Pyrindicin,a New Alkaloid from a Streptomyces Strain

In the course of our examination for the alkaloid productivities of Streptomyces strains, Streptomyces sp. NA–15 was found to produce a new alkaloid, pyrindicin, in the culture medium. The strain NA–15 was found to be a variant of Streptomyces griseoflavus and was designated as S. griseoflavus var. pyr indie us nov. var.

After the culture conditions for pyrindicin production were studied, pyrindicin was obtained as its hydrochloride (mp 145°C, decomp.) from the cultured broth. The compound was shown to possess weak antimicrobial and several pharmacological activities. The LD₅₀ of the hydrochloride (ip, in mice) was 87 mg/kg.     

Responses of carbon and oxygen stable isotopes in rice grain (Oryza sativa L.) to an increase in air temperature during grain filling in the Japanese archipelago

Stable isotopic compositions of carbon (δ¹³C) and oxygen (δ¹⁸O) in plants reflect growth conditions. Therefore, these isotopes might be good indicators of changes in environmental factors, such as variations in air temperature caused by climate change. It is predicted that climate change will lead to a greater increase in minimum air temperatures (primarily during the night) than in maximum air temperatures (primarily during the day) in many parts of Japan. In the present study, we investigated whether the δ¹³C and δ¹⁸O of the rice grain Koshihikari (Oryza sativa L.) from the northern latitudes (30.49°–37.14°) of Japan reflect variations in air temperature during grain filling and are related to the yield and proportion of first-grade rice (<15 % transparency, roundness, and cracking) as an indicator of quality. We revealed that rice δ¹³C was not correlated with mean maximum or minimum air temperatures for each prefecture. By contrast, rice δ¹⁸O was positively correlated with mean minimum air temperature, suggesting that rice δ¹⁸O reflects changes in night air temperature. We further showed that an increase in the mean minimum air temperature during grain filling had a negative effect on rice yield and quality. Our findings indicate that the δ¹⁸O of rice grain may be a good indicator of physiological changes in response to minimum air temperatures during grain filling.     

Effects of Metformin on Burn-Induced Hepatic Endoplasmic Reticulum Stress in Male Rats

Severe burn injury causes hepatic dysfunction that results in major metabolic derangements including insulin resistance and hyperglycemia and is associated with hepatic endoplasmic reticulum (ER) stress. We have recently shown that insulin reduces ER stress and improves liver function and morphology; however, it is not clear whether these changes are directly insulin mediated or are due to glucose alterations. Metformin is an antidiabetic agent that decreases hyperglycemia by different pathways than insulin; therefore, we asked whether metformin affects postburn ER stress and hepatic metabolism. The aim of the present study is to determine the effects of metformin on postburn hepatic ER stress and metabolic markers. Male rats were randomized to sham, burn injury and burn injury plus metformin and were sacrificed at various time points. Outcomes measured were hepatic damage, function, metabolism and ER stress. Burn-induced decrease in albumin mRNA and increase in alanine transaminase (p < 0.01 versus sham) were not normalized by metformin treatment. In addition, ER stress markers were similarly increased in burn injury with or without metformin compared with sham (p < 0.05). We also found that gluconeogenesis and fatty acid metabolism gene expressions were upregulated with or without metformin compared with sham (p < 0.05). Our results indicate that, whereas thermal injury results in hepatic ER stress, metformin does not ameliorate postburn stress responses by correcting hepatic ER stress.     

WNT10B functional dualism: beta-catenin/Tcf-dependent growth promotion or independent suppression with deregulated expression in cancer

We found aberrant DNA methylation of the WNT10B promoter region in 46% of primary hepatocellular carcinoma (HCC) and 15% of colon cancer samples. Three of 10 HCC and one of two colon cancer cell lines demonstrated low or no expression, and 5-aza-2'deoxycytidine reactivated WNT10B expression with the induction of demethylation, indicating that WNT10B is silenced by DNA methylation in some cancers, whereas WNT10B expression is up-regulated in seven of the 10 HCC cell lines and a colon cancer cell line. These results indicate that WNT10B can be deregulated by either overexpression or silencing in cancer. We found that WNT10B up-regulated beta-catenin/Tcf activity. However, WNT10B-overexpressing cells demonstrated a reduced growth rate and anchorage-independent growth that is independent of the beta-catenin/Tcf activation, because mutant beta-catenin-transduced cells did not suppress growth, and dominant-negative hTcf-4 failed to alleviate the growth suppression by WNT10B. Although WNT10B expression alone inhibits cell growth, it acts synergistically with the fibroblast growth factor (FGF) to stimulate cell growth. WNT10B is bifunctional, one function of which is involved in beta-catenin/Tcf activation, and the other function is related to the down-regulation of cell growth through a different mechanism. We suggest that FGF switches WNT10B from a negative to a positive cell growth regulator.     

