Paradoxical Sensitivity to an Integrated Stress Response Blocking Mutation in Vanishing White Matter Cells

The eukaryotic translation initiation factor eIF2B promotes mRNA translation as a guanine nucleotide exchange factor (GEF) for translation initiation factor 2 (eIF2). Endoplasmic reticulum (ER) stress-mediated activation of the kinase PERK and the resultant phosphorylation of eIF2’s alpha subunit (eIF2α) attenuates eIF2B GEF activity thereby inducing an integrated stress response (ISR) that defends against protein misfolding in the ER. Mutations in all five subunits of human eIF2B cause an inherited leukoencephalopathy with vanishing white matter (VWM), but the role of the ISR in its pathogenesis remains unclear. Using CRISPR-Cas9 genome editing we introduced the most severe known VWM mutation, EIF2B4^A391D, into CHO cells. Compared to isogenic wildtype cells, GEF activity of cells with the VWM mutation was impaired and the mutant cells experienced modest enhancement of the ISR. However, despite their enhanced ISR, imposed by the intrinsic defect in eIF2B, disrupting the inhibitory effect of phosphorylated eIF2α on GEF by a contravening EIF2S1/eIF2α^S51A mutation that functions upstream of eIF2B, selectively enfeebled both EIF2B4^A391D and the related severe VWM EIF2B4^R483W cells. The basis for paradoxical dependence of cells with the VWM mutations on an intact eIF2α genotype remains unclear, as both translation rates and survival from stressors that normally activate the ISR were not reproducibly affected by the VWM mutations. Nonetheless, our findings support an additional layer of complexity in the development of VWM, beyond a hyperactive ISR.     

Synthesis and evaluation of a [18F]formyl–Met–Leu–Phe derivative: A positron emission tomography imaging probe for bacterial infections

The tripeptide formyl–Met–Leu–Phe (fMLF) is a prototype of N-formylated chemotactic peptides for neutrophils owing to its ability to bind and activate the G protein-coupled formyl peptide receptor (FPR). Here, we developed an ¹⁸F-labeled fMLF derivative targeting FPR as a positron emission tomography (PET) imaging probe for bacterial infections. The study demonstrates that the fMLF derivative fMLFXYk(FB)k (X?=?Nle) has a high affinity for FPR (Ki?=?0.62?±?0.13?nM). The radiochemical yield and purity of [¹⁸F]fMLFXYk(FB)k were 16% and >96%, respectively. The in vivo biodistribution study showed that [¹⁸F]fMLFXYk(FB)k uptake was higher in the bacterial infected region than in the non-infected region. We observed considerably higher infection-to-muscle ratio of 4.6 at 60?min after [¹⁸F]fMLFXYk(FB)k injection. Furthermore, small-animal PET imaging studies suggested that [¹⁸F]fMLFXYk(FB)k uptake in the bacterial infected region was clearly visualized 60?min after injection.     

Analysis of inphase interaction pattern in EEG

Inphase interactions among EEG signals recorded using eight electrodes were investigated. The inphase interaction parameters are presented in two ways: (1) matrix form in which the number of inphase interactions are tabulated; and (2) histogram in which the number of inphase interactions are plotted pair-wise between two sites as a function of phase delays in milliseconds. The highest number of interactions occurs between 0 and 8 ms in normal brains. The values of interaction parameters are enhanced by various activities. For example, inphase interaction parameters increase in the motor area in the right hemisphere if the EEG is recorded during repeated left fist clenching. Inphase interactions are drastically altered by the presence of a tumor. We studied the inphase interactions of the EEG of a patient having an occipital tumor. The interaction parameters are greatly diminished in this area, indicating a severe impairment of neuronal communications between both hemispheres in the occipital region. The confidence limits of the changes in inphase interaction parameters during fist clenching are tested statistically using the Student's t test. The test shows that the interaction parameters increase, in general, with 1–5% confidence limits in respective cortex areas as a result of fist clenching.     

