Site-directed spin labeling electron paramagnetic resonance study of the calcium-induced structural transition in the N-domain of human cardiac troponin C complexed with troponin I

Calcium-induced structural transition in the amino-terminal domain of troponin C (TnC) triggers skeletal and cardiac muscle contraction. The salient feature of this structural transition is the movement of the B and C helices, which is termed the "opening" of the N-domain. This movement exposes a hydrophobic region, allowing interaction with the regulatory domain of troponin I (TnI) as can be seen in the crystal structure of the troponin ternary complex [Takeda, S., Yamashita, A., Maeda, K., and Maeda, Y. (2003) Nature 424, 35-41]. In contrast to skeletal TnC, Ca(2+)-binding site I (an EF-hand motif that consists of an A helix-loop-B helix motif) is inactive in cardiac TnC. The question arising from comparisons with skeletal TnC is how both helices move according to Ca(2+) binding or interact with TnI in cardiac TnC. In this study, we examined the Ca(2+)-induced movement of the B and C helices relative to the D helix in a cardiac TnC monomer state and TnC-TnI binary complex by means of site-directed spin labeling electron paramagnetic resonance (EPR). Doubly spin-labeled TnC mutants were prepared, and the spin-spin distances were estimated by analyzing dipolar interactions with the Fourier deconvolution method. An interspin distance of 18.4 A was estimated for mutants spin labeled at G42C on the B helix and C84 on the D helix in a Mg(2+)-saturated monomer state. The interspin distance between Q58C on the C helix and C84 on the D helix was estimated to be 18.3 A under the same conditions. Distance changes were observed by the addition of Ca(2+) ions and the formation of a complex with TnI. Our data indicated that the C helix moved away from the D helix in a distinct Ca(2+)-dependent manner, while the B helix did not. A movement of the B helix by interaction with TnI was observed. Both Ca(2+) and TnI were also shown to be essential for the full opening of the N-domain in cardiac TnC.     

Interaction between methyl CpG-binding protein and ran GTPase during cell division in tobacco cultured cells

BACKGROUND AND AIMS: Methyl CpG-binding proteins are considered to play critical roles in epigenetic control of gene expression by recognizing and interacting with 5-methylcytosine (m(5)C) in eukaryotes. However, among 13 corresponding genes in Arabidopsis thaliana, designated as featuring a methyl-binding domain (MBD), only four have so far been shown actually to bind to m(5)C. One example, AtMBD5, was selected here to screen for interacting proteins. METHODS: Yeast two-hybrid assays were used for screening, and physical interaction was confirmed by pull-down and bimolecular fluorescence complementation (BiFC) assays. Cellular localization was analysed by fluorescence-tagged fusion proteins using tobacco (Nicotiana tabacum) cultured bright yellow 2 cells. KEY RESULTS: A gene finally identified was found to encode AtRAN3, a protein that belongs to the Ran GTPase family, which plays a critical role in nucleocytoplasmic transport and spindle bipolarization during cell division. AtMBD5 and AtRAN3 were clearly shown to interact in the nucleus by BiFC. On co-expression of AtMBD5-cyan fluorescence protein and yellow fluorescence protein-AtRAN3 in tobacco cells, both localized to the nucleus in the resting stage, migrating to the cytoplasm, primarily around chromatin, during mitosis, particularly at metaphase. CONCLUSIONS: These results suggest that AtMBD5 becomes localized to the vicinity of chromosomes with the aid of AtRAN3 during cell division, and may play an important role not only in maintenance of chromatin structures by binding to m(5)C, but also in progress through mitosis by detaching from m(5)C. The present findings also shed light on the physiological function of Ran GTPases, direct target proteins of which have not thus far been well defined, suggesting their key role in chromatin movements in plant cells.     

Adult Mortality Attributable to Preventable Risk Factors for Non-Communicable Diseases and Injuries in Japan: A Comparative Risk Assessment

Background

The population of Japan has achieved the longest life expectancy in the world. To further improve population health, consistent and comparative evidence on mortality attributable to preventable risk factors is necessary for setting priorities for health policies and programs. Although several past studies have quantified the impact of individual risk factors in Japan, to our knowledge no study has assessed and compared the effects of multiple modifiable risk factors for non-communicable diseases and injuries using a standard framework. We estimated the effects of 16 risk factors on cause-specific deaths and life expectancy in Japan.

Methods and Findings

We obtained data on risk factor exposures from the National Health and Nutrition Survey and epidemiological studies, data on the number of cause-specific deaths from vital records adjusted for ill-defined codes, and data on relative risks from epidemiological studies and meta-analyses. We applied a comparative risk assessment framework to estimate effects of excess risks on deaths and life expectancy at age 40 y. In 2007, tobacco smoking and high blood pressure accounted for 129,000 deaths (95% CI: 115,000–154,000) and 104,000 deaths (95% CI: 86,000–119,000), respectively, followed by physical inactivity (52,000 deaths, 95% CI: 47,000–58,000), high blood glucose (34,000 deaths, 95% CI: 26,000–43,000), high dietary salt intake (34,000 deaths, 95% CI: 27,000–39,000), and alcohol use (31,000 deaths, 95% CI: 28,000–35,000). In recent decades, cancer mortality attributable to tobacco smoking has increased in the elderly, while stroke mortality attributable to high blood pressure has declined. Life expectancy at age 40 y in 2007 would have been extended by 1.4 y for both sexes (men, 95% CI: 1.3–1.6; women, 95% CI: 1.2–1.7) if exposures to multiple cardiovascular risk factors had been reduced to their optimal levels as determined by a theoretical-minimum-risk exposure distribution.

