Preparation of glycosylated amino acid derivatives for glycoprotein synthesis by in vitro translation system

General preparation of glycosylated amino acylated nucleotide for in vitro peptide synthesis was described. Both O-glycosylated amino acyl nucleotides and C-glycosylated amino acyl nucleotide were synthesized by choosing the appropriate protecting group.     

Learning to simulate others' decisions

A fundamental challenge in social cognition is how humans learn another person's values to predict their decision-making behavior. This form of learning is often assumed to require simulation of the other by direct recruitment of one's own valuation process to model the other's process. However, the cognitive and neural mechanism of simulation learning is not known. Using behavior, modeling, and fMRI, we show that simulation involves two learning signals in a hierarchical arrangement. A simulated-other's reward prediction error processed in ventromedial prefrontal cortex mediated simulation by direct recruitment, being identical for valuation of the self and simulated-other. However, direct recruitment was insufficient for learning, and also required observation of the other's choices to generate?a simulated-other's action prediction error encoded in dorsomedial/dorsolateral prefrontal cortex. These findings show that simulation uses a core prefrontal circuit for modeling the other's valuation to generate prediction and an adjunct circuit for tracking behavioral variation to refine prediction.     

Thermotolerant cyclamen with reduced acrolein and methyl vinyl ketone

Reduced levels of trienoic fatty acids (TAs) in chloroplast membranes induce thermotolerance in several plant species, but the underlying mechanisms remain unclear. TA peroxidation in plant cell membranes generates cytotoxic, TA-derived compounds containing α,β-unsaturated carbonyl groups. The relationship between low TA levels and the amounts of cytotoxic TA-derived compounds was examined using thermotolerant transgenic cyclamen (Cyclamen persicum Mill.) with low TA contents. Changes in the levels of the cytotoxic TA-derived acrolein (ACR), methyl vinyl ketone (MVK), (E)-2-hexenal, 4-hydroxy-2-nonenal, and malondialdehyde were analysed in the leaf tissues of wild-type (WT) and thermotolerant transgenic cyclamen under heat stress. Levels of ACR and MVK in the WT increased in parallel with the occurrence of heat-induced tissue damage, whereas no such changes were observed in the thermotolerant transgenic lines. Furthermore, exogenous ACR and MVK infiltrated into leaves to concentrations similar to those observed in heat-stressed WT leaves caused similar disease symptoms. These results suggest that thermotolerance in transgenic cyclamen depends on reduced production rates of ACR and MVK under heat stress, due to the low level of TAs in these plants.     

Pyrrolysine analogs as substrates for bacterial pyrrolysyl-tRNA synthetase in vitro and in vivo

Pyrrolysine-tRNA(Pyl) complex is produced by pyrrolysyl-tRNA synthetase (PylRS). In this study, we investigated the substrate specificity of Desulfitobacterium hafnience PylRS. PylRS incorporated various L-lysine derivatives into tRNA(Pyl) in vitro. In addition, the PylRS/tRNA(Pyl) pair introduced these lysine derivatives into the recombinant protein by the Escherichia coli expression system, indicating that this PylRS/tRNA(Pyl) pair can be used in protein engineering technology.     

Genetic and functional analyses of SHANK2 mutations suggest a multiple hit model of autism spectrum disorders

Autism spectrum disorders (ASD) are a heterogeneous group of neurodevelopmental disorders with a complex inheritance pattern. While many rare variants in synaptic proteins have been identified in patients with ASD, little is known about their effects at the synapse and their interactions with other genetic variations. Here, following the discovery of two de novo SHANK2 deletions by the Autism Genome Project, we identified a novel 421 kb de novo SHANK2 deletion in a patient with autism. We then sequenced SHANK2 in 455 patients with ASD and 431 controls and integrated these results with those reported by Berkel et al. 2010 (n = 396 patients and n = 659 controls). We observed a significant enrichment of variants affecting conserved amino acids in 29 of 851 (3.4%) patients and in 16 of 1,090 (1.5%) controls (P = 0.004, OR = 2.37, 95% CI = 1.23–4.70). In neuronal cell cultures, the variants identified in patients were associated with a reduced synaptic density at dendrites compared to the variants only detected in controls (P = 0.0013). Interestingly, the three patients with de novo SHANK2 deletions also carried inherited CNVs at 15q11–q13 previously associated with neuropsychiatric disorders. In two cases, the nicotinic receptor CHRNA7 was duplicated and in one case the synaptic translation repressor CYFIP1 was deleted. These results strengthen the role of synaptic gene dysfunction in ASD but also highlight the presence of putative modifier genes, which is in keeping with the “multiple hit model” for ASD. A better knowledge of these genetic interactions will be necessary to understand the complex inheritance pattern of ASD.     

Catheter ablation of ventricular tachycardia originating from the left posterior papillary muscle guided by the shadow of a multipolar catheter

A 62-year-old man without structural heart disease underwent electrophysiological testing for ventricular tachycardia (VT). Hemodynamically unstable VT was induced after isoproterenol (ISP) provocation. Electroanatomical mapping using a multipolar catheter identified the earliest activation originating from the posterior papillary muscle (PPM) where prepotentials preceding the local ventricular electrogram were observed. Irrigated radiofrequency current guided by the shadow of a multipolar catheter eliminated the VT. This case suggested that multipolar catheters may be helpful for identifying tachycardia origins arising from the PPM.     

