Impaired amino acid transport at the blood brain barrier is a cause of autism spectrum disorder

Autism spectrum disorders (ASD) are a group of genetic disorders often overlapping with other neurological conditions. We previously described abnormalities in the branched-chain amino acid (BCAA) catabolic pathway as a cause of ASD. Here, we show that the solute carrier transporter 7a5 (SLC7A5), a large neutral amino acid transporter localized at the blood brain barrier (BBB), has an essential role in maintaining normal levels of brain BCAAs. In mice, deletion of Slc7a5 from the endothelial cells of the BBB leads to atypical brain amino acid profile, abnormal mRNA translation, and severe neurological abnormalities. Furthermore, we identified several patients with autistic traits and motor delay carrying deleterious homozygous mutations in the SLC7A5 gene. Finally, we demonstrate that BCAA intracerebroventricular administration ameliorates abnormal behaviors in adult mutant mice. Our data elucidate a neurological syndrome defined by SLC7A5 mutations and support an essential role for the BCAA in human brain function

Reduction and Gasification Characteristics of A Unique Iron Ore/ carbon Composite Prepared from Robe River and A Coal Tar Vacuum Residue

We have prepared a unique iron ore/carbon composites (IOC) from a low grade iron ore called Robe River and a thermoplastic carbonaceous material. When Robe River which contains iron as goethite, FeOOH, is heated up to 250 to 300°C, the OH groups are removed as H2O, leaving flat pore spaces of 0.8 nm wide between 2.0 nm thick Fe₂O₃ layers. The pore spaces are, however, closed over 300°C by the sintering of the Fe₂O₃ layers. The idea proposed is to insert the thermoplastic carbonaceous material into the pore space of 0.8 nm wide while the pore spaces are opened and to carbonize it to form carbon in the pore space below 500°C. The iron oxide in the IOC thus prepared is reduced very rapidly in inert atmosphere and the carbon retained in the pore space is gasified by CO₂ very rapidly also. In this work the reaction characteristics of the unique iron ore/carbon composite prepared from Rove River and a coal tar vacuum residue, CTVR, were examined for its direct reduction, indirect reduction in a H₂ atmosphere, and coke gasification in a CO₂ atmosphere from the viewpoints of reaction enthalpies and rate parameters. The examinations clarified that the carbonaceous material retained as coke in the pore space of iron ore are very reactive and show reaction characteristics different from bulk carbon

Investigation on conversion pathways in degradative solvent extraction of rice straw by using liquid membrane-FTIR spectroscopy

Degradative solvent extraction (DSE) is effective in both dewatering and upgrading biomass wastes through the selective removal of oxygen functional groups. However, this conversion mechanism has yet to be elucidated. Here, liquid membrane-FTIR spectroscopy was utilized to examine the main liquid product (Solvent-soluble) without sample modification. Rice straw (RS) and 1-methylnaphthalene (as a non-hydrogen donor solvent) were used as materials, and measurements were performed at treatment temperatures of 200, 250, 300, and 350 °C for 0 min, and at 350 °C for 60 min. The Solvent-soluble spectra were quantitatively analyzed, and changes in the oxygen-containing functional groups and hydrogen bonds at each temperature were used to characterize the DSE mechanism. It was determined that the DSE reaction process can be divided into three stages. During the first stage, 200–300 °C (0 min), oxygen was removed via dehydration, and aromaticity was observed. In the second stage, 300–350 °C (0 min), deoxygenation reactions involving dehydration and decarboxylation were followed by reactions for aromatization. For the third stage, 350 °C (0–60 min), further aromatization and dehydration reactions were observed. Intramolecular reactions are indicated as the predominant mechanism for dehydration in RS DSE, and the final product is composed of smaller molecular compounds

Structure–activity characteristics of phenylalanine analogs selectively transported by L-type amino acid transporter 1 (LAT1)