The TRK-Fused Gene Is Mutated in Hereditary Motor and Sensory Neuropathy with Proximal Dominant Involvement

Hereditary motor and sensory neuropathy with proximal dominant involvement (HMSN-P) is an autosomal-dominant neurodegenerative disorder characterized by widespread fasciculations, proximal-predominant muscle weakness, and atrophy followed by distal sensory involvement. To date, large families affected by HMSN-P have been reported from two different regions in Japan. Linkage and haplotype analyses of two previously reported families and two new families with the use of high-density SNP arrays further defined the minimum candidate region of 3.3 Mb in chromosomal region 3q12. Exome sequencing showed an identical c.854C>T (p.Pro285Leu) mutation in the TRK-fused gene (TFG) in the four families. Detailed haplotype analysis suggested two independent origins of the mutation. Pathological studies of an autopsied patient revealed TFG- and ubiquitin-immunopositive cytoplasmic inclusions in the spinal and cortical motor neurons. Fragmentation of the Golgi apparatus, a frequent finding in amyotrophic lateral sclerosis, was also observed in the motor neurons with inclusion bodies. Moreover, TAR DNA-binding protein 43 kDa (TDP-43)-positive cytoplasmic inclusions were also demonstrated. In cultured cells expressing mutant TFG, cytoplasmic aggregation of TDP-43 was demonstrated. These findings indicate that formation of TFG-containing cytoplasmic inclusions and concomitant mislocalization of TDP-43 underlie motor neuron degeneration in HMSN-P. Pathological overlap of proteinopathies involving TFG and TDP-43 highlights a new pathway leading to motor neuron degeneration.     

The Mutant Form of Acetolactate Synthase Genomic DNA from Rice is an Efficient Selectable Marker for Genetic Transformation

The proper use of a marker gene in a transformation process is critical for the production of transgenic plants. However, consumer concerns and regulatory requirements raise an objection to the presence of exogenous DNA in transgenic plants, especially antibiotic-resistant genes and promoters derived from viruses. One approach to overcome this problem is the elimination of marker genes from the plant genome by using several site-specific recombination systems. We propose an alternative method to solve this problem using a marker gene exclusively derived from the host plant DNA. We cloned a genomic DNA fragment containing regulatory and coding sequences of acetolactate synthase (ALS) gene from rice, and mutagenized the ALS gene into a herbicide-resistant form. After transfer of this construct to the rice genome, transgenic plants were efficiently selected with a herbicide, bispyribac-sodium salt, which inhibits the activity of wild type ALS. We also analyzed the regulatory feature of the rice ALS gene promoter with the gusA reporter gene and revealed that GUS expression was observed constitutively in aerial parts of rice seedlings and root tips. The marker system consisted exclusively of host plant DNA and enabled efficient selection in a monocot crop plant, rice. The selection system can potentially be applied to generate transgenic plants of other crop species and can be expected to be publicly acceptable.     

Constitutive expression of human lactoferrin and its N-lobe in rice plants to confer disease resistance.

The milk protein, lactoferrin, is known to have antibacterial, antiviral, and antifungal activities. To explore the possibility of conferring disease resistance in plants by expressing this protein, the gene for the full-length human lactoferrin (HLF), as well as the N-lobe, the N-terminal half molecule (HLFN), was introduced into rice plants and expressed constitutively under the control of the cauliflower mosaic virus 35S promotor. Western blot analysis of leaves from HLF-transgenic rice plants showed an 80 kDa-band, which was about 1-2 kDa less than human milk lactoferrin. HLFN was expressed as a 45-kDa protein and retained its heparin-binding property. Deglycosylation experiments suggested that both proteins produced by the plants had plant-type oligosaccharide chains. The transgenic rice plants were assessed for resistance against disease-causing bacteria, virus, and fungi. Of the pathogens tested, significant resistance against Burkholderia (Pseudomonas) plantarii, the causative agent of bacterial seedling blight disease, was observed in the transgenic plants expressing HLF or HLFN.     

Complete Sequence and Gene Organization of the Genome of a Hyper-thermophilic Archaebacterium, Pyrococcus horikoshii OT3

The complete sequence of the genome of a hyper-thermophilicarchaebacterium, Pyrococcus horikoshii OT3, has been determinedby assembling the sequences of the physical map-based contigsof fosmid clones and of long polymerase chain reaction (PCR)products which were used for gap-filling. The entire lengthof the genome was 1,738,505 bp. The authenticity of the entiregenome sequence was supported by restriction analysis of longPCR products, which were directly amplified from the genomicDNA. As the potential protein-coding regions, a total of 2061open reading frames (ORFs) were assigned, and by similaritysearch against public databases, 406 (19.7%) were related togenes with putative function and 453 (22.0%) to the sequencesregistered but with unknown function. The remaining 1202 ORFs(58.3%) did not show any significant similarity to the sequencesin the databases. Sequence comparison among the assigned ORFsin the genome provided evidence that a considerable number ofORFs were generated by sequence duplication. By similarity search,11 ORFs were assumed to contain the intein elements. The RNAgenes identified were a single 16S-23S rRNA operon, two 5S rRNAgenes and 46 tRNA genes including two with the intron structure.All the assigned ORFs and RNA coding regions occupied 91.25%of the whole genome. The data presented in this paper are availableon the internet at http://www.nite.go.jp.