MURC, a muscle-restricted coiled-coil protein that modulates the Rho/ROCK pathway, induces cardiac dysfunction and conduction disturbance

We identified a novel muscle-restricted putative coiled-coil protein, MURC, which is evolutionarily conserved from frog to human. MURC was localized to the cytoplasm with accumulation in the Z-line of the sarcomere in the murine adult heart. MURC mRNA expression in the heart increased during the developmental process from the embryonic stage to adulthood. In response to pressure overload, MURC mRNA expression increased in the hypertrophied heart. Using the yeast two-hybrid system, we identified the serum deprivation response (SDPR) protein, a phosphatidylserine-binding protein, as a MURC-binding protein. MURC induced activation of the RhoA/ROCK pathway, which modulated serum response factor-mediated atrial natriuretic peptide (ANP) expression and myofibrillar organization. SDPR augmented MURC-induced transactivation of the ANP promoter in cardiomyocytes, and RNA interference of SDPR attenuated the action of MURC on the ANP promoter. Transgenic mice expressing cardiac-specific MURC (Tg-MURC) exhibited cardiac contractile dysfunction and atrioventricular (AV) conduction disturbances with atrial chamber enlargement, reduced thickness of the ventricular wall, and interstitial fibrosis. Spontaneous episodes of atrial fibrillation and AV block were observed in Tg-MURC mice. These findings indicate that MURC modulates RhoA signaling and that MURC plays an important role in the development of cardiac dysfunction and conduction disturbance with increased vulnerability to atrial arrhythmias.     

Effects of a novel Y5 antagonist in obese mice: combination with food restriction or sibutramine

Objective: To further address the function of the Y5 receptor in energy homeostasis, we investigated the effects of a novel spironolactone Y5 antagonist in diet-induced obese (DIO) mice. Methods and Procedures: Male C57BL/6 or Npy5r^−/− mice were adapted to high-fat (HF) diet for 6–10 months and were submitted to three experimental treatments. First, the Y5 antagonist at a dose of 10 or 30 mg/kg was administered for 1 month to DIO C57BL/6 or Npy5r^−/− mice. Second, the Y5 antagonist at 30 mg/kg was administered for 1.5 months to DIO C57BL/6 mice, and insulin sensitivity was evaluated using an insulin tolerance test. After a recovery period, nuclear magnetic resonance measurement was performed to evaluate body composition. Third, DIO mice were treated with the Y5 antagonist alone, or in combination with 10% food restriction, or with another anorectic agent, sibutramine at 10 mg/kg, for 1.5 months. Plasma glucose, insulin, and leptin levels, and adipose tissue weights were quantified. Results: The spironolactone Y5 antagonist significantly reduced body weight in C57BL DIO mice, but not in Npy5r^−/− DIO mice. The Y5 antagonist produced a fat-selective loss of body weight, and ameliorated obesity-associated insulin resistance in DIO mice. In addition, the Y5 antagonist combined with either food restriction or sibutramine tended to produce greater body weight loss, as compared with single treatment. Discussion: These findings demonstrate that the Y5 receptor is an important mediator of energy homeostasis in rodents.     

Antagonizing dopamine D1-like receptor inhibits Th17 cell differentiation: preventive and therapeutic effects on experimental autoimmune encephalomyelitis

Five types of dopamine receptors, termed D1 to D5, have been identified to date. The D1 and D5 receptors form the D1-like group that couples with the Gαs class of G proteins, while D2, D3 and D4 form the D2-like group that couples with the Gαi class of G proteins. A D2-like-receptor (D2-like-R) antagonist L750667 induced dendritic cell (DC)-mediated Th17 differentiation. In contrast, a D1-like-R antagonist SCH23390 inhibited DC-mediated Th17 differentiation. The D1-like-Rs were expressed on both DCs and T cells, whereas D2-like-Rs were marginally expressed on CD4⁺CD45RA⁺ naïve T cells. In addition, SCH23390 had the ability to prevent experimental autoimmune encephalomyelitis (EAE) in mice. Spleen cells from EAE mice showed decreased IL-17 production, when SCH23390 was administered. Adoptive transfer of DCs treated with SCH23390 successfully prevented EAE. These findings indicate that antagonizing D1-like-Rs on DCs inhibits Th17 differentiation, thereby leading to an amelioration of EAE.     

Identification of proteins encoded in Escherichia coli hydA, hydB and analysis of the hydA locus

The hydB gene of Escherichia coli, which is related with the expression of hydrogenase activity, was cloned into the plasmid (pES1). Using the maxicell protein-labeling method, the molecular weight of hydB gene product was estimated. Comparing between the gene products from the mutant strains and that of the hydB genes cloned strains, the molecular weight of the gene product was 35,000 Mr. Similarly, the molecular weight of the gene product of hydA, which had been previously cloned, was determined by maxicell analysis. The molecular weight of hydA gene product was estimated to be 80,000 Mr. Using deletion analysis and Tn1000 insertional inactivation of hydA's function, the hydA coding region was estimated between 2.2 kb and 2.8 kb in a 3.1 kb EcoRI-MluI fragment on the recombinant plasmid pEH3.     

