Expression analysis of the type I keratin protein keratin 33A in goat coat hair

The coat of a goat, like that of many mammalian species, consists of an outer coat of coarse hairs and an under coat of fine, downy hairs. The coarse guard hairs are produced by primary follicles and the finer cashmere hairs by secondary follicles. We previously reported that hair keratins are components of cashmere hair, and proteomic analysis revealed that their expression is elevated in winter coat hair. To determine detailed characterization, we have cloned keratin 33A gene, a major highly expressed keratin in winter, and then analyzed the expression of goat hair coat. By Western analysis, we detected that keratin 33A protein is expressed only in hair coat among the various goat tissues. Moreover, the expression level in winter has increased in cashmere high‐producing Korean native breed, whereas the expression levels between summer and winter had not changed in cashmere low‐producing Saanen. In addition, by immunohistochemistry we determined that keratin 33A is localized in the major cortex portion of cashmere fiber. These results confirm that keratin 33A is a structural protein of goat cashmere hair fiber.     

Effects of growth factors on development of fetal islet B-cells in vitro

To investigate the role of growth factors (epidermal growth factor [EGF], betacellulin, and activin A) in the development of islet B cells of rat fetal pancreatic explants in vitro, pancreases from rat fetuses at day 18 of gestation were cultured for 96 hr, with or without these growth factors. Culture medium was changed every 24 hr, and the level of insulin released in the culture medium was measured. After 72 hr of culture, pancreases were examined histologically. As a result, EGF promoted cell proliferation, but reduced B cell volume. Whereas, betacellulin and activin A inhibited cell division, but promoted increased B cell volume and insulin secretion, especially activin A, which stimulated insulin release in a time dependent manner. These results suggest that EGF, betacellulin, and activin A promote pancreatic cell proliferation, islet B-cell differentiation, and islet B-cell differentiation and functional maturation, respectively, and that EGF, betacellulin, and activin A, in this order, regulate islet B-cell neogenesis.     

Spatial coupling of mTOR and autophagy augments secretory phenotypes

Protein synthesis and autophagic degradation are regulated in an opposite manner by mammalian target of rapamycin (mTOR), whereas under certain conditions it would be beneficial if they occurred in unison to handle rapid protein turnover. We observed a distinct cellular compartment at the trans side of the Golgi apparatus, the TOR-autophagy spatial coupling compartment (TASCC), where (auto)lysosomes and mTOR accumulated during Ras-induced senescence. mTOR recruitment to the TASCC was amino acid- and Rag guanosine triphosphatase-dependent, and disruption of mTOR localization to the TASCC suppressed interleukin-6/8 synthesis. TASCC formation was observed during macrophage differentiation and in glomerular podocytes; both displayed increased protein secretion. The spatial coupling of cells' catabolic and anabolic machinery could augment their respective functions and facilitate the mass synthesis of secretory proteins.     

The Process and Development Mechanism of Age-related Fibrosis in the Pancreatic Islets of Sprague-Dawley Rats: Immunohistochemical Detection of Myofibroblasts and Suppression Effect by Estrogen Treatment

The mechanism of spontaneous islet fibrosis in Sprague-Dawley rats was investigated. Using sections of the pancreas in naive males aged 26 to 102 weeks old and 26-week-old males injected with β-estradiol 3-benzoate (EB), the incidence of lesions and histological scores of fibrosis were examined in conjunction with immunohistochemistry for α-smooth muscle actin (α-SMA), platelet-derived growth factor receptor-α (PDGFRα) and estrogen receptor-α (ERα). The incidence of islet fibrosis increased in 78-week-old animals compared to the 26-week-old animals, and the incidence of atrophy in the fibrotic islet increased in animals over 52 weeks old. α-SMA and PDGFRα were positively stained mainly in fibrotic/inflammatory islets, and the histological score of α-SMA in the fibrotic islet decreased age-dependently. Notably, α-SMA and PDGFRα were co-expressed in inflammatory islets with a high score at all ages. The positive index of ERα in the EB-treated group increased when compared with that of the naive group. However, it was independent of the existence of fibrosis. In contrast, the score of α-SMA and PDGFRα decreased in the EB-treated group. In conclusion, it was clarified that a part of age-related fibrosis in islets became atrophy with age, and α-SMA-positive myofibroblasts were considered to contribute to the development of fibrosis. Strong PDGFRα stainability in fibrotic/inflammatory islets may imply that myofibroblasts were stimulated by PDGF to produce an extracellular matrix. Although estradiol has been known to suppress fibrosis/inflammation in the islet, nuclear-located ER-dependent signaling was considered not to be involved in the suppression mechanism. EB possibly affected the inhibition of the appearance of myofibroblasts.     