Abstract L-type amino acid transporter 1 (LAT1) is a transmembrane protein responsible for transporting large neutral amino acids. While numerous LAT1-targeted compound delivery for the brain and tumors have been investigated, their LAT1 selectivity often remains ambiguous despite high LAT1 affinity. This study assessed the LAT1 selectivity of phenylalanine (Phe) analogs, focusing on their structure–activity characteristics. We discovered that 2-iodo-l-phenylalanine (2-I-Phe), with an iodine substituent at position 2 in the benzene ring, markedly improves LAT1 affinity and selectivity compared to parent amino acid Phe, albeit at the cost of reduced transport velocity. l-Phenylglycine (Phg), one carbon shorter than Phe, was found to be a substrate for LAT1 with a lower affinity, exhibiting a low level of selectivity for LAT1 equivalent to Phe. Notably, (R)-2-amino-1,2,3,4-tetrahydro-2-naphthoic acid (bicyclic-Phe), with an α-methylene moiety akin to the α-methyl group in α-methyl-l-phenylalanine (α-methyl-Phe), a known LAT1-selective compound, showed similar LAT1 transport maximal velocity to α-methyl-Phe, but with higher LAT1 affinity and selectivity. In vivo studies revealed tumor-specific accumulation of bicyclic-Phe, underscoring the importance of LAT1-selectivity in targeted delivery. These findings emphasize the potential of bicyclic-Phe as a promising LAT1-selective component, providing a basis for the development of LAT1-targeting compounds based on its structural framework

Organellar Na+/H+ Exchangers:Novel Players in Organelle pH Regulation and Their Emerging Functions

Mammalian Na+/H+ exchangers (NHEs) play a fundamental role in cellular ion homeostasis. NHEs exhibit an appreciable variation in expression, regulation, and physiological function, dictated by their dynamics in subcellular localization and/or interaction with regulatory proteins. In recent years, a subgroup of NHEs consisting of four isoforms has been identified, and its members predominantly localize to the membranes of the Golgi apparatus and endosoines. These organellar NHEs constitute a family of transporters with an emerging function in the regulation of luminal pH and in intracellular membrane trafficking as expressed, for example, in cell polarity development. Moreover, specific roles of a variety of cofactors, regulating the intracellular dynamics of these transporters, are also becoming apparent, thereby providing further insight into their mechanism of action and overall functioning. Interestingly, organellar NHEs have been related to mental disorders, implying a potential role in the brain, thus expanding the physiological significance of these transporters

Ratiometric fluorescence imaging of cell surface pH by poly(ethylene glycol)-phospholipid conjugated with fluorescein isothiocyanate

Various physiological and pathological processes are accompanied with the alteration of pH at extracellular juxtamembrane region. Accordingly, the methods to analyze the cell surface pH have been demanded in biological and medical sciences. In this study, we have established a novel methodology for cell surface pH imaging using poly(ethylene glycol)-phospholipid (PEG-lipid) as a core structure of ratiometric fluorescent probes. PEG-lipid is a synthetic amphiphilic polymer originally developed for the cell surface modification in transplantation therapy. Via its hydrophobic alkyl chains of the phospholipid moiety, PEG-lipid is, when applied extracellularly, spontaneously inserted into the plasma membrane and retained at the surface of the cells. We have demonstrated that the PEG-lipid conjugated with fluorescein isothiocyanate (FITC-PEG-lipid) can be used as a sensitive and reversible cell-surfacea-nchored pH probe between weakly alkaline and acidic pH with an excellent spatiotemporal resolution. The remarkably simple procedure for cell-surface labeling with FITC-PEG-lipid would also be advantageous when considering its application to high-throughput in vitro assay. This study further indicates that various probes useful for the investigation of juxtamembrane environments could also be developed by using PEG-lipid as the core structure for bio-membrane anchoring

Linkage of N-cadherin to multiple cytoskeletal elements revealed by a proteomic approach in hippocampal neurons

