffer but was ineffective in distilled water. LPC had no influence on the activity in vitro of partially purified autolytic enzymes.Correspondence to: T. Tsuchido     

Purification of cytochrome P450 and ferredoxin, involved in bisphenol A degradation, from Sphingomonas sp. strain AO1

In a previous study (M. Sasaki, J. Maki, K. Oshiman, Y. Matsumura, and T. Tsuchido, Biodegradation 16:449-459, 2005), the cytochrome P450 monooxygenase system was shown to be involved in bisphenol A (BPA) degradation by Sphingomonas sp. strain AO1. In the present investigation, we purified the components of this monooxygenase, cytochrome P450 (P450bisd), ferredoxin (Fd(bisd)), and ferredoxin reductase (Red(bisd)). We demonstrated that P450bisd and Fd(bisd) are homodimeric proteins with molecular masses of 102.3 and 19.1 kDa, respectively, by gel filtration chromatography analysis. Spectroscopic analysis of Fd(bisd) revealed the presence of a putidaredoxin-type [2Fe-2S] cluster. P450(bisd), in the presence of Fd(bisd), Red(bisd), and NADH, was able to convert BPA. The K(m) and kcat values for BPA degradation were 85 +/- 4.7 microM and 3.9 +/- 0.04 min(-1), respectively. NADPH, spinach ferredoxin, and spinach ferredoxin reductase resulted in weak monooxygenase activity. These results indicated that the electron transport system of P450bisd might exhibit strict specificity. Two BPA degradation products of the P450(bisd) system were detected by high-performance liquid chromatography analysis and were thought to be 1,2-bis(4-hydroxyphenyl)-2-propanol and 2,2-bis(4-hydroxyphenyl)-1-propanol based on mass spectrometry-mass spectrometry analysis. This is the first report demonstrating that the cytochrome P450 monooxygenase system in bacteria is involved in BPA degradation.     

Interactions between the Nucleus and Cytoplasmic Organelles during the Cell Cycle of Euglena gracilis in Synchronized Cultures III. Association between the Nucleus and Chloroplasts at an Early Stage in the Cell Cycle under Photoorganotrophic Conditions (Part II)

Three-dimensional models of chloroplasts before and during theirassociation with the nucleus in Euglena gracilis Z were constructedbased on serial sections of cells taken during an early phaseof the cell cycle in synchronized cultures under photoorganotrophiccondition. Before association, the cell contained two to threechloroplasts which were composed of elongated tubular bodiesextending in different directions toward the cell periphery.These tubular arms of chloroplasts were drawn toward the nucleus,and folded, with concurrent coalescence, into a single giantbody surrounding the nucleus. A DAPI-fluorescence photomicrographof a giant chloroplast revealed that the chloroplast-nucleoidswere in the form of a continuous strand lying throughout thechloroplast body, and that some protuberances from the nucleoidstrand were in especially close proximity to the nuclear periphery. A new mode of the chloroplast-nucleus connection was observed.The nucleus had conspicuous protrusions, whose distal ends wereconnected with the chloroplast. The outer membrane of the nuclearenvelope was continuous with the outer chloroplast membrane,and at some sites, an open space was formed between the innernuclear membrane and the inner chloroplast membrane. ¹ This work was reported at the 48th Annual Meeting of the BotanicalSociety of Japan, held in Kyoto, in October, 1983. For PartI see Ehara et al. (1984).     

A method for the direct identification of differentiating muscle cells by a fluorescent mitochondrial dye

Identification of differentiating muscle cells generally requires fixation, antibodies directed against muscle specific proteins, and lengthy staining processes or, alternatively, transfection of muscle specific reporter genes driving GFP expression. In this study, we examined the possibility of using the robust mitochondrial network seen in maturing muscle cells as a marker of cellular differentiation. The mitochondrial fluorescent tracking dye, MitoTracker, which is a cell-permeable, low toxicity, fluorescent dye, allowed us to distinguish and track living differentiating muscle cells visually by epi-fluorescence microscopy. MitoTracker staining provides a robust and simple detection strategy for living differentiating cells in culture without the need for fixation or biochemical processing.     

Properties of the electrogenic activity of bacteriorhodopsin

In this study, we analyzed the photoelectric current generated by bacteriorhodopsin adsorbed on a polymer film, “Lumirror” (Muneyuki et al. in FEBS Lett 427:109–114, 1998). We could examine the photoelectric current over a wide range of light intensity and pH values using the same membrane owing to the mechanical and chemical stability of the thin polymer film. We analyzed the photoelectric current by comparison with a simple equivalent electric circuit. Analysis of experimental results obtained at different light intensities suggested that the electromotive force of the bacteriorhodopsin was independent of light intensity. The pH dependence of the photoelectric current suggested that the bacteriorhodopsin could generate a maximum electromotive force at approximately pH 6.     

