Aquatic invertebrate’s Carbohydrate-binding module assists environmental cellulase to immobilize in wetland sediments

Carbohydrate-binding modules (CBMs) are non-catalytic protein domains that bind to carbohydrates, and have been well studied in microorganisms. Endogenous CBMs in aquatic invertebrates, however, have not yet been identified, and little is known about their ecological significance to wetland environments. Using an approach of characterizing a recombinant CBM (CjCel9A) from a brackish bivalve, Corbicula japonica, this work identified CjCel9A-CBM’s cellulose-binding activity. Scatchard plot analysis in the study of CjCel9A-CBM binding to α-cellulose showed a high corresponding partitioning coefficient (Kr) of 20.33, indicating CjCel9A-CBM’s high affinity for cellulose. In addition, this affinity tolerated a high ion concentration buffer system, consistent with C. japonica’s adaption to brackish wetland environments. Moreover, immuno-scanning electron microscopy (immuno-SEM) suggested that CjCel9A-CBM binds to α-cellulose unevenly, which was further determined to be caused by its higher affinity for crystalline cellulose (Cellulose I, mostly seen in plant leaves). Together, these findings suggest that CjCel9A-CBM is capable of immobilizing its associated catalytic domain on environmental crystalline cellulose (i.e., fallen leaves) in wetland sediments. Most importantly, they could provide a reasonable answer to a question recognized broadly in wetland ecologists, namely, why many wetland sediments have constant cellulase activities, although the sediments are being washed almost every day

A Novel Method for Rearing Zebrafish by Using Freshwater Rotifers (Brachionus calyciflorus)

The zebrafish (Danio rerio) has become a powerful model organism for studying developmental processes and genetic diseases. However, there remain several problems in previous rearing methods. In this study, we demonstrate a novel method for rearing zebrafish larvae by using a new first food, freshwater rotifers (Brachionus calyciflorus). Feeding experiments indicated that freshwater rotifers are suitable as the first food for newly hatched larval fish. In addition, we revisited and improved a feeding schedule from 5 to 40 days postfertilization. Our feeding method using freshwater rotifers accelerated larval growth. At 49 dpf, one pair out of 10 pairs successfully produced six fertilized eggs. At 56, 63, and 71 dpf, 6 out of the 10 pairs constantly produced normal embryos. Our method will improve the husbandry of the zebrafish

Different effects of methylphenidate and atomoxetine on the behavior and brain transcriptome of zebrafish

2種類のADHD治療薬が対照的な作用を持つことを発見 --ゼブラフィッシュで行動や脳での働きを探る--. 京都大学プレスリリース. 2020-05-18.Attention deficit-hyperactivity disorder (ADHD) is a prevalent neuropsychiatric disorder found in children. It is characterized by inattention, hyperactivity, and impulsivity. Methylphenidate (MPH) and atomoxetine (ATX) are commonly prescribed for the treatment of ADHD. In the present study, we examined the behavioral and brain transcriptome changes in MPH-treated and ATX-treated zebrafish. In behavioral analysis, zebrafish showed opposite response to each treatment. MPH-treated fish showed higher anxiety-like behavior while ATX-treated fish showed lower anxiety-like behavior. Further, we performed RNA sequencing analysis of zebrafish brain to elucidate the underlying biological pathways associated with MPH and ATX treatment. Interestingly, we found that shared differentially expressed genes in MPH-treated and ATX-treated fish were instrumental in cholesterol biosynthesis pathway and were regulated in opposite manner. Our findings highlight the contrast between MTH and ATX, and may suggest the alterations in clinical practice for these medications and drug development for ADHD

Cooling-Sensitive TRPM8 Is Thermostat of Skin Temperature against Cooling.

We have shown that cutaneous cooling-sensitive receptors can work as thermostats of skin temperature against cooling. However, molecule of the thermostat is not known. Here, we studied whether cooling-sensitive TRPM8 channels act as thermostats. TRPM8 in HEK293 cells generated output (y) when temperature (T) was below threshold of 28.4°C. Output (y) is given by two equations: At T >28.4°C, y = 0; At T <28.4°C, y  =  -k(T - 28.4°C). These equations show that TRPM8 is directional comparator to elicits output (y) depending on negative value of thermal difference (ΔT  =  T - 28.4°C). If negative ΔT-dependent output of TRPM8 in the skin induces responses to warm the skin for minimizing ΔT recursively, TRPM8 acts as thermostats against cooling. With TRPM8-deficient mice, we explored whether TRPM8 induces responses to warm the skin against cooling. In behavioral regulation, when room temperature was 10°C, TRPM8 induced behavior to move to heated floor (35°C) for warming the sole skin. In autonomic regulation, TRPM8 induced activities of thermogenic brown adipose tissue (BAT) against cooling. When menthol was applied to the whole trunk skin at neutral room temperature (27°C), TRPM8 induced a rise in core temperature, which warmed the trunk skin slightly. In contrast, when room was cooled from 27 to 10°C, TRPM8 induced a small rise in core temperature, but skin temperature was severely reduced in both TRPM8-deficient and wild-type mice by a large heat leak to the surroundings. This shows that TRPM8-driven endothermic system is less effective for maintenance of skin temperature against cooling. In conclusion, we found that TRPM8 is molecule of thermostat of skin temperature against cooling

