Phosphorylation and Inactivation of Brain Glycogen Synthase by a Multifunctional Calmodulin-Dependent Protein Kinase

Glycogen synthase was partially purified from canine brain to about 70% purity. The purified enzyme showed differences from the properties of the skeletal muscle enzyme with respect to molecular weights of the holoenzyme and subunit and phosphopeptide mapping. The multifunctional calmodulin-dependent protein kinase from the brain phosphorylated brain glycogen synthase with concomitant inactivation of the enzyme. Although about 1.3 mol of phosphate/mol subunit was maximally incorporated into glycogen synthase, 0.4 mol of phosphate/mol subunit was sufficient for the maximal inactivation of the enzyme. The results indicate that brain glycogen synthase is regulated in a calmodulin-dependent manner similarly to the skeletal muscle enzyme, but that the brain enzyme is different from the skeletal muscle enzyme.     

Cloning of a bovine orphan transporter and its short splicing variant

We have isolated a cDNA (bv7-3) encoding a member of the Na+,Cl(-)-dependent transporter family and its short splicing variant (bv7-3s) by screening a bovine retina cDNA library. Sequence analysis revealed that bv7-3 encodes a protein of 729 amino acids and is a bovine homologue of the rat orphan transporter v7-3-2. bv7-3s contains 265 amino acids, sharing 252 N-terminal amino acids with bv7-3. Both mRNAs for bv7-3 and bv7-3s were detected in nervous system by Northern blot analysis. In immunofluorescence analysis in transfected HEK 293T cells, myc-tagged bv7-3 was mainly detected on the plasma membrane, whereas myc-tagged bv7-3s showed a pattern of intracellular membrane staining.     

Analysis of Petal Anthocyanins to Investigate Flower Coloration of Zhongyuan (Chinese) and Daikon Island (Japanese) Tree Peony Cultivars

Pn, Pg; Pn, Pg > Cy ; Pn, Cy and Pn, Cy > Pg groups. Each group consequently specified significant features among CIELAB color notation and petal pigmentation, being adequate to characterize tree peony flowers as similar between Zhongyuan and Daikon Island cultivars, thus the cultivars of the two areas are suggested to be related to one another. Received 25 April 2000/ Accepted in revised form 21 December 2000     

Production of Lysine by Pyruvate Dehydrogenase Mutants of Brevibacterium flavum

Mutants with low pyruvate dehydrogenase (PD) activities were derived from a pyruvate kinase-deficient lysine-producing mutant of Brevibacterium flavum, No. 22. They were selected as prototrophic revertants of the acetate auxotrophs of strain No. 22. Among them strain KD-11 produced 55g/liter of lysine as its HCI salt when cultured for 72 hr in a medium containing lOOg/liter of glucose, soybean-meal hydrolysate and methionine. The lysine yield of strain KD-11 was the highest ever reported (55%). The mutant required a higher concentration of methionine for maximum production and gave a smaller amount of cell mass in cultivation than its parent. PD activity of strain No. 22 was stimulated by cysteine, stabilized by glycerol, and gave apparent Kms of 89, 22, 380, 83 μM for pyruvate, coenzyme A, 3-acetylpyridine adenine dinucleotide, and NAD, respectively, under standard conditions. The apparent Km for NAD of PD from strain KD-11 was 10-times higher than that from No. 22. When the concentration of NAD was low, the cell extracts of strain KD-11 showed low PD activity. The specific activity of phosphoenolpyruvate carboxylase of strain KD-11 was slightly higher than that of strain No. 22, while the inhibition by aspartate of the former enzyme was weaker than that of the latter.     

Plasma B-type Natriuretic Peptide as a Predictor of Cardiovascular Events in Subjects with Atrial Fibrillation: A Community-Based Study

Objectives

Atrial fibrillation (AF) is a significant public health issue due to its high prevalence in the general population, and is associated with an increased risk of cardiovascular (CV) events including systemic thrombo-embolism, heart failure, and coronary artery disease. The relationship between plasma B-type natriuretic peptide (BNP) and CV risk in real world AF subjects remains unknown.

