Prebiotic origin of glycolytic metabolism: histidine and cysteine can produce acetyl CoA from glucose via reactions homologous to non-phosphorylated Entner-Doudoroff pathway

Conversion of glucose to pyruvate via reactions homologous to the non-phosphorylated Entner-Doudoroff (non-P ED) pathway could be achieved in the presence of two amino acid catalysts, cysteine and histidine: cystine oxidizes glucose to gluconic acid by the reaction homologous to glucose dehydrogenase and histidine changes gluconic acid to 2-keto-3-deoxy gluconic acid, then to pyruvate by the reaction homologous to gluconic acid dehydratase and 2-keto-3-deoxy gluconate aldolase, respectively. Pyruvate can be converted to acetyl CoA by the reaction with CoA, TPP and FAD in the presence of cysteine and histidine, which resembles pyruvate dehydrogenase reaction. It was found that gluconic acid dehydration alone is non-specific, in contrast to other reactions. The non-P ED pathway is used by some extreme thermophiles in bacteria and archaea, usually thought as the oldest among the contemporary organisms. This study suggests the possible contribution of amino acid to the origin of the glycolytic pathway.     

Expression of neolactoglycolipids: sialosyl-, disialosyl-,O-acetyldisialosyl- and fucosyl- derivatives of neolactotetraosyl ceramide and neolactohexaosyl ceramide in the developing cerebral cortex and cerebellum

The following neolacto glycolipids were identified and their developmental expression was studied in the rat cerebral cortex and cerebellum: Fuc1-³IIInLcOse₄Cer,Fuc1-³VnLcOse₆Cer and (Fuc)₂1-³III,³VnLcOse₆Cer, as well as acidic glycolipids, NeuAc2-³IVnLcOse₄Cer [nLM1], (NeuAc)₂2-³IVnLcOse₄Cer [nLD1],O-acetyl (NeuAc)₂2-³IVnLcOse₄Cer [OAc-nLD1] and their higher neolactosaminyl homologues NeuAc2-³VInLcOse₆Cer [nHM1] and (NeuAc)₂2-³VInLcOse₆Cer [nHD1]. These glycolipids were expressed in the cerebral cortex only during embryonic stages and disappeared postnatally. This loss was ascribed to the down regulation of the synthesis of the key precursor LcOse₃Cer which is synthesized by the enzyme lactosylceramide:N-acetylglucosaminyl transferase. On the other hand in the cerebellum, these glycolipids increased with postnatal development due to increasing availability of LcOse₃Cer. In the cerebellum, only nLM1 and fucosyl-neolactoglycolipids declined after postnatal day 10–15, perhaps due to regulation by other glycosyltransferases. Also, in the cerebellum, nLD1 and nHD1 were shown to be specifically associated with Purkinje cells and their dendrites in the molecular layer and with their axon terminals in the deep cerebellar nuclei, similar to other neolactoglycolipids shown previously.     

Glucose deprivation accelerates VLDL receptor-mediated TG-rich lipoprotein uptake by AMPK activation in skeletal muscle cells

Glucose and fatty acids are major energy sources in skeletal muscle. Very low-density lipoprotein receptor (VLDL-R), which is highly expressed in heart, skeletal muscle and adipose tissue, plays a crucial role in metabolism of triglyceride (TG)-rich lipoproteins. To explore energy switching between glucose and fatty acids, we studied expression of VLDL-R and lipoprotein uptake in rat L6 myoblasts. l-Glucose or d-glucose deprivation in the medium noticeably induced the AMPK (AMP-activated protein kinase) activation and VLDL-R expression. Dose-dependent induction of VLDL-R expression was observed when d-glucose was less than 4.2 mM. The same phenomenon was also observed in rat primary skeletal myoblasts and cultured vascular smooth muscle cells. The uptake of β-VLDL but not LDL was accompanied by induction of VLDL-R expression. Our study suggests that the VLDL-R-mediated uptake of TG-rich lipoproteins might compensate for glucose shortfall through AMPK activation in skeletal muscle.     

