Intracellular Transport of Fat‐Soluble Vitamins A and E

Vitamins are compounds that are essential for the normal growth, reproduction and functioning of the human body. Of the 13 known vitamins, vitamins A, D, E and K are lipophilic compounds and are therefore called fat‐soluble vitamins. Because of their lipophilicity, fat‐soluble vitamins are solubilized and transported by intracellular carrier proteins to exert their actions and to be metabolized properly. Vitamin A and its derivatives, collectively called retinoids, are solubilized by intracellular retinoid‐binding proteins such as cellular retinol‐binding protein (CRBP), cellular retinoic acid‐binding protein (CRABP) and cellular retinal‐binding protein (CRALBP). These proteins act as chaperones that regulate the metabolism, signaling and transport of retinoids. CRALBP‐mediated intracellular retinoid transport is essential for vision in human. α‐Tocopherol, the main form of vitamin E found in the body, is transported by α‐tocopherol transfer protein (α‐TTP) in hepatic cells. Defects of α‐TTP cause vitamin E deficiency and neurological disorders in humans. Recently, it has been shown that the interaction of α‐TTP with phosphoinositides plays a critical role in the intracellular transport of α‐tocopherol and is associated with familial vitamin E deficiency. In this review, we summarize the mechanisms and biological significance of the intracellular transport of vitamins A and E.      

Analysis of Δ12-fatty acid desaturase function revealed that two distinct pathways are active for the synthesis of PUFAs in T. aureum ATCC 34304

Thraustochytrids are known to synthesize PUFAs such as docosahexaenoic acid (DHA). Accumulating evidence suggests the presence of two synthetic pathways of PUFAs in thraustochytrids: the polyketide synthase-like (PUFA synthase) and desaturase/elongase (standard) pathways. It remains unclear whether the latter pathway functions in thraustochytrids. In this study, we report that the standard pathway produces PUFA in Thraustochytrium aureum ATCC 34304. We isolated a gene encoding a putative Δ12-fatty acid desaturase (TauΔ12des) from T. aureum. Yeasts transformed with the tauΔ12des converted endogenous oleic acid (OA) into linoleic acid (LA). The disruption of the tauΔ12des in T. aureum by homologous recombination resulted in the accumulation of OA and a decrease in the levels of LA and its downstream PUFAs. However, the DHA content was increased slightly in tauΔ12des-disruption mutants, suggesting that DHA is primarily produced in T. aureum via the PUFA synthase pathway. The transformation of the tauΔ12des-disruption mutants with a tauΔ12des expression cassette restored the wild-type fatty acid profiles. These data clearly indicate that TauΔ12des functions as Δ12-fatty acid desaturase in the standard pathway of T. aureum and demonstrate that this thraustochytrid produces PUFAs via both the PUFA synthase and the standard pathways.     

Galectin-8 and galectin-9 are novel substrates for thrombin

Galectin-8 and galectin-9, which each consist of two carbohydrate recognition domains (CRDs) joined by a linker peptide, belong to the tandem-repeat-type subclass of the galectin family. Alternative splicing leads to the formation of at least two and three distinct splice variants (isoforms) of galectin-8 and galectin-9, respectively, with tandem-repeat-type structures. The isoforms share identical CRDs and differ only in the linker region. In a search for differences in biological activity among the isoforms, we found that their isoforms with the longest linker peptide, that is, galectin-8L and galectin-9L (G8L and G9L), are highly susceptible to thrombin cleavage, whereas the predominant isoforms, galectin-8M and galectin-9M (G8M and G9M), and other members of human galectin family so far examined were resistant to thrombin. Amino acid sequence analysis of proteolytic fragments and site-directed mutagenesis showed that the thrombin cleavage sites (-IAPRT- and -PRPRG- for G8L and G9L, respectively) resided within the linker peptides. Although intact G8L stimulated neutrophil adhesion to substrate more efficiently than G8M, the activity of G8L but not that of G8M decreased on thrombin digestion. Similarly, thrombin treatment almost completely abolished eosinophil chemoattractant (ECA) activity of G9L. These observations suggest that G8L and G9L play unique roles in relation to coagulation and inflammation.     