Methods

The subject of the study (n = 228; mean age = 69 years) was unselected individuals with AF in a community-based population (n = 15,394; AF prevalence rate = 1.5%). The CV event free rate within each BNP tertile was estimated, and Cox regression analysis was performed to examine the relative risk of the onset of CV events among the tertiles. The prognostic ability of BNP was compared to an established risk score for embolic events (CHADS2 score). In addition, to determine the usefulness of BNP as a predictor in addition to CHADS2 score, we calculated Net Reclassification Improvement (NRI) and Integrated Discrimination Improvement (IDI) indices.

Results

During the follow-up period 58 subjects experienced CV events (52 per 1,000 person-years). The event-free ratio was significantly lower in the highest tertile (p < 0.02). After adjustment for established CV risk factors, the hazard ratio (HR) of the highest tertile was significantly higher than that of the lowest tertile (HR = 2.38; p < 0.02). The predictive abilities of plasma BNP in terms of sensitivity and specificity for general CV events were comparable to those of CHADS2 score. Adding BNP to the CHADS2 score only model improved the NRI (0.319; p < 0.05) and the IDI (0.046; p < 0.05).

Conclusion

Plasma BNP is a valuable biomarker both singly or in combination with an established scoring system for assessing general CV risk including stroke, heart failure and acute coronary syndrome in real-world AF subjects.     

Different effects of two exotic herbivores on the pollinator-mediated effect of an exotic plant on a native plant

Exotic plants can affect native plants indirectly through various biotic interactions. However, combinations of the multiple indirect effects of exotic plants on native plants have been rarely evaluated. Herbivory can either positively or negatively influence plant–pollinator interactions. Here, we addressed whether the pollinator-mediated plant interaction between exotic and native plants is altered through the introduction of exotic herbivores by conducting a 2-year common garden experiment. We compared the effects of pollinator-mediated indirect effects of an exotic plant, Solidago altissima, on the co-flowering native plant Aster microcephalus in geographically different populations reflecting differences in insect herbivore communities. We found a positive effect of co-flowering S. altissima on pollinator visitation of A. microcephalus, which varied between gardens and years. The co-flowering S. altissima did not significantly affect the seed set of A. microcephalus in the first year but had a negative effect in the second year. The facilitative effect of S. altissima on A. microcephalus pollination was suggested to be negatively affected by an exotic aphid, while it was not significantly affected by an exotic lace bug. Our study suggests that the phenology and feeding guilds of the herbivores may be critical for predicting the effect of exotic plants on native plants through herbivore–pollinator interactions. Integrated effects between plant interactions via multiple species interactions under different abiotic and biotic environments are necessary to understand the impact of exotic plants under complex interactions in nature.     

Preparation of metaphase chromatin of Physarumpolycephalum without the loss of repressed RNA synthesis

A novel method for the preparation of intact chromatin from the slime mold $\text{Physarum}\text{polycephalum>}$ which retains the $\text{in}\text{vivo}$ property of RNA synthesis is described. Preparations from G₂-cells were highly active, while those from metaphase-cells were inactive. The plasmodial cells were disrupted by gentle homogenization on a polyethylene sieve in a neutral isotonic sucrose medium containing Mg⁺⁺, deoxycholate and EGTA, a Ca⁺⁺-chelating agent. The nuclei were lysed in a hypotonic buffer without use of EDTA and chromatin was precipitated by centrifugation after addition of Mg⁺⁺.     

Sense-overlapping lncRNA as a decoy of translational repressor protein for dimorphic gene expression

Long noncoding RNAs (lncRNAs) are vastly transcribed and extensively studied but lncRNAs overlapping with the sense orientation of mRNA have been poorly studied. We analyzed the lncRNA DAPALR overlapping with the 5´ UTR of the Doublesex1 (Dsx1), the male determining gene in Daphnia magna. By affinity purification, we identified an RNA binding protein, Shep as a DAPALR binding protein. Shep also binds to Dsx1 5´ UTR by recognizing the overlapping sequence and suppresses translation of the mRNA. In vitro and in vivo analyses indicated that DAPALR increased Dsx1 translation efficiency by sequestration of Shep. This regulation was impaired when the Shep binding site in DAPALR was deleted. These results suggest that Shep suppresses the unintentional translation of Dsx1 by setting a threshold; and when the sense lncRNA DAPALR is expressed, DAPALR cancels the suppression caused by Shep. This mechanism may be important to show dimorphic gene expressions such as sex determination and it may account for the binary expression in various developmental processes.