Changes in Triterpenoid Content during the Growth Cycle of Cultured Plant Cells

Similar changes in the pentacyclic triterpenoid contents wereobserved during the growth cycle of Datura innoxia, Luffa cylindricaand Lycopersicon esculentum seedling callus cells in batch culture.Triterpenoid contents decreased for several days after callusinoculation, then increased rapidly during the mid and lateexponential phases of growth. (Received May 28, 1984; Accepted September 13, 1984)     

The Accumulation of Cyclin-Dependent Kinase Inhibitor p27Is a Primary Response to Staurosporine and Independent of G1 Cell Cycle Arrest

The staurosporine-induced G1 cell cycle arrest was analyzed in a variety of cell lines which includes human tumor cell lines and oncogene-transformed NIH3T3 cell lines. All the cell lines which were sensitive to staurosporine-induced G1 arrest contained a functional retinoblastoma protein (pRB). However, when pRB-lacking fibroblast cells derived from pRB knockout mice were tested they were also sensitive to G1 arrest by staurosporine, indicating that the inactivation of pRB alone is not sufficient for the abrogation of staurosporine-induced G1 arrest. In searching for a common event caused by staurosporine, the cyclin-dependent kinase (CDK) inhibitor protein p27^kip1but not p21^cip1was found to accumulate after staurosporine treatment in all the cell lines examined. This accumulation occurred regardless of the induction of the G1 arrest. The result indicates that the accumulation of p27^kip1is the cell's primary response to staurosporine and that the capability of staurosporine to induce G1 arrest depends on the integrity of cell cycle regulatory components which are downstream of p27^kip1.     

Enzyme Aided Regioselective Acylation of Nucleosides

Abstract

Selective protection of the three hydroxyl groups of sugar moiety of nucleosides have been studied by enzyme-catalyzed esterification in organic solvents. Selectively protected products were obtained. The reaction provides an efficient method for selective protection of nucleosides.     

Transmembrane molecules for phylogenetic analyses of pathogenic protists: Leishmania-specific informative sites in hydrophilic loops of trans- endoplasmic reticulum N-acetylglucosamine-1-phosphate transferase

A sequence database was created for the Leishmania N-acetylglucosamine-1-phosphate transferase (nagt) gene from 193 independent isolates. PCR products of this single-copy gene were analyzed for restriction fragment length polymorphism based on seven nagt sequences initially available. We subsequently sequenced 77 samples and found 19 new variants (genotypes). Alignment of all 26 nagt sequences is gap free, except for a single codon addition or deletion. Phylogenetic analyses of the sequences allow grouping the isolates into three subgenera, each consisting of recognized species complexes, i.e., subgenus Leishmania (L. amazonensis-L. mexicana, L. donovani-L. infantum, L. tropica, L. major, and L. turanica-L. gerbilli), subgenus Viannia (L. braziliensis, L. panamensis), and one unclassified (L. enriettii) species. This hierarchy of grouping is also supported by sequence analyses of selected samples for additional single-copy genes present on different chromosomes. Intraspecies divergence of nagt varies considerably with different species complexes. Interestingly, species complexes with less subspecies divergence are more widely distributed than those that are more divergent. The relevance of this to Leishmania evolutionary adaptation is discussed. Heterozygosity of subspecies variants contributes to intraspecies diversity, which is prominent in L. tropica but not in L. donovani-L. infantum. This disparity is thought to result from the genetic recombination of the respective species at different times as a rare event during their predominantly clonal evolution. Phylogenetically useful sites of nagt are restricted largely to several extended hydrophilic loops predicted from hypothetical models of Leishmania NAGT as an endoplasmic reticulum transmembrane protein. In silico analyses of nagt from fungi and other protozoa further illustrate the potential value of this and, perhaps, other similar transmembrane molecules for phylogenetic analyses of single-cell eukaryotes.     

Synthesis and antibacterial activity of 6-O-(heteroaryl-isoxazolyl)propynyl 2-fluoro ketolides

Macrolide antibiotics are widely prescribed for the treatment of respiratory tract infections; however, the increasing prevalence of macrolide-resistant pathogens is a public health concern. Therefore, the development of new macrolide derivatives with activities against resistant pathogens is urgently needed. A series of novel 6-O-(heteroaryl-isoxazolyl)propynyl 2-fluoro ketolides has been synthesized from erythromycin A. These compounds have shown very promising in vitro and in vivo antibacterial activities against key respiratory pathogens including erythromycin-susceptible/resistant strains.     

