A new species ofCortinarius sectionBolares from Japan

Cortinarius reticulisporus sp. nov., found in deciduous forests, is described and illustrated. It closely resemblesCortinarius rubicundulusin its pileus color and the vivid yellowing of the context on bruising, but differs from the latter in its subglobose to broadly ellipsoid basidiospores with quotient of the length/width smaller than 1.4 and particularly fine lines connecting warts when observed under SEM. The differences betweenCortinarius reticulisporus and other similar taxa are briefly discussed.     

Intra- and intermolecular beta-pleated sheet formation in glutamine-repeat inserted myoglobin as a model for polyglutamine diseases

An aberrant structure of the expanded polyglutamine might be involved in the formation of aggregates in CAG repeat diseases. To elucidate structural properties of the expanded polyglutamine, we prepared sperm whale myoglobin (Mb) mutants, in which 12, 28, 35, and 50 repeats of glutamine were inserted at the corner between the C and D helices (Gln(12), Gln(28), Gln(35), and Gln(50), respectively). Circular dichroism and IR spectroscopies showed that the expanded polyglutamine, which was recognized by the monoclonal antibody 1C2 in Gln(28), Gln(35), and Gln(50) Mb forms an antiparallel beta-pleated sheet structure. Gln(50) Mb aggregates were found to comprise an intermolecular antiparallel beta-pleated sheet. Fluorescence together with (1)H NMR spectra revealed partial unfolding of the protein surface in Gln(35) and Gln(50) Mb, although the structural changes in the protein core were rather small. The present results indicate that the fluctuating beta-pleated sheet of the expanded polyglutamine exposed on the protein surface facilitates the formation of aggregates through intermolecular interactions. The present study has first established and characterized structural properties of a molecular model for polyglutamine diseases in which various lengths of polyglutamine including a pathologically expanded glutamine repeat were inserted into a structurally known protein.     

Ultrastructural evidence for dendritic release of acetylcholinesterase in the rat substantia nigra

Morphological evidence for dendritic secretion of acetylcholinesterase (AChE) in rat substantia nigra--a physiologically known phenomenon--was searched by means of a modified cytochemical method devised for fine localization of AChE activity at the electron microscopic level. DAB precipitate was observed in cluster of small vesicles in contact with the plasma membrane and in the extracellular space in the vicinity of the vesicles. Single coated or uncoated large vesicles filled with stained material were found in the cytoplasm of the dendrites at distance from or in contact with the plasma membrane. Immunoperoxidase staining with specific anti-serum against rat AChE gave similar localization of AChE. These results suggest that AChE is released from the dendrites of the nigral neurons by a process of vesicular exocytosis and captured by endocytosis. The relation of this process to a putative release from the smooth endoplasmic reticulum remains to be elucidated.     

Endoscopic endonasal reduction for blowout fracture of the medial orbital wall

Endoscopic endonasal reductions have been addressed in 63 patients with blowout fracture of the medial orbital wall since 1992. The operations were carried out under general anesthesia with a magnified operative space projected on a television monitor by a charge coupled device video camera attached to the endoscope. The middle nasal turbinate was fractured toward the nasal septum, the uncinate process was cut off, and the bulla was opened. The ethmoidal bony partition and the mucous membrane were removed; however, the fractured bone chips of the medial orbital wall were preserved. The herniated orbital contents were pressed back into the orbital cavity, and the medial wall was set with 2-mm-thick bent silicone plates placed in the ethmoidal sinus. The plates were removed in the outpatient clinic 2 months after the operation. The surgical results of 21 patients treated with endoscopic reduction were compared with those of four patients treated with transfacial reduction with an iliac bone graft. All of the patients had isolated medial wall fracture and became aware of diplopia within 15 degrees in any direction from the primary position (straight gaze) before the operation; the follow-up period covered 6 months. The patients were classified into two categories according to postoperative double vision: "good," indicating no double vision or diplopia of more than 45 degrees, and "poor," diplopia of less than 45 degrees. Improvement of diplopia was observed in all patients without any complication. Of the 21 patients who underwent endoscopic reductions, 17 were classified as "good" and four as "poor." On the other hand, of the four patients who underwent transfacial reductions, three were classified as "good" and one as "poor." Significant differences were not observed between the surgical results of our two methods. Endoscopic endonasal reduction showed greater aesthetic advantages and, moreover, required no grafting. This technique is suggested as one of the most reasonable treatments of medial orbital wall fractures.     

Cloning and characterization of the deoxyribonuclease sd alpha gene from Streptococcus pyogenes

The pathogenesis of Streptococcus pyogenes infection, especially toxic shock-like syndrome (TSLS), is still not fully understood; however, the exoproteins have been considered to play a role. We analyzed the culture supernatant proteins (exoproteins) from a TSLS-related isolate belonging to M3 serotype S. pyogenes by two-dimensional gel electrophoresis and characterized a single protein spot by using BLAST database. We cloned the gene of this protein and named it sdα, which was similar to the deoxyribonuclease (DNase) sdc of S. equisimilis. We showed that the recombinant protein from the sdα gene had DNase activity. By polymerase chain reaction, we found that the sdα gene was present in most clinically isolated S. pyogenes including TSLS-related isolates. We thus conclude that Sdα is a new DNase of S. pyogenes. Received: 27 August 2001 / Accepted: 19 October 2001     

