Leucine aminopeptidase during meiotic development.

We found a leucine aminopeptidase (LAP; EC 3.4.11.1) to be abundant in meiotic prophase tissue of a basidiomycete, Coprinus cinereus. After direct purification of the aminopeptidase component from meiocytes, we cloned the gene by degenerate PCR using partial amino-acid sequences of the purified enzyme and 5' and 3' RACE. It was homologous to the eukaryotic leucine aminopeptidase gene. The recombinant protein possesses the characteristic activities of a Coprinus leucine aminopeptidase (CoLAP) with a molecular mass of 52.4 kDa, and forms a homohexamer. Northern blot and spatial distribution analysis by immunohistochemical staining indicated CoLAP to be abundant in meiotic prophase cells and the supporting cells around meiocytes, but scarce in mycelium cells. Interestingly, from zygotene to pachytene, CoLAP was mostly present in supporting cells around meiocytes, but from diplotene onwards, it was plentiful in meiocytes themselves, suggesting that its expression is required to control some of the biochemical events at meiotic prophase. Moreover, the strong expression of CoLAP mRNA immediately after treatment with methyl methanesulfonate in mycelium implies that CoLAP has a role in somatic DNA repair.     

Short-step syntheses of naturally occurring polyoxygenated aromatics based on site-selective transformation

Wogonin and astringin were synthesized from inexpensive chrysin and piceid in short steps. The key feature of these syntheses is site-selective transformation. The target molecules were obtained in 27 and 62% yields from the starting materials, respectively.     

Tim4 recognizes carbon nanotubes and mediates phagocytosis leading to granuloma formation

1. Download : Download high-res image (187KB)
2. Download : Download full-size image

     

Improved preparation of vitexin from hot water extract of Basella alba,the commercially available vegetable Malabar spinach (“Tsurumurasaki” in Japanese) and the application to semisynthesis of chafuroside B

ABSTRACT

Hot water extraction of D-arabinofuranosylvitexin from the raw leaves of commercially available Basella alba “Tsurumurasaki” and subsequent acidic hydrolysis was improved to be a procedure using a high-pressure steam sterilizer to afford vitexin. The amount was estimated to be 14.1 mg from 1 g of dry weight of the raw leaves, whose recovery was calculated to be 95% based on the estimated content of D-arabinofuranosylvitexin in B. alba raw leaves. The product was dehydratively cyclized between hydroxy groups on the carbohydrate and flavone skeletons under modified Mitsunobu reaction conditions in N,N-dimethylformamide to give chafuroside B, which is known to be a bioactive Oolong tea polyphenol. Through these transformations, 10.2 mg of chafuroside B could be semisynthesized from 1 g of dry weight of the raw leaves, and the efficiency was improved compared to that from the extraction from Oolong tea (3.4 μg from 1 g of dry weight).     

Directed acyclic graph kernels for structural RNA analysis

Background  

Recent discoveries of a large variety of important roles for non-coding RNAs (ncRNAs) have been reported by numerous researchers. In order to analyze ncRNAs by kernel methods including support vector machines, we propose stem kernels as an extension of string kernels for measuring the similarities between two RNA sequences from the viewpoint of secondary structures. However, applying stem kernels directly to large data sets of ncRNAs is impractical due to their computational complexity.     

Structure and expression properties of the endo-β-1,4-glucanase A gene from the filamentous fungus Aspergillus nidulans

Endo-beta-1,4-glucanase A (EG A) of Aspergillus nidulans was purified to homogeneity, and its genomic gene (eglA) was cloned based on partial amino acid sequences of the purified enzyme and sequenced. The eglA gene comprised 1228 bp with four putative introns and encoded a polypeptide of 326 amino acids bearing high homology to the family A cellulases. The eglA promoter activity in A. nidulans was examined using the A. oryzae Taka-amylase A gene as a reporter. Expression of the reporter gene was induced by carboxymethylcellulose and cellobiose, and repressed by glucose, galactose, mannose, xylose, sorbitol, glycerol and succinate. Lactose neither induced nor repressed the expression.     

Gene cloning of the pink-colored protein from Pleurotus salmoneostramineus (PsPCP) and its species-specific chromoprotein are effective for colorization of the fruit body

Pleurotus salmoneostramineus is a pink mushroom. This pink color is a protein and forms a complex with 3H-indol-3-one. The gene encoding the pink-colored protein from P. salmoneostramineus (PsPCP) was cloned, and its sequence was elucidated as a 681-bp. The ORF encodes 226 amino acid residues. The amino acid sequence of the protein did not show any significant homology in the DDBJ/EMBL/GenBank databases. Recombinant PsPCP was expressed as the soluble form in E. coli. The reaction mixture of purified recombinant PsPCP and 3H-indol-3-one showed a pink color as the native pigment. A real-time PCR analysis revealed the strong expression of PsPCP in the primordium formation stage of the life cycle of the fungus; however, its expression decreased with the maturation of the fruit body. A comparison of PsPCP gene expression profiles between two strains revealed high levels in the dark-colored strain.     

Ddhd1 knockout mouse as a model of locomotive and physiological abnormality in familial spastic paraplegia

We have previously reported a novel homozygous 4-bp deletion in DDHD1 as the responsible variant for spastic paraplegia type 28 (SPG28; OMIM#609340). The variant causes a frameshift, resulting in a functionally null allele in the patient. DDHD1 encodes phospholipase A₁ (PLA₁) catalyzing phosphatidylinositol to lysophosphatidylinositol (LPI). To clarify the pathogenic mechanism of SPG28, we established Ddhd1 knockout mice (Ddhd1[−/−]) carrying a 5-bp deletion in Ddhd1, resulting in a premature termination of translation at a position similar to that of the patient. We observed a significant decrease in foot–base angle (FBA) in aged Ddhd1(−/−) (24 months of age) and a significant decrease in LPI 20:4 (sn-2) in Ddhd1(−/−) cerebra (26 months of age). These changes in FBA were not observed in 14 months of age. We also observed significant changes of expression levels of 22 genes in the Ddhd1(−/−) cerebra (26 months of age). Gene Ontology (GO) terms relating to the nervous system and cell–cell communications were significantly enriched. We conclude that the reduced signaling of LPI 20:4 (sn-2) by PLA₁ dysfunction is responsible for the locomotive abnormality in SPG28, further suggesting that the reduction of downstream signaling such as GPR55 which is agonized by LPI is involved in the pathogenesis of SPG28.