Upregulation of cAMP is a new functional signal pathway of Klotho in endothelial cells

We measured angiotensin I-converting enzyme (ACE) activity in a human endothelial cell to characterize the intracellular signal pathways of Klotho. COS-1 cells transfected with naked mouse membrane-form klotho plasmid DNA (pCAGGS-klotho) translated proper Klotho protein. This translated Klotho protein was secreted into the culture medium. Furthermore, ACE activity in human umbilical vein endothelial cells (HUVEC) was upregulated when HUVEC were co-cultured with COS-1 cells that were pre-transfected with pCAGGS-klotho. The conditioned medium from COS-1 cells pre-transfected with pCAGGS-klotho also dose-dependently upregulated ACE in HUVEC. In addition, the conditioned medium induced time- and dose-dependent enhancement of cAMP production in HUVEC. Rp-cAMP, an inhibitor of cAMP-dependent protein kinase A (PKA), inhibited the upregulation of ACE by Klotho protein. Our results suggest that mouse membrane-form Klotho protein acts as a humoral factor to increase ACE activity in HUVEC via a cAMP-PKA-dependent pathway. These findings may provide a new insight into the mechanism of Klotho protein.     

Effects of docosahexaenoic acid on in vitro amyloid beta peptide 25–35 fibrillation

Amyloid β peptide_25–35 (Aβ_25–35) encompasses one of the neurotoxic domains of full length Aβ_1–40/42, the major proteinaceous component of amyloid deposits in Alzheimer's disease (AD). We investigated the effect of docosahexaenoic acid (DHA, 22:6, n-3), an essential brain polyunsaturated fatty acid, on the in vitro fibrillation of Aβ_25–35 and found that it significantly reduced the degree of fibrillation, as shown by a decrease in the intensity of both the thioflavin T and green fluorescence in confocal microscopy. Transmission electron microscopy revealed that DHA-incubated samples were virtually devoid of structured fibrils but had an amorphous-like consistency, whereas the controls contained structured fibers of various widths and lengths. The in vitro fibrillation of Aβ_25–35 appeared to be pH-dependent, with the strongest effect seen at pH 5.0. DHA inhibited fibrillation at all pHs, with the strongest effect at pH 7.4. It also significantly decreased the levels of Aβ_25–35 oligomers. Nonreductive gradient gel electrophoresis revealed that the molecular size of the oligomers of Aβ_25–35 was 10 kDa (equivalent to decamers of Aβ_25–35) and that DHA dose-dependently reduced these decamers. These results suggest that DHA decreases the in vitro fibrillation of Aβ_25–35 by inhibiting the oligomeric amyloid species and, therefore, Aβ_25–35-related neurotoxicity or behavioral impairment could be restrained by DHA.     

Brief Communication: Orphaned male Chimpanzees die young even after weaning

If a social‐living animal has a long life span, permitting different generations to co‐exist within a social group, as is the case in many primate species, it can be beneficial for a parent to continue to support its weaned offspring to increase the latter's survival and/or reproductive success. Chimpanzees have an even longer period of dependence on their mothers' milk than do humans, and consequently, offspring younger than 4.5–5 years old cannot survive if the mother dies. Most direct maternal investments, such as maternal transportation of infants and sharing of night shelters (beds or nests), end with nutritional weaning. Thus, it had been assumed that a mother's death was no longer critical to the survival of weaned offspring, in contrast to human children, who continue to depend on parental care long after weaning. However, in theory at least, maternal investment in a chimpanzee son after weaning could be beneficial because in chimpanzees' male‐philopatric society, mother and son co‐exist for a long time after the offspring's weaning. Using long‐term demographic data for a wild chimpanzee population in the Mahale Mountains, Tanzania, we show the first empirical evidence that orphaned chimpanzee sons die younger than expected even if they lose their mothers after weaning. This suggests that long‐lasting, but indirect, maternal investment in sons continues several years after weaning and is vital to the survival of the sons. The maternal influence on males in the male‐philopatric societies of hominids may be greater than previously believed. Am J Phys Anthropol, 153:139–143, 2014. © 2013 Wiley Periodicals, Inc.     

Cloning and Characterization of Two Novel Genes, cry24B and s1orf2, from a Mosquitocidal Strain of Bacillus thuringiensis serovar sotto

Two new crystal protein genes, cry24B and s1orf2, were cloned from a mosquitocidal Bacillus thuringiensis serovar sotto strain. The cry24B and s1orf2 genes encoded a 76-kDa and 62-kDa protein, respectively. The Cry24B protein retained five conserved regions commonly found in the existing Cry proteins. The amino acid sequence of the S1ORF2 had a high homology to that of the ORF2 protein of B. thuringiensis serovar jegathesan. Southern hybridization experiments with a cry24B gene-specific probe revealed that these genes are located on two large plasmids of > 100 kb. When the two genes, cry24B and s1orf2, were expressed in an acrystalliferous B. thuringiensis host, the proteins were synthesized and accumulated as inclusions. These inclusions exhibited no larvicidal activities against three mosquito species: Aedes aegypti, Anopheles stephensi, and Culex pipiens molestus. Likewise, the inclusions contained no cytocidal activity against HeLa cells.     

