Formation of coniferyl alcohol from ferulic acid by the white rot fungus Trametes

The white rot fungus, Trametes sp., was cultivated in a medium containing ferulic acid, glucose and ethanol under aerobic conditions in submerged culture. The ferulic acid was transformed into coniferyl alcohol, coniferylaldehyde, dihydroconiferyl alcohol, vanillic acid, vanillyl alcohol, 2-methoxyhydroquinone and 2-methoxyquinone during 48–120 hr of cultivation. The amount of coniferyl alcohol in the culture reached a maximum after 90 hr with ca 40% of the initial amount of ferulic acid. Cinnamic acid, p-methoxycinnamic acid, 3,4-dimethoxycinnamic acid, p -coumaric acid and sinapic acid were also transformed into the corresponding alcohols, benzoic acids and benzyl alcohols in the fungus culture.     

CO2 fixation in leaves of a CAM plant without lower epidermis and the effect of CO2 on their deacidification

The effects of peeling the epidermis off Bryophyllum daigremontianumleaves on CO₂ uptake in light and darkness were investigated.Light-induced CO₂ uptake in the daytime was markedly enhancedin the peeled leaves, but dark fixation of CO₂ carried out atmidnight was not. The difference in promotion of CO₂ uptakein light and darkness was due to stomatal closing in the dayand opening at night. Also, deacidification was strikingly inhibitedby CO₂ in peeled leaves. (Received February 3, 1977; )     

Effect of alcohols on the collagen-phosphatidylcholine interaction

The interaction of phosphatidylcholine dispersions with acid soluble collagen separated from the skin of one month-old swine was studied to define the conditions facilitating the association of the collagen with lipids. When acid soluble collagen and phosphatidylcholine dispersions were incubated in 75 mM citrate buffer of pH 3.7 at 25 degrees C, the reisolated collagen fibrils did not contain appreciable amounts of phosphatidylcholine. However, the presence of n-propanol greatly promoted the retention of phosphatidylcholine, the amount of phosphatidylcholine associated being nearly 30% of collagen on a weight basis under optimal conditions. In contrast, methanol, ethanol, isopropanol, and n-butanol did not appreciably enhance the association of phosphatidylcholine with collagen. A limited inhibition of phosphatidylcholine retention was observed upon addition of sodium chloride to the propanol medium. The interaction of phosphatidylcholine with acid soluble collagen decreased sharply when temperature was increased above 30 degrees C; almost no phosphatidylcholine-collagen association occured at 40 degrees C. It appears that the enhanced association in the presence of n-propanol is due to a looseing of the collagen triple helix that exposes hydrophobic sites necessary for the interaction. However, the conversion of the triple helical structure to the random coil conformation by heating prevents the association of phosphatidylcholine with acid soluble collagen.     

A revision of the genusCyathorachis in Japan

Cyathorachis, a genus of Cretaceous tree fern rachises is revised. In addition to the type specimens of the sole species,C. fujiiana Ogura, two specimens collected near the type locality were examined histologically. The histological data being used for the taxonomical comparison were obtained from anatomical studies on the rachises of extant tree ferns. As a result of the study three species are recognized, two of which are new. As a result of this study, it appears that species differentiation of the tree ferns in the late Cretaceous age of Hokkaido was rather high, as it is now in tropical and subtropical regions. Contributions from the Laboratory of Phylogenetic Botany, Chiba University, No. 74.     

A scanning electron microscopy of the interstitial tissue of the boar testis

In this study, the architecture of the interstitial tissue of the boar testis was examined by using scanning and transmission electron microscopes. The boar testis was remarkable for the abundance of interstitial tissue, and Leydig cells having many microvilli in their surface were almost round in shape. Both bundles of collagen fibers and networks of reticular fibers were observed around the Leydig cells. The capillary in the interstitial tissue of the boar was a muscle type, and both pericytes and collagen fibers were observed around the capillaries. The lymphatic capillary was poorly developed in the interstitial tissues of the boar testis. Endothelial cells were the only component of the capillary wall, and anchoring filaments were often observed on the abluminal surface of the endothelium.     

Role of lysine-67 in the active site of class C beta-lactamase from Citrobacter freundii GN346

Citrobacter freundii GN346 produces a class C beta-lactamase exhibiting the substrate profile of a typical cephalosporinase. The structural and promoter regions of the cephalosporinase gene, comprising 1408 nucleotides, were completely sequenced. The amino acid sequence of the mature enzyme, comprising 361 amino acids, and its molecular mass, 39,878 Da, were determined. The active site was confirmed to be Ser-64. The amino acid sequence of the enzyme differs from that of the cephalosporinase of C. freundii OS60 by nine residues. The nucleotide sequence of the promoter region suggests a possible attenuator structure. Lys-67, one of the most conserved residues found in class A and C beta-lactamases and penicillin-binding proteins, was converted into arginine, threonine or glutamic acid through site-directed mutagenesis. The Glu-67 enzyme had lost the catalytic activity and the Thr-67 enzyme only showed a trace of activity. The Arg-67 enzyme, which retained a significant amount of the activity, was purified. The Km values of the Arg-67 enzyme for cephalothin, cephaloridine and benzylpenicillin are 13-19 times those of the wild-type enzyme; the kcat values for the three substrates are 37%, 3%, and 36% those of the wild-type enzyme, respectively.     