Essential Elements as Biomarkers of Acute Kidney Injury and Spontaneous Reversion

Acute kidney injury (AKI) is an important health problem and can be caused by number of factors. The use of aminoglycosides, such as gentamicin, is one of these factors. Recently, an effort has been made to find biomarkers to guide treatment protocols. Inductively coupled plasma optical emission spectroscopy (ICP-OES) was used to estimate the contents of Ca, Cu, Fe, K, Mg, Mn, Na, P, and Zn in serum and urine of the healthy, AKI, and spontaneous recovery (SR) groups of animals. The animal model of AKI and SR was validated by measuring serum and urinary urea and creatinine. The quantitative determination of the elements showed a decrease in serum levels of Ca, and Fe in the AKI group (P<0.01 vs. healthy), with a return to normal levels in the SR group, without a significant difference between the healthy and SR groups. In the urine samples, there was a decrease in P and Na levels in the AKI group (P<0.001 and P<0.01 vs. healthy), but Ca levels were increased in this group compared with the healthy and SR groups (P<0.01). These findings indicate that mineral elements might be useful as biomarkers for AKI.     

Vasopressin Stimulates the Induction of Heat Shock Protein 27 and αB-Crystallin via Protein Kinase C Activation in Vascular Smooth Muscle Cells

In the present study, we examined the effect of vasopressin on the induction of the low-molecular-weight heat shock proteins heat shock protein 27 (HSP27) and αB-crystallin in an aortic smooth muscle cell line, A10 cells. Vasopressin induced a time-dependent accumulation of HSP27 and αB-crystallin. The stimulatory effects of vasopressin were dose-dependent over the range 0.1 nmol/L to 0.1 μmol/L. The EC₅₀values for vasopressin were 2 (HSP27) and 4 nmol/L (αB-crystallin). Vasopressin induced increases in the levels of the mRNAs for HSP27 and αB-crystallin. 12-O-Tetradecanoylphorbol 13-acetate (TPA), a protein kinase C (PKC)-activating phorbol ester, induced an accumulation of HSP27 (EC₅₀, 20 nmol/L) and αB-crystallin (EC₅₀, 2 nmol/L). In contrast, 4α-phorbol 12,13-didecanoate, a non-PKC-activating phorbol ester, had no such effect. Staurosporine and calphostin C, inhibitors of PKC, significantly reduced the vasopressin-induced accumulation of HSP27 and αB-crystallin as well as that induced by TPA. BAPTA/AM and TMB-8, inhibitors of intracellular Ca²⁺mobilization, significantly reduced the vasopressin-induced accumulation of HSP27 and αB-crystallin. These results strongly suggest that vasopressin stimulates the induction of HSP27 and αB-crystallin via PKC activation in vascular smooth muscle cells and that this effect of vasopressin is dependent on intracellular Ca²⁺mobilization.     

Extrinsic proteins of photosystem II: an intermediate member of PsbQ protein family in red algal PS II.

The oxygen-evolving photosystem II (PS II) complex of red algae contains four extrinsic proteins of 12 kDa, 20 kDa, 33 kDa and cyt c-550, among which the 20 kDa protein is unique in that it is not found in other organisms. We cloned the gene for the 20-kDa protein from a red alga Cyanidium caldarium. The gene consists of a leader sequence which can be divided into two parts: one for transfer across the plastid envelope and the other for transfer into thylakoid lumen, indicating that the gene is encoded by the nuclear genome. The sequence of the mature 20-kDa protein has low but significant homology with the extrinsic 17-kDa (PsbQ) protein of PS II from green algae Volvox Carteri and Chlamydomonas reinhardtii, as well as the PsbQ protein of higher plants and PsbQ-like protein from cyanobacteria. Cross-reconstitution experiments with combinations of the extrinsic proteins and PS IIs from the red alga Cy. caldarium and green alga Ch. reinhardtii showed that the extrinsic 20-kDa protein was functional in place of the green algal 17-kDa protein on binding to the green algal PS II and restoration of oxygen evolution. From these results, we conclude that the 20-kDa protein is the ancestral form of the extrinsic 17-kDa protein in green algal and higher plant PS IIs. This provides an important clue to the evolution of the oxygen-evolving complex from prokaryotic cyanobacteria to eukaryotic higher plants. The gene coding for the extrinsic 20-kDa protein was named psbQ' (prime).     

Induction by ozone of ethylene production and an ACC oxidase cDNA in rice (Oryza sativa L.) leaves

Exposure to ozone at 1 µl l^–1 for 6 h induced ethylene production in rice (Oryza sativa L. cv. Hitomebore) leaves. The stimulation of ethylene production was detectable 2 h after the start of the exposure to ozone, and lasted for 6 h after the exposure. A 429-bp cDNA fragment encoding ACC oxidase was obtained by RT-PCR from ozone-treated rice leaves. Its nucleotide sequence and deduced amino-acid sequence had 97.2% and 94.4% identity, respectively, to those of OS1A1COX, which was previously obtained from deepwater rice. The abundance of the cDNA increased in accordance with the induction of ethylene production by the exposure to ozone.