A viral noncoding RNA generated by cis-element-mediated protection against 5'->3' RNA decay represses both cap-independent and cap-dependent translation

Positive-strand RNA viruses use diverse mechanisms to regulate viral and host gene expression for ensuring their efficient proliferation or persistence in the host. We found that a small viral noncoding RNA (0.4 kb), named SR1f, accumulated in Red clover necrotic mosaic virus (RCNMV)-infected plants and protoplasts and was packaged into virions. The genome of RCNMV consists of two positive-strand RNAs, RNA1 and RNA2. SR1f was generated from the 3′ untranslated region (UTR) of RNA1, which contains RNA elements essential for both cap-independent translation and negative-strand RNA synthesis. A 58-nucleotide sequence in the 3′ UTR of RNA1 (Seq1f58) was necessary and sufficient for the generation of SR1f. SR1f was neither a subgenomic RNA nor a defective RNA replicon but a stable degradation product generated by Seq1f58-mediated protection against 5′→3′ decay. SR1f efficiently suppressed both cap-independent and cap-dependent translation both in vitro and in vivo. SR1f trans inhibited negative-strand RNA synthesis of RCNMV genomic RNAs via repression of replicase protein production but not via competition of replicase proteins in vitro. RCNMV seems to use cellular enzymes to generate SR1f that might play a regulatory role in RCNMV infection. Our results also suggest that Seq1f58 is an RNA element that protects the 3′-side RNA sequences against 5′→3′ decay in plant cells as reported for the poly(G) tract and stable stem-loop structure in Saccharomyces cerevisiae.     

Changes in immunological parameters in lung cancer patients undergoing immunotherapy with streptococcal preparation OK-432

Summary We have studied the immunological status of patients treated with streptococcal preparation OK-432. Two KE of OK-432 was injected intramuscularly once every week for more than three years unless the patients died. The natural killer (NK) activity in those patients who underwent curative surgery for lung cancer and had no sign of recurrence was significantly increased (P < 0.01) during the OK-432 treatment. However, the NK activity in the patients who had persistent disease (non-resected cases, incompletely resected cases or recurrent cases) was not significantly increased in comparison with that before the immunotherapy. Also, in the cases with no clinical symptoms of recurrence, both the lymphoblastogenetic reactions to the mitogens and the IL-2 production were significantly enhanced(P < 0.01) during the administration of OK-432. Reactions in the SU-PS (polysaccharide taken from the cell-wall fraction of theStreptococcus pyogenes SU strain and containing 7.2% of protein) skin-test appeared to significantly correlate with the immunological status of the patients under OK-432 therapy, but the PHA and PPD skin reactions showed no definitive enhancement. The survival rate of the patients whose SU-PS skin tests were positive during the OK-432 immunotherapy was significantly higher (P < 0.01) than that of the patients with negative reactions.     

Cryptic fragment alpha4 LG4-5 derived from laminin alpha4 chain inhibits de novo adipogenesis by modulating the effect of fibroblast growth factor-2

Cleavage of the extracellular matrix (ECM) by proteolysis unmasks cryptic sites and generates novel fragments with biological activities functionally distinct from those of the intact ECM molecule. The laminin G-like (LG)4-5 fragment has been shown to be excised from the laminin α 4 chain in various tissues. However, the functional role of this fragment has remained unknown to date. To investigate this, we prepared α 4 LG1-3 and α 4 LG4-5 fragments by elastase digestion of recombinant α 4 LG1-5, and examined their effects on de novo adipogenesis in mice at the site of injection of basement membrane extract (Matrigel) and fibroblast growth factor (FGF)-2. Although the addition of whole α 4 LG1-5 suppressed adipogenesis to some extent, the α 4 LG4-5 fragment could strongly suppress adipogenesis at a concentration of less than 20 n m . Addition of the α 4 LG4 module, which contains a heparin-binding region, had a suppressive effect, but this was lost in mutants with reduced heparin-binding activity. In addition, antibodies against the extracellular domain of syndecan-2 and -4, which are known receptors for the α 4 LG4 module, suppressed adipogenesis. Thus, these results suggest that the cryptic α 4 LG4-5 fragment derived from the laminin α 4 chain inhibits de novo adipogenesis by modulating the effect of FGF-2 through syndecans.     