In situ observation of a cell adhesion and metabolism using surface infrared spectroscopy

In this study, we report on an in situ monitoring system of living cultured cells using infrared absorption spectroscopy in the geometry of multiple internal reflections (MIR-IRAS). In order to observe living cultured cells, the temperature in the sample chamber of a FT-IR spectrometer was maintained at 37 °C and a humidified gas mixture containing 5% CO₂ was introduced into the sample chamber. Human breast cell line MCF-7 cultured on Si MIR prisms were placed in the sample chamber and infrared spectra of MCF-7 cells were collected for 5 h. It was found that the adhesion and metabolism of MCF-7 cells could be monitored by the absorption intensity of amide-II protein band (1,545 cm⁻¹) and also by the absorption intensities of CH _x bands (2,700–3,100 cm⁻¹). These results suggest that our system is useful for a nondestructive and non-label monitoring of cell viability. Our method based on infrared absorption spectroscopy has a potential for bioscreening application.     

cDNA microarray analyses reveal candidate marker genes for the detection of ascidian disease in Korea

A serious disease of the ascidian Halocynthia roretzi has been spread extensively among Korean aquaculture sites. To reveal the cause of the disease and establish a monitoring system for it, we constructed a cDNA microarray spotted with 2,688 cDNAs derived from H. roretzi hemocyte cDNA libraries to detect genes differentially expressed in hemocytes between diseased and non-diseased ascidians. We detected 21 genes showing increased expression and 16 genes showing decreased expression in hemocytes from diseased ascidians compared with those from non-diseased ascidians. RT-PCR analyses confirmed that the expression levels of genes encoding astacin, lysozyme, ribosomal protein PO, and ubiquitin-ribosomal protein L40e fusion protein were increased in hemocytes from diseased ascidians, while those of genes encoding HSP40, HSP70, fibronectin, carboxypeptidase and lactate dehydrogenase were decreased. These genes were expressed not only in hemocytes but also in various other tissues in ascidians. Furthermore, the expression of glutathione-S transferase omega, which is known to be up-regulated in H. roretzi hemocytes during inflammatory responses, was strongly increased in hemocytes from diseased ascidians. These gene expression profiles suggest that immune and inflammatory reactions occur in the hemocytes of diseased ascidians. These genes will be good markers for detecting and monitoring this disease of ascidians in Korean aquaculture sites.     

High reproductive rates result in high predation risks: a mechanism promoting the coexistence of competing prey in spatially structured populations

I tested the hypothesis that spatial structure provides a trade-off between reproduction and predation risk and thereby facilitates predator-mediated coexistence of competing prey species. I compared a cellular automata model to a mean-field model of two prey species and their common predator. In the mean-field model, the prey species with the higher reproductive rate (the superior competitor) always outcompeted the other species (the inferior competitor), both in the presence of and the absence of the predator. In the cellular automata model, both prey species, which differed only in their reproductive rates, coexisted for a long time in the presence of their common predator at intermediate levels of predation. At low predation rates, the superior competitor dominated, while high predation rates favored the inferior competitor. This discrepancy in the results of the different models was due to a trade-off that spontaneously emerged in spatially structured populations; that is, the more clustered distribution of the superior competitor made it more susceptible to predation. In addition, coexistence of competing prey species declined with increasing dispersal ranges of either prey or predator, which suggests that the trade-off that results from spatial structure becomes less important as either prey or predator disperse over a broader range.     

Sensing of cytoplasmic pH by bacterial chemoreceptors involves the linker region that connects the membrane-spanning and the signal-modulating helices.

The two major chemoreceptors of Escherichia coli, Tsr and Tar, mediate opposite responses to the same changes in cytoplasmic pH (pH(i)). We set out to identify residues involved in pH(i) sensing to gain insight into the general mechanisms of signaling employed by the chemoreceptors. Characterization of various chimeras of Tsr and Tar localized the pH(i)-sensing region to Arg(259)-His(267) of Tar and Gly(261)-Asp(269) of Tsr. This region of Tar contains three charged residues (Arg(259)-Ser(261), Asp(263), and His(267)) that have counterparts of opposite charge in Tsr (Gly(261)-Glu(262), Arg(265), and Asp(269)). The replacement of all of the three charged residues in Tar or Arg(259)-Ser(260) alone by the corresponding residues of Tsr reversed the polarity of pH(i) response, whereas the replacement of Asp(263) or His(267) did not change the polarity but altered the time course of pH(i) response. These results suggest that the electrostatic properties of a short cytoplasmic region within the linker region that connects the second transmembrane helix to the first methylation helix is critical for switching the signaling state of the chemoreceptors during pH sensing. Similar conformational changes of this region in response to external ligands may be critical components of transmembrane signaling.