In vivo targeted single-nucleotide editing in zebrafish

To date, several genome editing technologies have been developed and are widely utilized in many fields of biology. Most of these technologies, if not all, use nucleases to create DNA double-strand breaks (DSBs), raising the potential risk of cell death and/or oncogenic transformation. The risks hinder their therapeutic applications in humans. Here, we show that in vivo targeted single-nucleotide editing in zebrafish, a vertebrate model organism, can be successfully accomplished with the Target-AID system, which involves deamination of a targeted cytidine to create a nucleotide substitution from cytosine to thymine after replication. Application of the system to two zebrafish genes, chordin (chd) and one-eyed pinhead (oep), successfully introduced premature stop codons (TAG or TAA) in the targeted genomic loci. The modifications were heritable and faithfully produced phenocopies of well-known homozygous mutants of each gene. These results demonstrate for the first time that the Target-AID system can create heritable nucleotide substitutions in vivo in a programmable manner, in vertebrates, namely zebrafish

Simple Prediction Method for Rubber Adhesive Friction by the Combining Friction Test and FE Analysis

In the design and development of rubber products, it is important to evaluate the contact load dependency of the friction coefficient. In particular, since the pressure distribution varies depending on the dimensions of sliding bodies and the pattern of the contact surface, a simplified and accurate evaluation method that can take these influences into account is desired. In this study, we proposed a prediction method for the adhesive friction between rubber specimens of arbitrary shapes with arbitrary roughness and a smooth hard surface, by combining the: (1) friction theory considering the influence of roughness; (2) basic friction test; and (3) finite element analysis. Further, we verified the effectiveness of the proposed method by comparing the predicted results with the measurement results of friction between a hemispherical PDMS specimen and a PMMA flat plate and between a PDMS block specimen with a grooved surface and a flat prism. Results show that the prediction accuracy of the contact load dependency of the friction coefficient is reasonably good

A novel silk-like shell matrix gene is expressed in the mantle edge of the Pacific oyster prior to shell regeneration.

During shell formation, little is known about the functions of organic matrices, especially about the biomineralization of shell prismatic layer. We identified a novel gene, shelk2, from the Pacific oyster presumed to be involved in the shell biosynthesis. The Pacific oyster has multiple copies of shelk2. Shelk2 mRNA is specifically expressed on the mantle edge and is induced during shell regeneration, thereby suggesting that Shelk2 is involved in shell biosynthesis. To our surprise, the database search revealed that it encodes a spider silk-like alanine-rich protein. Interestingly, most of the Shelk2 primary structure is composed of two kinds of poly-alanine motifs-GXNA(n)(S) and GSA(n)(S)-where X denotes Gln, Arg or no amino acid. Occurrence of common motifs of Shelk2 and spider silk led us to the assumption that shell and silk are constructed under similar strategies despite of their living environments

The Germ Cell Lineage Identified by vas-mRNA during the Embryogenesis in Goldfish

vas RNA has been identified in germ-line cells and its precursors in zebrafish, with the result that the germ-line lineage can be traced throughout embryogenesis. In the present study, we described vas localization and the migration of vas-positive cells in goldfish, using whole mount in situ hybridization. The signals of vas mRNA localization appeared at the marginal part of the first to third cleavage planes. The eight signals were detected during the period from the 8- cells to the 512-cell stage. At the late-blastula stage, additional numbers of vas-positive cells were observed, suggesting the proliferation of these cells. At the segmentation period, vas-positive cells showed a long extended distribution along the embryonic axis, but did not form any clusters. vas-positive cells were occasionally distributed at the head region, especially around the future otic vesicle. These signals were inherited to the primordial germ cells, suggesting that vas-positive cells were primordial germ cells (PGCs) in goldfish