Clinical significance on MicroScan Rapid plus series using various antibiotic-resistant bacteria

MicroScan Rapid plus Neg II Series and MicroScan Rapid plus Pos Series by Siemens Healthcare Diagnostics K.K. are the panels which enable to measure identification and antimicrobial susceptibility testing quickly and we have confirmed that it is useful for detecting drug resistance bacteria. As the identification result of comparing Rapid plus series with the current panel by using 143 strains of various drug resistance bacteria, Gram positive cocci was 87. 7%, glucose fermenter was 100% and glucose non-fermenter was 77.3 in Gram negative bacilli. On the evaluation of antimicrobial susceptibility testing, Rapid plus series, in comparison with the current panel, confirmed the lower tendency of MIC value on some drugs, but it basically presented the good concordance rate. In terms of the reporting time of antimicrobial agent, non-fermenter or MRCNS reported the result as needed after 8 hours and it took a little longer time for the report of antimicrobial agent. On the other hand, 80% or higher of antimicrobial agent on panel was reported for intestinal bacteria in 4.5 hours and for MRSA in 6.5 hours. It enabled to report the testing result on the same day. Due to the results above, Rapid plus series was highly valued on the usability, such as the early detection of drug resistance bacteria and the selection of therapeutic agents.     

Evicting cuckoo nestlings from the nest: a new anti-parasitism behaviour

As avian brood parasitism usually reduces hosts'' reproductive success, hosts often exhibit strong defence mechanisms. While such host defences at the egg stage (especially egg rejection) have been extensively studied, defence mechanisms at the nestling stage have been reported only recently. We found a previously unknown anti-parasitism behaviour in the large-billed Gerygone, which is a host species of the little bronze-cuckoo, a host-evicting brood parasite. The hosts forcibly pulled resisting nestlings out of their nests and dumped them. Although it has been suggested that defence mechanisms at the nestling stage may evolve when host defence at the egg stage is evaded by the parasite, the studied host seems to lack an anti-parasitism strategy at the egg stage. This suggests that the evolutionary pathway may be quite different from those of previously studied cuckoo–host systems. Future research on this unique system may give us new insights into the evolution of avian brood parasitism.     

Prediction of liquid chromatographic retention times of peptides generated by protease digestion of the Escherichia coli proteome using artificial neural networks

We developed a computational method to predict the retention times of peptides in HPLC using artificial neural networks (ANN). We performed stepwise multiple linear regressions and selected for ANN input amino acids that significantly affected the LC retention time. Unlike conventional linear models, the trained ANN accurately predicted the retention time of peptides containing up to 50 amino acid residues. In 834 peptides, there was a strong correlation (R2 = 0.928) between measured and predicted retention times. We demonstrated the utility of our method by the prediction of the retention time of 121,273 peptides resulting from LysC-digestion of the Escherichia coli proteome. Our approach is useful for the proteome-wide characterization of peptides and the identification of unknown peptide peaks obtained in proteome analysis.     

SPLITS: a new program for predicting split and intron-containing tRNA genes at the genome level

In the archaea, some tRNA precursors contain intron(s) not only in the anticodon loop region but also in diverse sites of the gene (intron-containing tRNA or cis-spliced tRNA). The parasite Nanoarchaeum equitans, a member of the Nanoarchaeota kingdom, creates functional tRNA from separate genes, one encoding the 5'-half and the other the 3'-half (split tRNA or trans-spliced tRNA). Although recent genome projects have revealed a huge amount of nucleotide sequence data in the archaea, a comprehensive methodology for intron-containing and split tRNA searching is yet to be established. We therefore developed SPLITS, which is aimed at searching for any type of tRNA gene and is especially focused on intron-containing tRNAs or split tRNAs at the genome level. SPLITS initially predicts the bulge-helix-bulge splicing motif (a well-known, required structure in archaeal pre-tRNA introns) to determine and remove the intronic regions of tRNA genes. The intron-removed DNA sequences are automatically queried to tRNAscan-SE. SPLITS can predict known tRNAs with single introns located at unconventional sites on the genes (100%), tRNAs with double introns (85.7%), and known split tRNAs (100%). Our program will be very useful for identifying novel tRNA genes after completion of genome projects. The SPLITS source code is freely downloadable at http://splits.iab.keio.ac.jp/.