Identification of Distinct Ligands for the C-type Lectin Receptors Mincle and Dectin-2 in the Pathogenic Fungus Malassezia

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Highlights? Mincle and Dectin-2 cooperatively recognize Malassezia through distinct ligands ? The Mincle ligands in Malassezia are glucosyl- and mannosylglycolipids ? The Dectin-2 ligand in Malassezia is an O-linked mannobiose-rich glycoprotein ? Cytokine response to the respective ligands was impaired in Mincle^?/? and Dectin-2^?/? DCs     

Periarticular Osteophytes as an Appendicular Joint Stress Marker (JSM): Analysis in a Contemporary Japanese Skeletal Collection

Objective

The aim of this study was to investigate the possibility that periarticular osteophytes plays a role as a appendicular joint stress marker (JSM) which reflects the biomechanical stresses on individuals and populations.

Methods

A total of 366 contemporary Japanese skeletons (231 males, 135 females) were examined closely to evaluate the periarticular osteophytes of six major joints, the shoulder, elbow, wrist, hip, knee, and ankle and osteophyte scores (OS) were determined using an original grading system. These scores were aggregated and analyzed statistically from some viewpoints.

Results

All of the OS for the respective joints were correlated logarithmically with the age-at-death of the individuals. For 70 individuals, in whom both sides of all six joints were evaluated without missing values, the age-standardized OS were calculated. A right side dominancy was recognized in the joints of the upper extremities, shoulder and wrist joints, and the bilateral correlations were large in the three joints on the lower extremity. For the shoulder joint and the hip joint, it was inferred by some distinctions that systemic factors were relatively large. All of these six joints could be assorted by the extent of systemic and local factors on osteophytes formation. Moreover, when the age-standardized OS of all the joints was summed up, some individuals had significantly high total scores, and others had significantly low total scores; namely, all of the individuals varied greatly in their systemic predisposition for osteophytes formation.

Conclusions

This study demonstrated the significance of periarticular osteophytes; the evaluating system for OS could be used to detect differences among joints and individuals. Periarticular osteophytes could be applied as an appendicular joint stress marker (JSM); by applying OS evaluating system for skeletal populations, intra-skeletal and inter-skeletal variations in biomechanical stresses throughout the lives could be clarified.     

Design and synthesis of C35-fluorinated solamins and their growth inhibitory activities against human cancer cell lines

The convergent synthesis of C35-fluorinated analogues of solamin, a mono-THF Annonaceous acetogenin, has been achieved by the Sonogashira coupling of the THF ring fragment and the fluorinated γ-lactone fragment. It was revealed that the number of fluorine atoms on the γ-lactone moiety affects the growth inhibitory activities against human cancer cell lines.     

N-cadherin regulates p38 MAPK signaling via association with JNK-associated leucine zipper protein: implications for neurodegeneration in Alzheimer disease

Synaptic loss, which strongly correlates with the decline of cognitive function, is one of the pathological hallmarks of Alzheimer disease. N-cadherin is a cell adhesion molecule essential for synaptic contact and is involved in the intracellular signaling pathway at the synapse. Here we report that the functional disruption of N-cadherin-mediated cell contact activated p38 MAPK in murine primary neurons, followed by neuronal death. We further observed that treatment with Aβ(42) decreased cellular N-cadherin expression through NMDA receptors accompanied by increased phosphorylation of both p38 MAPK and Tau in murine primary neurons. Moreover, expression levels of phosphorylated p38 MAPK were negatively correlated with that of N-cadherin in human brains. Proteomic analysis of human brains identified a novel interaction between N-cadherin and JNK-associated leucine zipper protein (JLP), a scaffolding protein involved in the p38 MAPK signaling pathway. We demonstrated that N-cadherin expression had an inhibitory effect on JLP-mediated p38 MAPK signal activation by decreasing the interaction between JLP and p38 MAPK in COS7 cells. Also, this study demonstrated a novel physical and functional association between N-cadherin and p38 MAPK and suggested neuroprotective roles of cadherin-based synaptic contact. The dissociation of N-cadherin-mediated synaptic contact by Aβ may underlie the pathological basis of neurodegeneration such as neuronal death, synaptic loss, and Tau phosphorylation in Alzheimer disease brain.     

Casein kinase 2 is the major enzyme in brain that phosphorylates Ser129 of human alpha-synuclein: Implication for alpha-synucleinopathies

In Lewy body diseases and multiple system atrophy, α-synuclein is hyperphosphorylated at Ser129, suggesting a role in pathogenesis. Here, we report purification of the protein kinase in rat brain that phosphorylates Ser129 and its identification as casein kinase-2 (CK2). We show that most of the activity can be inhibited by heparin, an inhibitor of CK2. Phosphorylated Ser129 was detected in primary cultured neurons and inhibited by CK2 inhibitors. In some cases of Lewy body disease, CK2-like immunoreactivity was recovered in the sarkosyl-insoluble fraction, which was enriched in phosphorylated α-synuclein. Taken together, these findings suggest that CK2 may be involved in the hyperphosphorylation of α-synuclein in α-synucleinopathies.