Conclusions

Tobacco smoking and high blood pressure are the two major risk factors for adult mortality from non-communicable diseases and injuries in Japan. There is a large potential population health gain if multiple risk factors are jointly controlled. Please see later in the article for the Editors'' Summary     

Fgf19 is required for zebrafish lens and retina development

Fgf signaling plays crucial roles in morphogenesis. Fgf19 is required for zebrafish forebrain development. Here, we examined the roles of Fgf19 in the formation of the lens and retina in zebrafish. Knockdown of Fgf19 caused a size reduction of the lens and the retina, failure of closure of the choroids fissure, and a progressive expansion of the retinal tissue to the midline of the forebrain. Fgf19 expressed in the nasal retina and lens was involved in cell survival but not cell proliferation during embryonic lens and retina development. Fgf19 was essential for the differentiation of lens fiber cells in the lens but not for the neuronal differentiation and lamination in the retina. Loss of nasal fate in the retina caused by the knockdown of Fgf19, expansion of nasal fate in the retina caused by the overexpression of Fgf19 and eye transplantation indicated that Fgf19 in the retina was crucial for the nasal-temporal patterning of the retina that is critical for the guidance of retinal ganglion cell axons. Knockdown of Fgf19 also caused incorrect axon pathfinding. The present findings indicate that Fgf19 positively regulates the patterning and growth of the retina, and the differentiation and growth of the lens in zebrafish.     

COX-2 and CCR2 induced by CD40 ligand and MCP-1 are linked to VEGF production in endothelial cells

Recent studies have reported that expression of MCP-1 and its receptor, CCR2; and CD40-CD40 ligand (CD40L) interaction on mesenchymal cells play important roles in tumor development. Studies have also connected MCP-1, CCR2, and CD40L to COX-2 expression. The aim of this study was to examine the effect of MCP-1/CCR2 and CD40-CD40L interaction on COX-2 and VEGF expression in endothelial cells. We also investigated the localization of these proteins in gastric cancer tissue. COX-2 and CCR2 levels were evaluated in CD40L-stimulated HUVECs by Western blot and real-time PCR. VEGF secreted in the culture media was quantified by ELISA. Localizations of MCP-1, CD40L, CD34, CD40 and CCR2 in 34 gastric cancer tissue specimens were evaluated by immunohistochemistry. CD40-CD40L interaction-induced COX-2 production and subsequently, upregulated COX-2 production contributed to elevated VEGF and CCR2 levels in CD40L-stimulated HUVECs. CD40L-stimulated VEGF production was COX-2 but not COX-1 dependent. RS-102895, a CCR2-specific antagonist, significantly reduced VEGF production in CD40L- and MCP-1-stimulated HUVECs. MCP-1 had a synergistic effect on COX-2, CCR2 and VEGF levels in CD40L-stimulated HUVECs. In gastric cancer tissue, there was significant correlation between microvessel density and scores for CD40L, MCP-1 and CCR2 protein expression. Thus, MCP-1 had a synergistic effect on COX-2 and CCR2 protein expression in CD40L-stimulated HUVECs and thereby stimulated VEGF production in these cells.     

Cancer Mortality Studies in Misara,a Radon Hot Spring in Japan: A Summary up to 2003

Misasa, which is well known for its radon hot springs, is a historical town located in Tottori Prefecture, Japan. The presence of the hot springs is noted in an 1164 entry in town records. The presence of radioactivity in the hot springs was recognized in 1914. Since ancient times, the spring water has been believed to be healthful. The town of Misasa has 90 radon hot spring sources, yielding a total volume of 3,000 tons of spring water a day. The concentration of ²²²Rn in the spring water ranges from 17.4 to 9,361 Bq per liter. A series of cancer mortality studies on the radon hot springs were conducted through the collaboration of scientists scattered throughout different Japanese cities. At present, no definite evidence has been discovered that indicates an increase of cancer mortality in the Misasa radon hot spring area. Even a lower risk was found for stomach cancers in both radon and no-radon hot springs.     

Synthesis of zinc bacteriochlorophyll-d analogues with various 17-substituents and their chlorosomal self-aggregates in non-polar organic solvents

Zinc 3(1)-demethyl-bacteriopheophorbides-d possessing various 17-propionate residue (17(2)-COOCH2-) were prepared as models of light-harvesting pigments in major photosynthetic antennae of green bacteria. The synthetic compounds were monomeric in polar organic solvents to give the same visible absorption spectra, but self-aggregated in non-polar organic solvents to afford large oligomers with broadened and red-shifted absorption bands. The peak positions were identical for all the self-aggregates but the widths of the redmost (Qy) bands were dependent upon the substituents. More hydrophobic and less sterically hindered hydrocarbon moieties as the esterifying group sharpened oligomeric Qy bands, giving less inhomogeneous supramolecular structures due to stabilization of the self-aggregates. In the reverse esters (17(2)-CH2OCO-) structurally isomeric to the above compounds, the same optical behavior was observed. Movement of the oxo group from the 17(3)- to 17(5)-position (17(2)-COOCH2--->17(2)-CH2OCO-) slightly increased the widths of self-aggregated Qy bands without change of the peak positions, where minor less self-aggregated components would enhance in the solution due to a little more stabilization of such species by the 17(4)-carbonyl group.