CHIP is associated with Parkin,a gene responsible for familial Parkinson's disease,and enhances its ubiquitin ligase activity

Unfolded Pael receptor (Pael-R) is a substrate of the E3 ubiquitin ligase Parkin. Accumulation of Pael-R in the endoplasmic reticulum (ER) of dopaminergic neurons induces ER stress leading to neurodegeneration. Here, we show that CHIP, Hsp70, Parkin, and Pael-R formed a complex in vitro and in vivo. The amount of CHIP in the complex was increased during ER stress. CHIP promoted the dissociation of Hsp70 from Parkin and Pael-R, thus facilitating Parkin-mediated Pael-R ubiquitination. Moreover, CHIP enhanced Parkin-mediated in vitro ubiquitination of Pael-R in the absence of Hsp70. Furthermore, CHIP enhanced the ability of Parkin to inhibit cell death induced by Pael-R. Taken together, these results indicate that CHIP is a mammalian E4-like molecule that positively regulates Parkin E3 activity.     

Aberrant tau phosphorylation by glycogen synthase kinase-3beta and JNK3 induces oligomeric tau fibrils in COS-7 cells

Neurofibrillary tangles (NFTs) are found in a wide range of neurodegenerative disorders, including Alzheimer's disease. The major component of NFTs is aberrantly hyperphosphorylated microtubule-associated protein tau. Because appropriate in vivo models have been lacking, the role of tau phosphorylation in NFTs formation has remained elusive. Here, we describe a new model in which adenovirus-mediated gene expression of tau, DeltaMEKK, JNK3, and GSK-3beta in COS-7 cells produces most of the pathological phosphorylation epitopes of tau including AT100. Furthermore, this co-expression resulted in the formation of tau aggregates having short fibrils that were detergent-insoluble and Thioflavin-S-reactive. These results suggest that aberrant tau phosphorylation by the combination of these kinases may be involved in "pretangle," oligomeric tau fibril formation in vivo.     

Examining the bases of the J3/4 domain of Escherichia coli ribonuclease P

We prepared several mutants of the J3/4 and P4 domains of Escherichia coli ribonuclease P (RNase P): A62G, A62U, G63C/G64C, A65G, A67G, U69A, U69G, U69C, U69Delta, and U69UU. Comparison of the ribozyme and holo enzyme reactions at various concentrations of magnesium ions showed that the presence of a bulge at U69 in the P4 domain was important in the holo enzyme. The results also showed that the conserved bases G63 and G64 in the J3/4 domain were important for efficient ribozyme reactions but were replaceable in the presence of the protein component. Our data showed that the bases in the J3/4 and P4 domains displayed different responses to the metal ions that were affected by the presence of the protein component.     

Mitochondrial protease Omi/HtrA2 enhances caspase activation through multiple pathways

Omi/HtrA2 is a mitochondrial serine protease that is released into the cytosol during apoptosis and promotes cytochrome c (Cyt c)dependent caspase activation by neutralizing inhibitor of apoptosis proteins (IAPs) via its IAP-binding motif. The protease activity of Omi/HtrA2 also contributes to the progression of both apoptosis and caspase-independent cell death. In this study, we found that wild-type Omi/HtrA2 is more effective at caspase activation than a catalytically inactive mutant of Omi/HtrA2 in response to apoptotic stimuli, such as UV irradiation or tumor necrosis factor. Although similar levels of Omi/HtrA2 expression, XIAP-binding activity, and Omi/HtrA2 mitochondrial release were observed among cells transfected with catalytically inactive and wild-type Omi/HtrA2 protein, XIAP protein expression after UV irradiation was significantly reduced in cells transfected with wild-type Omi/HtrA2. Recombinant Omi/HtrA2 was observed to catalytically cleave IAPs and to inactivate XIAP in vitro, suggesting that the protease activity of Omi/HtrA2 might be responsible for its IAP-inhibiting activity. Extramitochondrial expression of Omi/HtrA2 indirectly induced permeabilization of the outer mitochondrial membrane and subsequent Cyt c-dependent caspase activation in HeLa cells. These results indicate that protease activity of Omi/HtrA2 promotes caspase activation through multiple pathways.     

CIP98, a novel PDZ domain protein,is expressed in the central nervous system and interacts with calmodulin-dependent serine kinase

Receptors and various molecules in neurons are localized at precise locations to perform their respective functions, especially in synaptic sites. Among synaptic molecules, PDZ domain proteins play major roles in scaffolding and anchoring membrane proteins for efficient synaptic transmission. In the present study, we isolated CIP98, a novel protein (98 kDa) consisting of three PDZ domains and a proline-rich region, which is widely expressed in the central nervous system. In situ hybridization and immunohistochemical staining patterns demonstrate that CIP98 is expressed strongly in certain types of neurons, i.e. pyramidal cells in layers III-V of the cerebral cortex, projecting neurons in the thalamus and interneurons in the cerebellum. The results of immunocytochemical staining and electron microscopy revealed that CIP98 is localized both in dendrites and axons. Interestingly, CIP98 interacts with CASK (calmodulin-dependent serine kinase), a member of the membrane-associated guanylate kinase (MAGUK) family that plays important roles in the molecular organization of proteins at synapses. CIP98 was shown to co-localize with CASK along the dendritic processes of neurons. In view of its direct association with CASK, CIP98 may be involved in the formation of CASK scaffolding proteins complex to facilitate synaptic transmission in the CNS.