Cellular communication network factor 3 in cartilage development and maintenance

Cellular communication network factor (CCN) 3 is one of the classical members of the CCN family, which are characterized by common molecular structures and multiple functionalities. Although this protein was discovered as a gene product overexpressed in a truncated form in nephroblastoma, recent studies have revealed its physiological roles in the development and homeostasis of mammalian species, in addition to its pathological association with a number of diseases. Cartilage is a tissue that creates most of the bony parts and cartilaginous tissues that constitute the human skeleton, in which CCN3 is also differentially produced to exert its molecular missions therein. In this review article, after the summary of the molecular structure and function of CCN3, recent findings on the regulation of ccn3 expression and the roles of CCN3 in endochondral ossification, cartilage development, maintenance and disorders are introduced with an emphasis on the metabolic regulation and function of this matricellular multifunctional molecule.     

Involvement of tyrosine kinase in the hypoxia/reoxygenation-induced gap junctional intercellular communication abnormality in cultured human umbilical vein endothelial cells

Vascular endothelial cells (EC), communicating with one another across gap junctions, are usually made dysfunctional by hypoxia and reoxygenation (H/R); however, very limited information exists regarding the effects of H/R on the endothelial gap junctions. We investigated whether H/R interferes with endothelial gap junctional intercellular communication (GJIC). After human umbilical vein EC had grown to confluence, they were exposed to hypoxia (pO₂ < 0.1%) for 12–16 h and then returned to normal atmospheric conditions for reoxygenation. At 0-, 2-, 4-, 6-h reoxygenation, GJIC was detected by means of a fluorescence recovery after a photobleaching technique. The results demonstrated that a GJIC reduction (about 20% less than that under normoxia) was induced after 2 h of reoxygenation; after 4 h of reoxygenation, it began to recover (to about 10% less than that under normoxia); and after 6 h of reoxygenation, GJIC was restored to the normal level. Calphostin C (1 × 10⁻⁷ mol/l), a specific protein kinase C inhibitor, partially inhibited the reduction in GJIC (resulting in a level about 10% less than that under normoxia), whereas the tyrosine kinase inhibitor genistein (10 µmol/L) completely blocked the reduction in GJIC. Vanadate (1.5 mmol/l), a tyrosine phosphatase inhibitor, amplified the inhibitory effect of H/R on GJIC (to about 40% less than that under normoxia). Immunofluorescence and immunoprecipitation showed that 2-h reoxygenation significantly stimulated tyrosine protein phosphorylation, and this phosphorylation event was obviously enhanced by vanadate. The results of Western blotting showed that the gap junctional protein connexin 43 (Cx43) was phosphorylated by H/R; moreover, immunoprecipitation demonstrated that 2-h reoxygenation induced a prominent increase of tyrosine phosphorylation of Cx43 compared with that under normoxia. These data indicate that H/R induces a transient endothelial GJIC dysfunction through the activation of tyrosine kinase and phosphorylation of tyrosine residues of Cx43. J. Cell. Physiol. 180:305–313, 1999. © 1999 Wiley-Liss, Inc.     

Connective tissue growth factor induces the proliferation, migration, and tube formation of vascular endothelial cells in vitro, and angiogenesis in vivo.

Connective tissue growth factor (CTGF) is a novel cysteine-rich, secreted protein. Recently, we found that inhibition of the endogenous expression of CTGF by its antisense oligonucleotide and antisense RNA suppresses the proliferation and migration of vascular endothelial cells. In the present study, the following observations demonstrated the angiogenic function of CTGF in vitro and in vivo: (i) purified recombinant CTGF (rCTGF) promoted the adhesion, proliferation and migration of vascular endothelial cells in a dose-dependent manner under serum-free conditions, and these effects were inhibited by anti-CTGF antibodies; (ii) rCTGF markedly induced the tube formation of vascular endothelial cells, and this effect was stronger than that of basic fibroblast growth factor or vascular endothelial growth factor; (iii) application of rCTGF to the chicken chorioallantoic membrane resulted in a gross angiogenic response, and this effect was also inhibited by anti-CTGF antibodies. (iv) rCTGF injected with collagen gel into the backs of mice induced strong angiogenesis in vivo. These findings indicate that CTGF is a novel, potent angiogenesis factor which functions in multi-stages in this process.