Effects of dehydration rate on physiological responses and survival after rehydration in larvae of the anhydrobiotic chironomid

Strategies to combat desiccation are critical for organisms living in arid and semi-arid areas. Larvae of the Australian chironomid Paraborniella tonnoiri resist desiccation by reducing water loss. In contrast, larvae of the African species Polypedilum vanderplanki can withstand almost complete dehydration, referred to as anhydrobiosis. For successful anhydrobiosis, the dehydration rate of P. vanderplanki larvae has to be controlled. Here, we desiccated larvae by exposing them to different drying regimes, each progressing from high to low relative humidity, and examined survival after rehydration. In larvae of P. vanderplanki, reactions following desiccation can be categorized as follows: (I) no recovery at all (direct death), (II) dying by unrepairable damages after rehydration (delayed death), and (III) full recovery (successful anhydrobiosis). Initial conditions of desiccation severely affected survival following rehydration, i.e. P. vanderplanki preferred 100% relative humidity where body water content decreased slightly. In subsequent conditions, unfavorable dehydration rate, such as more than 0.7 mg water lost per day, resulted in markedly decreased survival rate of rehydrated larvae. Slow dehydration may be required for the synthesis and distribution of essential molecules for anhydrobiosis. Larvae desiccated at or above maximum tolerable rates sometimes showed temporary recovery but died soon after.     

Calcineurin is involved in the early activation of NMDA-mediated cell death in mutant huntingtin knock-in striatal cells

Excitotoxicity has been proposed as one of the mechanisms involved in the specific loss of striatal neurons that occurs in Huntington's disease. Here, we studied the role of calcineurin in the vulnerability of striatal neurons expressing mutant huntingtin to excitotoxicity. To this end, we induced excitotoxicity by adding NMDA to a striatal precursor cell line expressing full-length wild-type (STHdh^Q7/Q7) or mutant (STHdh^Q111/Q111) huntingtin. We observed that cell death appeared earlier in STHdh^Q111/Q111 cells than in STHdh^Q7/Q7 cells. Interestingly, these former cells expressed higher levels of calcineurin A that resulted in a greater increase of its activity after NMDA receptor stimulation. Moreover, transfection of full-length mutant huntingtin in different striatal-derived cells (STHdh^Q7/Q7, M213 and primary cultures) increased calcineurin A protein levels. To determine whether high levels of calcineurin A might account for the earlier activation of cell death in mutant huntingtin knock-in cells, wild-type cells were transfected with calcineurin A. Calcineurin A-transfected STHdh^Q7/Q7 cells displayed a significant increase in cell death compared with that recorded in green fluorescent protein-transfected cells after NMDA treatment. Notably, addition of the calcineurin inhibitor FK-506 produced a more robust reduction in cell death in mutant huntingtin knock-in cells than it did in wild-type cells. These results suggest that high levels of calcineurin A could account for the increased vulnerability of striatal cells expressing mutant huntingtin to excitotoxicity.     

A food-derived synergist of NGF signaling: identification of protein tyrosine phosphatase 1B as a key regulator of NGF receptor-initiated signal transduction

Neurotrophins, such as the nerve growth factor (NGF), play an essential role in the growth, development, survival and functional maintenance of neurons in the central and peripheral systems. They also prevent neuronal cell death under various stressful conditions, such as ischemia and neurodegenerative disorders. NGF induces cell differentiation and neurite outgrowth by binding with and activating the NGF receptor tyrosine kinase followed by activation of a variety of signaling cascades. We have investigated the NGF-dependent neuritogenesis enhancer potential of a food-derived small molecule contained in Brassica vegetables and identified the protein tyrosine phosphatase (PTP) 1B as a key regulator of the NGF receptor-initiated signal transduction. Based on an extensive screening of Brassica vegetable extracts for the neuritogenic-promoting activity in the rat pheochromocytoma cell line PC12, we found the Japanese horseradish, wasabi (Wasabia japonica, syn. Eutrema wasabi), as the richest source and identified 6-methylsulfinylhexyl isothiocyanate (6-HITC), an analogue of sulforaphane isolated from broccoli, as one of the major neuritogenic enhancers in the wasabi. 6-HITC strongly enhanced the neurite outgrowth and neurofilament expression elicited by a low-concentration of NGF that alone was insufficient to induce neuronal differentiation. 6-HITC also facilitated the sustained-phosphorylation of the extracellular signal-regulated kinase and the autophosphorylation of the NGF receptor TrkA. It was found that PTP1B act as a phosphatase capable of dephosphorylating Tyr-490 of TrkA and was inactivated by 6-HITC in a redox-dependent manner. The identification of PTP1B as a regulator of NGF signaling may provide new clues about the chemoprotective potential of food components, such as isothiocyanates.