Urea-dependent unfolding of HIV-1 protease studied by circular dichroism and small-angle X-ray scattering

HIV-1 protease is responsible for the maturation of infective virions, and is one of the targets of drugs against AIDS. It is an aspartic protease with a 99-resiude polypeptide dimerized. Previous study with fluorescence and sedimentation measurements revealed that the protein was unfolded with concomitant dissociation of the subunits. In the present study, we investigated urea-dependent unfolding of HIV-1 protease with CD and SAXS in order to monitor the secondary structure and the global size and shape of the molecule, respectively. The unfolding parameters estimated by both methods were almost the same, indicating that the dissociation of the subunits accompanied the disruption of their internal structures. This is in line with the previous results, and moreover some residual structures were suggested to be present in the unfolded state. The distinct difference, as compared with the unfolding of pepsin, was interpreted from the point of their molecular architectures.     

Desert hedgehog‐patched 2 expression in peripheral nerves during Wallerian degeneration and regeneration

Hedgehog proteins are important in the development of the nervous system. As Desert hedgehog (Dhh) is involved in the development of peripheral nerves and is expressed in adult nerves, it may play a role in the maintenance of adult nerves and degeneration and regeneration after injury. We firstly investigated the Dhh‐receptors, which are expressed in mouse adult nerves. The Dhh receptor patched(ptc)2 was detected in adult sciatic nerves using RT‐PCR, however, ptc1 was undetectable under the same experimental condition. Using RT‐PCR in purified cultures of mouse Schwann cells and fibroblasts, we found ptc2 mRNA in Schwann cells, and at much lower levels, in fibroblasts. By immunohistochemistry, Ptc2 protein was seen on unmyelinated nerve fibers. Then we induced crush injury to the sciatic nerves of wild‐type (WT) and dhh‐null mice and the distal stumps of injured nerves were analyzed morphologically at different time points and expression of dhh and related receptors was also measured by RT‐PCR in WT mice. In dhh‐null mice, degeneration of myelinated fibers was more severe than in WT mice. Furthermore, in regenerated nerves of dhh‐null mice, minifascicular formation was even more extensive than in dhh‐null intact nerves. Both dhh and ptc2 mRNA levels were down‐regulated during the degenerative phase postinjury in WT mice, while levels rose again during the phase of nerve regeneration. These results suggest that the Dhh‐Ptc2 signaling pathway may be involved in the maintenance of adult nerves and may be one of the factors that directly or indirectly determines the response of peripheral nerves to injury. © 2005 Wiley Periodicals, Inc. J Neurobiol, 2006     

Synthesis, SAR studies, and evaluation of 1,4-benzoxazepine derivatives as selective 5-HT1A receptor agonists with neuroprotective effect: Discovery of Piclozotan

A new series of 1,4-benzoxazepine derivatives was designed, synthesized, and evaluated for binding to 5-HT1A receptor and cerebral anti-ischemic effect. A lot of compounds exhibited nanomolar affinity for 5-HT1A receptor with good selectivity over both dopamine D2 and alpha1-adrenergic receptors. Among these compounds, 3-chloro-4-[4-[4-(2-pyridinyl)-1,2,3,6-tetrahydropyridin-1-yl]butyl]-1, 4-benzoxazepin-5(4H)-one (50: SUN N4057 (Piclozotan) as 2HCl salt) showed remarkable neuroprotective activity in a transient middle cerebral artery occlusion (t-MCAO) model.     

Nuclear compartmentalization is abolished during fission yeast meiosis

In eukaryotic cells, the nuclear envelope partitions the nucleus from the cytoplasm. The fission yeast Schizosaccharomyces pombe undergoes closed mitosis in which the nuclear envelope persists rather than being broken down, as in higher eukaryotic cells. It is therefore assumed that nucleocytoplasmic transport continues during the cell cycle. Here we show that nuclear transport is, in fact, abolished specifically during anaphase of the second meiotic nuclear division. During that time, both nucleoplasmic and cytoplasmic proteins disperse throughout the cell, reminiscent of the open mitosis of higher eukaryotes, but the architecture of the S. pombe nuclear envelope itself persists. This functional alteration of the nucleocytoplasmic barrier is likely induced by spore wall formation, because ectopic induction of sporulation signaling leads to premature dispersion of nucleoplasmic proteins. A photobleaching assay demonstrated that nuclear envelope permeability increases abruptly at the onset of anaphase of the second meiotic division. The permeability was not altered when sporulation was inhibited by blocking the trafficking of forespore-membrane vesicles from the endoplasmic reticulum to the Golgi. The evidence indicates that yeast gametogenesis produces vesicle transport-mediated forespore membranes by inducing nuclear envelope permeabilization.