The CNS synapse is an adhesive junction differentiated for chemical neurotransmission and is equipped with presynaptic vesicles and postsynaptic neurotransmitter receptors. Cell adhesion molecule cadherins not only maintain connections between pre- and postsynaptic membranes but also modulate the efficacy of synaptic transmission. Although the components of the cadherin-mediated adhesive apparatus have been studied extensively in various cell systems, the complete picture of these components, particularly at the synaptic junction, remains elusive. Here, we describe the proteomic assortment of the N-cadherin-mediated synaptic adhesion apparatus in cultured hippocampal neurons. N-cadherin immunoprecipitated from Triton X-100-solubilized neuronal extract contained equal amounts of beta- and alpha-catenins, as well as F-actin-related membrane anchor proteins such as integrins bridged with alpha-actinin-4, and Na+/K+-ATPase bridged with spectrins. A close relative of beta-catenin, plakoglobin, and its binding partner, desmoplakin, were also found, suggesting that a subset of the N-cadherin-mediated adhesive apparatus also anchors intermediate filaments. Moreover, dynein heavy chain and LEK1/CENPF/mitosin were found. This suggests that internalized pools of N-cadherin in trafficking vesicles are conveyed by dynein motors on microtubules. In addition, ARVCF and NPRAP/neurojungin/delta 2-catenin, but not p120ctn/delta 1-catenin or plakophilins-1, -2, -3, -4 (p0071), were found, suggesting other possible bridges to microtubules. Finally, synaptic stimulation by membrane depolarization resulted in an increased 93-kDa band, which corresponded to proteolytically truncated beta-catenin. The integration of three different classes of cytoskeletal systems found in the synaptic N-cadherin complex may imply a dynamic switching of adhesive scaffolds in response to synaptic activity.close

Inhibition of cancer-type amino acid transporter LAT1 suppresses B16-F10 melanoma metastasis in mouse models

Abstract Metastasis is the leading cause of mortality in cancer patients. L-type amino acid transporter 1 (LAT1, SLC7A5) is a Na+-independent neutral amino acid transporter highly expressed in various cancers to support their growth. Although high LAT1 expression is closely associated with cancer metastasis, its role in this process remains unclear. This study aimed to investigate the effect of LAT1 inhibition on cancer metastasis using B16-F10 melanoma mouse models. Our results demonstrated that nanvuranlat (JPH203), a high-affinity LAT1-selective inhibitor, suppressed B16-F10 cell proliferation, migration, and invasion. Similarly, LAT1 knockdown reduced cell proliferation, migration, and invasion. LAT1 inhibitors and LAT1 knockdown diminished B16-F10 lung metastasis in a lung metastasis model. Furthermore, nanvuranlat and LAT1 knockdown suppressed lung, spleen, and lymph node metastasis in an orthotopic metastasis model. We discovered that the LAT1 inhibitor reduced the cell surface expression of integrin αvβ3. Our findings revealed that the downregulation of the mTOR signaling pathway, induced by LAT1 inhibitors, decreased the expression of integrin αvβ3, contributing to the suppression of metastasis. These results highlight the critical role of LAT1 in cancer metastasis and suggest that LAT1 inhibition may serve as a potential target for anti-metastasis cancer therapy

A novel mutation in the SLCO2A1 gene, encoding a prostaglandin transporter, induces chronic enteropathy.

Chronic enteropathy associated with SLCO2A1 gene (CEAS) is caused by loss-of-function mutations in SLCO2A1, which encodes a prostaglandin (PG) transporter. In this study, we report a sibling case of CEAS with a novel pathogenic variant of the SLCO2A1 gene. Compound heterozygous variants in SLCO2A1 were identified in an 8-year-old boy and 12-year-old girl, and multiple chronic nonspecific ulcers were observed in the patients using capsule endoscopy. The splice site mutation (c.940 + 1G>A) of the paternal allele was previously reported to be pathogenic, whereas the missense variant (c.1688T>C) of the maternal allele was novel and had not yet been reported. The affected residue (p.Leu563Pro) is located in the 11th transmembrane domain (helix 11) of SLCO2A1. Because SLCO2A1 mediates the uptake and clearance of PGs, the urinary PG metabolites were measured by liquid chromatography coupled to tandem mass spectrometry. The urinary tetranor-prostaglandin E metabolite levels in the patients were significantly higher than those in unaffected individuals. We established cell lines with doxycycline-inducible expression of wild type SLCO2A1 (WT-SLCO2A1) and the L563P mutant. Immunofluorescence staining showed that WT-SLCO2A1 and the L563P mutant were dominantly expressed on the plasma membranes of these cells. Cells expressing WT-SLCO2A1 exhibited time- and dose-dependent uptake of PGE2, while the mutant did not show any uptake activity. Residue L563 is very close to the putative substrate-binding site in SLCO2A1, R561 in helix 11. However, in a molecular model of SLCO2A1, the side chain of L563 projected outside of helix 11, indicating that L563 is likely not directly involved in substrate binding. Instead, the substitution of Pro may twist the helix and impair the transporter function. In summary, we identified a novel pathogenic variant of SLCO2A1 that caused loss-of-function and induced CEAS