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     

Granular tau oligomers as intermediates of tau filaments

Neurofibrillary tangles (NFTs) are pathological hallmarks of several neurodegenerative disorders, including Alzheimer's disease (AD). NFTs are composed of microtubule-binding protein tau, which assembles to form paired helical filaments (PHFs) and straight filaments. Here we show by atomic force microscopy that AD brain tissue and in vitro tau form granular and fibrillar tau aggregates. CD spectral analysis and immunostaining with conformation-dependent antibodies indicated that tau may undergo conformational changes during fibril formation. Enriched granules generated filaments, suggesting that granular tau aggregates may be an intermediate form of tau fibrils. The amount of granular tau aggregates was elevated in prefrontal cortex of Braak stage I cases compared to that of Braak stage 0 cases, suggesting that granular tau aggregation precedes PHF formation. Thus, granular tau aggregates may be a relevant marker for the early diagnosis of tauopathy. Reducing the level of these aggregates may be a promising therapy for tauopathies and for promoting healthy brain aging.     

Trace elements influenced by environmental changes in Lake Biwa: (I) Seasonal variations under suboxic hypolimnion conditions during 2007 and 2009

Dissolved oxygen (DO) in the Lake Biwa hypolimnion reached its lowest level of <1 mg kg^?1 in 2007. In this paper, we report the variations in the total dissolvable (TD), dissolved (D), and labile particulate (LP) fractions of Al, Si, P, V, Cr, Mn, Fe, Ni, Cu, Zn, As, Mo, W, and U in Lake Biwa 2007 and 2009. Al and Fe species were predominantly in the form of LP-Al and LP-Fe and were strongly correlated with one another (r = 0.99), suggesting that the weathering of aluminous minerals and the supply of clay mineral particles are the main factors that influence the distributions of Al and Fe. Although D-Al increased in the summer epilimnion, D-Fe was relatively low, probably as a result of uptake by plants. Reductive release of Fe from the bottom was not seen. Mn was also dominated by LP-Mn, but this fraction showed a different distribution to those of LP-Al and LP-Fe. The D-Mn and LP-Mn concentrations varied by factors of 700–1000 and showed marked increases in the bottom water during stratification in 2007. We believe that Mn²⁺ was released from the sediments and oxidized by DO in the bottom water. Ni, Cu, Zn, and Cr, which exist as cationic species, had LP/TD ratios of 0.1–0.7 and relatively uniform distributions. Si, P, V, As, Mo, W, and U, which form oxoacid species, had LP/TD ratios of 0–0.8. Si, P, and As were characterized by nutrient-like profiles, V, W, and U showed summer maxima in the epilimnion, and Mo had a uniform distribution. TD-Mo increased in the bottom water along with TD-Mn, while TD-V and TD-W showed significant decreases. These results are likely attributable to differences in the adsorption of these elements onto manganese oxides and iron hydroxides.     

The effects of phosphorylation and dephosphorylation of brain myosin on its actin-activated Mg2+-ATPase and contractile activities

Purified bovine brain myosin contained approximately 1 and 3 mol of protein-bound phosphate/mol myosin in the light chains and heavy chains, respectively. Large portions of this light chain- and heavy chain-bound phosphate (about 0.8 and 2.4 mol, respectively) were removed by incubation with a brain phosphoprotein phosphatase and potato acid phosphatase, respectively. Upon phosphorylation of the dephosphorylated brain myosin with myosin light chain kinase and casein kinase II, about 1.6 and 3.0 mol of phosphate was incorporated into the light chains and heavy chains, respectively, while much lower levels of phosphate were incorporated into the non-dephosphorylated brain myosin under the same conditions. The actin-activated Mg2+-ATPase activity of brain myosin rephosphorylated with myosin light chain kinase was about twice as high as that of dephosphorylated brain myosin (about 30 and 15 nmol phosphate/mg/min, respectively). On the other hand, whereas the rephosphorylated brain myosin superprecipitated rapidly with F-actin, the rate of superprecipitation of the dephosphorylated brain myosin was extremely low. Under appropriate conditions, a loose network of tiny superprecipitates, which formed initially throughout the solution, contracted to form eventually a large and dense particle. These results indicate that phosphorylation of the light chains of brain myosin is a prerequisite for the contraction of brain actomyosin. The role of phosphorylation of the heavy chains by casein kinase II remains to be elucidated.