Identification and growth of Leuconostoc lactis SHO-54, producing high amounts of NAD-specific 6-phosphogluconate dehydrogenase

The G+C content of the DNA of strain SHO-54 which produces a large amount of NAD-specific 6-phosphogluconate dehydrogenase was 41.0 mol%. The extent of the DNA–DNA homology between this strain and Leuconostoc lactis NRIC 1540 ranged from 72.8% to 95.5%. A minimal medium in which strain SHO-54 grew well was determined, and a high activity of the enzyme (410 nkat/ml) was obtained.     

Sugar composition of the polysaccharide portion of lipopolysaccharides isolated from non-O1 Vibrio cholerae O2 to O41, O44, and O68

The sugar composition of the polysaccharide portion of lipopolysaccharides (LPS) was determined for 42 serovars of non-O1 Vibrio cholerae, i.e., from serogroups O2 to O41, O44, and O68. On the basis of their compositional sugar pattern, they were classified into 24 chemotypes. 2-Keto-3-deoxyoctonate (KDO) was totally undetectable by the conventional color test (Weissbach's reaction) under the conventional hydrolysis conditions. Instead, a kind of KDO-like substance, which was positive in the reaction but not identical to KDO, was found in serogroup O19. Fructose, a characteristic sugar constituent of O1 V. cholerae LPS, was found in 33 serogroups but was absent from nine serogroups, approximately 20% of the members of this group so far examined.     

Avoiding entry into intracellular protein degradation pathways by signal mutations increases protein secretion in Pichia pastoris

In our previous study, we serendipitously discovered that protein secretion in the methylotrophic yeast Pichia pastoris is enhanced by a mutation (V50A) in the mating factor alpha (MFα) prepro-leader signal derived from Saccharomyces cerevisiae. In the present study, we investigated 20 single-amino-acid substitutions, including V50A, located within the MFα signal peptide, indicating that V50A and several single mutations alone provided significant increase in production of the secreted proteins. In addition to hydrophobicity index analysis, both an unfolded protein response (UPR) biosensor analysis and a microscopic observation showed a clear difference on the levels of UPR induction and mis-sorting of secretory protein into vacuoles among the wild-type and mutated MFα signal peptides. This work demonstrates the importance of avoiding entry of secretory proteins into the intracellular protein degradation pathways, an observation that is expected to contribute to the engineering of strains with increased production of recombinant secreted proteins.     

Effects of advance regeneration and germinated seedlings on forest recovery after clearcutting of Chamaecyparis obtusa plantations in Southern Kyushu,Southwestern Japan

We clarified the effect of advance regeneration on natural forest recovery after clearcutting of conifer plantations over the mid-term observation period (12–14 years to allow formation of the forest canopy). We established understory-intact plot (UI-plot), where advance regeneration (AR; DBH more than 1 cm before clearcutting) were retained, and understory-cleared plot (UC-plot), where all ARs were removed at felling of conifer crops. We surveyed individual density, species richness, and tree height with reference to different regeneration origins 1 year after and 12 or 14 years after clearcutting. Then, we compared the structural changes in UI-plot with that in UC-plot. The resprouted AR in UC-plot has provided the same effect as the retained AR in UI-plot in forming their height distribution patterns and stratification over a mid-term recovery process. We also found that individual density and species richness attributed by the individuals of newly established seedlings after clearcutting (SE), have contributed to the stand development in the both plots. Furthermore, SE was found to have a large contribution to forest recovery in terms of tree density and species number, while its significance for the recovery of gravity-dispersal, lucidophyllous trees was limited. We concluded that the advantage of AR retention is limited to the fast recovery of forest structure at the early stage after clearcutting, and mostly disappeared over a mid-term observation period as far as abundant advance regeneration was sufficiently accumulated and surrounding seed sources were effectively secured.

     

Structural Insights into the Substrate Specificity Switch Mechanism of the Type III Protein Export Apparatus

1. Download : Download high-res image (420KB)
2. Download : Download full-size image

     

Structure-activity relationship of prenyl-substituted polyphenols from Artocarpus heterophyllus as inhibitors of melanin biosynthesis in cultured melanoma cells

A series of prenylated, flavone-based polyphenols, compounds 1-8, were isolated from the wood of Artocarpus heterophyllus. These compounds, which have previously been shown not to inhibit tyrosinase activity, were found to be active inhibitors of the in vivo melanin biosynthesis in B16 melanoma cells, with little or no cytotoxicity. To clarify the structural requirement for inhibition, some structure-activity relationships were studied, in comparison with related compounds lacking prenyl side chains. Our experiments indicate that both prenyl and OH groups, as well as the type of substitution pattern, are crucial for the inhibition of melanin production in B16 melanoma cells.     

Expression and physiological role of CCN4/Wnt-induced secreted protein 1 mRNA splicing variants in chondrocytes

CCN4/Wnt-induced secreted protein 1 (WISP1) is one of the CCN (CTGF/Cyr61/Nov) family proteins. CCN members have typical structures composed of four conserved cysteine-rich modules and their variants lacking certain modules, generated by alternative splicing or gene mutations, have been described in various pathological conditions. Several previous reports described a CCN4/WISP1 variant (WISP1v) lacking the second module in a few malignancies, but no information concerning the production of WISP1 variants in normal tissue is currently available. The expression of CCN4/WISP1 mRNA and its variants were analyzed in a human chondrosarcoma-derived chondrocytic cell line, HCS-2/8, and primary rabbit growth cartilage (RGC) chondrocytes. First, we found WISP1v and a novel variant of WISP1 (WISP1vx) to be expressed in HCS-2/8, as well as full-length WISP1 mRNA. This new variant was lacking the coding regions for the second and third modules and a small part of the first module. To monitor the expression of CCN4/WISP1 mRNA along chondrocyte differentiation, RGC cells were cultured and sampled until they were mineralized. As a result, we identified a WISP1v ortholog in normal RGC cells. Interestingly, the WISP1v mRNA level increased dramatically along with terminal differentiation. Furthermore, overexpression of WISP1v provoked expression of an alkaline phosphatase gene that is a marker of terminal differentiation in HCS-2/8 cells. These findings indicate that WISP1v thus plays a critical role in chondrocyte differentiation toward endochondral ossification, whereas HCS-2/8-specific WISP1vx may be associated with the transformed phenotypes of chondrosarcomas.