Crystal structure of cold-active protein-tyrosine phosphatase from a psychrophile, Shewanella sp

The cold-active protein-tyrosine phosphatase (CAPTPase) of a psychrophile, Shewanella sp., shows high catalytic activity below 20 degrees C. The catalytic residue of CAPTPase is histidine, as opposed to the cysteine of known protein-tyrosine phosphatases (PTPases), and the enzyme protein has three amino acid sequences, Asp-Xaa-His, Gly-Asp-Xaa-Xaa-Asp-Arg and Gly-Asn-His-Glu, that are observed in many protein-serine/threonine phosphatases (PS/TPases). We have determined the crystal structures of CAPTPase at 1.82 angstroms and the enzyme bound with a phosphate ion at 1.90 angstroms resolution using X-ray crystallography and the multiple isomorphous replacement method. The final refined models are comprised of 331 amino acid residues, two metal ions, 447 water molecules, and an acetate or phosphate ion in an asymmetric unit. The enzyme protein consists of three beta-sheets, termed Sheet I, Sheet I', and Sheet II, and 14 alpha-helices. The CAPTPase has a different overall structure from known protein-tyrosine phosphatases. The arrangement of two metal ions, a phosphate ion and the adjacent amino acid residues in the catalytic site of CAPTPase is identical to that of PS/TPases. Thus, it was confirmed that the CAPTPase was a novel PTPase with a conformation similar to the catalytic site of PS/TPase. We speculate that the hydrophobic moiety around the catalytic residue of CAPTPase might play an important role in eliciting high activity at low temperature.     

HERC1 polymorphisms: population‐specific variations in haplotype composition

Human HERC1 is one of six HERC proteins and may play an important role in intracellular membrane trafficking. The human HERC1 gene is suggested to have been affected by local positive selection. To assess the global frequency distributions of coding and non‐coding single nucleotide polymorphisms (SNPs) in the HERC1 gene, we developed a new simultaneous genotyping method for four SNPs, and applied this method to investigate 1213 individuals from 12 global populations. The results confirmed remarked differences in the allele and haplotype frequencies between East Asian and non‐East Asian populations. One of the three common haplotypes observed was found to be characteristic of East Asians, who showed a relatively uniform distribution of haplotypes. Information on haplotypes would be useful for testing the function of polymorphisms in the HERC1 gene. This is the first study to investigate the distribution of HERC1 polymorphisms in various populations. Copyright © 2009 John Wiley & Sons, Ltd.     

The novel lipid raft adaptor p18 controls endosome dynamics by anchoring the MEK–ERK pathway to late endosomes

The regulation of endosome dynamics is crucial for fundamental cellular functions, such as nutrient intake/digestion, membrane protein cycling, cell migration and intracellular signalling. Here, we show that a novel lipid raft adaptor protein, p18, is involved in controlling endosome dynamics by anchoring the MEK1–ERK pathway to late endosomes. p18 is anchored to lipid rafts of late endosomes through its N‐terminal unique region. p18^?/? mice are embryonic lethal and have severe defects in endosome/lysosome organization and membrane protein transport in the visceral endoderm. p18^?/? cells exhibit apparent defects in endosome dynamics through perinuclear compartment, such as aberrant distribution and/or processing of lysosomes and impaired cycling of Rab11‐positive recycling endosomes. p18 specifically binds to the p14–MP1 complex, a scaffold for MEK1. Loss of p18 function excludes the p14–MP1 complex from late endosomes, resulting in a downregulation of the MEK–ERK activity. These results indicate that the lipid raft adaptor p18 is essential for anchoring the MEK–ERK pathway to late endosomes, and shed new light on a role of endosomal MEK–ERK pathway in controlling endosome dynamics.