Evaluation of dry matter production and yield in early-sown wheat using near-isogenic lines for the vernalization locus Vrn-D1

ABSTRACT

Wheat (Triticum aestivum L.) grain yield is predicted to decrease in the future because of an increase in air temperature globally. To clarify the effects of the vernalization response gene in wheat to warmer winters, we compared dry matter production and grain yield between spring wheat ‘Asakazekomugi’ and its winter-type near-isogenic line (NIL) carrying different alleles of the vernalization response gene Vrn-D1 under early-, standard-, and late-sowing conditions. Under early-sowing conditions, dry matter production of the NIL carrying the winter allele of Vrn-D1, named Asa (Vrn-D1b), exceeded that of ‘Asakazekomugi’ from mid-March (after stem elongation in Asa (Vrn-D1b)) when the temperatures rose. Tiller number and leaf area index under early-sowing conditions were consistently higher in Asa (Vrn-D1b) than in ‘Asakazekomugi’ from mid-March onward. It was suggested that the early-sown ‘Asakazekomugi’ could not effectively absorb solar radiation to produce dry matter because of the acceleration of stem elongation caused by the Vrn-D1 gene during the cold season. The grain yield of Asa (Vrn-D1b) with early sowing was higher than with standard sowing. In contrast, the grain yield of ‘Asakazekomugi’ was lower in the early-sown crop than in the crop sown at the standard date. These results suggested that the higher grain yield of Asa (Vrn-D1b) than that of ‘Asakazekomugi’ under early-sown conditions could be due to Asa (Vrn-D1b) maintaining high dry matter production after the jointing stage by suppressing acceleration of growth caused by warm conditions after sowing.     

Automated analysis of fine‐root dynamics using a series of digital images

Information related to the growth of fine roots is important for understanding C allocation in trees and the mechanisms of C cycling in ecosystems. Observations using a camera or scanner embedded in the soil enabled us to obtain continuous images of fine‐root‐growth dynamics. However, these methods are still labor‐intensive because the image analysis has to be conducted manually. We developed an automated method for tracking movement or elongation of fine roots using a sequence of scanner images. We also show how data obtained with these methods can be used for calculating fine‐root behavior. Two A4‐size scanners were buried in a mixed forest in Japan and images were taken continuously from within the soil. We preprocessed these images by extracting the fine‐root area from the images and developed an automated calculation plug‐in we named A‐root for tracking growth movement of the tips of fine roots. A‐root and manual‐tracking results were compared using the same images. The results show the A‐root and manual‐tracking methods yielded similar levels of accuracy. The average growth rate of 17 fine roots tracked using the program was 0.16 mm h^–1. The observation of the direction of growth in fine roots showed the direction may be influenced by the original root's growth where the fine roots branched, distribution of soil particles, other roots, and the force of gravity. The A‐root analysis also suggested there may be an interaction between speed of growth and changes in direction of growing fine roots.     

Metabolite profiling of spinach (Spinacia oleracea L.) leaves by altering the ratio of in the culture solution

To observe the effect of different forms of N, we have applied metabolic profiling using gas chromatography mass spectrometry to evaluate the metabolite composition of spinach. The aim of the present study was to find an appropriate indicator of overall metabolic response to N source. The effect of the     ratio on spinach tissue was investigated, comparing two cultivars that differed in their ability to use N. There was wide variation in     absorption without any distinct growth inhibition in either cultivar. Statistical analysis revealed that the metabolites could be broadly divided into two types, correlating either positively or negatively with     uptake in the culture solution. Principal component analysis (PCA) was an effective tool that extracted the indexes of the effect of the ratio of N forms as PCA scores. Close relationships between metabolite profiles and     uptake were observed, indicating that metabolic profiling is able to distinguish the invisible metabolic change in mature leaves of spinach and may provide a new tool for the investigation of quality features or rhizosphere nutritional condition.     

Spatial variation in respiration from coarse woody debris in a temperate secondary broad-leaved forest in Japan

We measured the rates of respiration from snags and logs (“coarse woody debris”, CWD) of Japanese red pine (Pinus densiflora Sieb. et Zucc.) to examine the rate of decomposition and CO₂ efflux from these materials in a temperate secondary broad-leaved forest in Japan. At this site, a high quantity of CWD of P. densiflora had accumulated as a result of pine wilt disease during the 1970s. Respiration rates were measured using a dynamic closed chamber method combined with an infrared gas analyzer. We measured the respiration rate of 7 samples of snags and 10 samples of logs from August 2003 to January 2004. The responses of the respiration rates of snags (R_snag) and logs (R_log) to changing temperature were both exponential and the responses to water content were quadratic, and the same function could be used to estimate annual values of both R_snag and R_log. Intensive measurements of water contents of snags and logs showed a marked difference in water content. The mean water content of snags was 20% of log water content. This difference was likely responsible for the observed difference in annual R_snag and R_log. The decay rate constants estimated from the respiration rates measurement of snags and logs were 0.019 and 0.081 year⁻¹, respectively. Despite being lower than R_log, R_snag was a significant compartment of the CWD carbon budget at this site.     

PCR-DGGE analysis of bacterial communities in <Emphasis Type="Italic">funazushi</Emphasis>, fermented crucian carp with rice,during fermentation

Funazushi (fermented Crucian Carp with rice) is a fermented fish product found only around Lake Biwa in Shiga Prefecture, Japan. It is characterized by a unique cheese-like flavor and characteristic sour taste. We analyzed the changes in the microbial community during funazushi fermentation by denaturing gradient gel electrophoresis of PCR-amplified 16S rDNA fragments (PCR-DGGE) and by plate counts. The plate counts showed that lactic acid bacteria reached 8.0 log₁₀ CFU/g within 7 days of fermentation initiation before decreasing slowly to 4.0 log₁₀ CFU/g during the remainder of 1-year study period. PCR-DGGE revealed that the dominant bacteria in the initial (days 14 and 30) and latter (days 90, 180, and 360) periods of fermentation were Lactobacillus plantarum and L. acetotolerans. This is the first identification of L. acetotolerans in funazushi as traditional cultivation techniques have not been sufficiently sensitive. This is the first report of PCR-DGGE being used to assess the microbial community in funazushi. This technique was also found to be effective in profiling microbial diversity.