Suppression of the degradation of recombinant human apolipoprotein E by a protease inhibitor obtained from fetal bovine serum in serum-free culture

The recombinant human apolipoprotein E (Apo-E) produced by Chinese hamster ovary cells (CHO-322 cells) in serum free culture was degraded to 24K and 23K fragments that contained N-terminal amino acid. The degradation site of Apo-E to 24K fragment was between Arg180 and Leu181 and the C-terminal amino acid of 23K fragment was Gly169. In fetal bovine serum (FBS)-containing culture, the degradation was inhibited. However, in calf serum (CS) the inhibitory activity was not detected. Thus, we attempted the purification of the factor with this inhibitory activity from FBS. A protease inhibitor was purified to give a single peak from FBS by ammonium sulfate precipitation and combination of several column chromatographies. When this FBS-derived protease inhibitor (FBS-d-PI) was added to serum-free culture of CHO-322 cells, degradation of recombinant Apo-E to the 24K and 23K fragments was dose-dependently suppressed and accumulation of intact Apo-E in culture supernatant was observed. FBS-d-PI was found to be a glycoprotein with relative molecular size of 75K daltons under reducing condition, and 85K daltons under nonreducing condition by SDS-PAGE. A complex of FBS-d-PI and a cellular protease was also detected in culture supernatant by western blot analysis using mouse monoclonal antibodies against FBS-d-PI.     

稻瘟菌诱导的水稻几丁酶、β-1.3-葡聚糖酶活性及分布

稻瘟菌(Pyricularia oryzae C.)孢子感染或菌丝培养液处理后,水稻的几丁酶和β-1,3-葡聚糖酶活力被分别诱导提高6～9倍和3～5倍。用亲和层析初步纯化的几丁酶在体外能降解稻瘟菌细胞壁和抑制它的孢子萌发。这两个酶在细胞间隙和细胞内都有分布。     

Developmental interactions between the army wormLeucania separata [Lep.: Noctuidae] and its parasiteApanteles ruficrus [Hym.: Braconidae]

The developmental interactions between the gregarious endoparasitoidApanteles ruficrus Hal. and the army worm,Leucania separata Walker were investigated. The parasitoid preferred young host larvae and developed in 9.5 days irrespective of host age at the time of parasitization. The growth of parasitized host larvae were depressed. The net maximum weight of the host larva was positively correlated with the number of parasitoid eggs laid when the 2nd instar was parasitized. And when parasitizing in 2nd instar, the weight of parasitoid was negatively correlated with the number of eggs laid. The parasitoid has an ability to regulate the size of the host and the parasitoid itself according to the number of eggs laid when the host larva is very small.     

Utilization and requirements of amino acids by a cell line derived from the butterfly Papilio xuthus

The media, in which a butterfly cell line (Px 58), derived from pharate adult ovaries of Papilio xuthus cultured for 8 days, were analysed to examine the changes in free amino acids in the medium during cultivation. Beta-alanine, arginine, glycine, histidine, lysine, phenylalanine, proline, serine, and tryptophan did not change markedly. Asparagine, aspartic acid, cystine, glutamine, isoleucine, leucine, methionine, threonine, tyrosine, and valine decreased to some extent with culturing. Alpha-alanine increased markedly, and glutamic acid did so to a lesser extent. Requirements of amino acids by the cell line were examined by deleting amino acids one at a time. Deletion of alpha-alanine, beta-alanine, asparagine, glutamic acid, glycine, and phenylalanine did not cause deterioration of the cell. These amino acids were thought to be non-essential or required only a little. Deletion of other amino acids impaired the cell growth severely. These amino acids would appear to be essential for growth of the Px 58 cell line.     

Cryopreservation of infective larvae of Dipetalonema viteae.

infective larvae of Dipetalonema viteae produced infections in Mongolian jirds (Meriones unguiculatus) after storage of infected ticks (Ornithodoros tartakovskyi) in the presence of dimethyl sulfoxide (DMSO, 5%) for 7 or 595 days in liquid nitrogen (-196 C). Infectivity of these larvae was only partially impaired. Microfilaremias of test jirds were generally lower than those of control jirds given nonfrozen larvae; however, the majority of test jirds developed microfilarial counts suitable for use in infecting ticks. In contradistinction, larvae frozen free of the tick failed to retain infectivity. Apparently the tick, in conjunction with DMSO, protects the larvae during freezing and thawing.     

Experimental Approach Reveals the Role of alx1 in the Evolution of the Echinoderm Larval Skeleton

Over the course of evolution, the acquisition of novel structures has ultimately led to wide variation in morphology among extant multicellular organisms. Thus, the origins of genetic systems for new morphological structures are a subject of great interest in evolutionary biology. The larval skeleton is a novel structure acquired in some echinoderm lineages via the activation of the adult skeletogenic machinery. Previously, VEGF signaling was suggested to have played an important role in the acquisition of the larval skeleton. In the present study, we compared expression patterns of Alx genes among echinoderm classes to further explore the factors involved in the acquisition of a larval skeleton. We found that the alx1 gene, originally described as crucial for sea urchin skeletogenesis, may have also played an essential role in the evolution of the larval skeleton. Unlike those echinoderms that have a larval skeleton, we found that alx1 of starfish was barely expressed in early larvae that have no skeleton. When alx1 overexpression was induced via injection of alx1 mRNA into starfish eggs, the expression patterns of certain genes, including those possibly involved in skeletogenesis, were altered. This suggested that a portion of the skeletogenic program was induced solely by alx1. However, we observed no obvious external phenotype or skeleton. We concluded that alx1 was necessary but not sufficient for the acquisition of the larval skeleton, which, in fact, requires several genetic events. Based on these results, we discuss how the larval expression of alx1 contributed to the acquisition of the larval skeleton in the putative ancestral lineage of echinoderms.     

Extensive size homoplasy at a microsatellite locus in the Japanese bumblebee, Bombus diversus

An extensive size homoplasy was found at microsatellite locus B11 of the bumblebee, Bombus diversus, in northern to central Honshu, Japan. A total of 16 alleles of different nucleotide sequences in five length morphs was obtained at B11 for this species. Of these alleles, five were 141 base pairs (bp) in length, five were 137 bp and four were 133 bp. Allele diversity in each length morph was high compared with previous studies. It is noteworthy that this extensive size homoplasy was found in a relatively small geographic area, in contrast to results from previous studies. Reconstruction of a median‐joining network revealed the complicated evolutionary process of the locus, involving insertion/deletion and point mutations. Preliminary estimation of the mutation rate of the B11 locus in B. diversus gives a value comparable to those estimated from experimental Drosophila populations. Effects of the extensive size homoplasy in population genetic studies is discussed.