Genetic modification of lignin biosynthesis for improved biofuel production

The energy in cellulosic biomass largely resides in plant cell walls. Cellulosic biomass is more difficult than starch to break down into sugars because of the presence of lignin and the complex structure of cell walls. Transgenic down-regulation of major lignin genes led to reduced lignin content, increased dry matter degradability, and improved accessibility of cellulases for cellulose degradation. This review provides background information on lignin biosynthesis and focuses on genetic manipulation of lignin genes in important monocot species as well as the dicot potential biofuel crop alfalfa. Reduction of lignin in biofuel crops by genetic engineering is likely one of the most effective ways of reducing costs associated with pretreatment and hydrolysis of cellulosic feedstocks, although some potential fitness issues should also be addressed.     

Carbohydrate structures of human interleukin 5 expressed in Chinese hamster ovary cells.

The asparagine-linked sugar chains of recombinant human interleukin 5 produced by Chinese hamster ovary cells were released quantitatively as oligosaccharides by hydrazinolysis. After N-acetylation followed by NaB3H4 reduction, each oligosaccharide was isolated by paper electrophoresis and serial lectin column chromatography. Study of their structures by sequential exoglycosidase digestion in combination with methylation analysis, revealed that they are bi-, tri-, and tetraantennary complex-type with fucosylated and non-fucosylated trimannosyl cores and high mannose type sugar chains. More than 80% of the sugar chains occur as biantennary complex-type sugar chains. Although acidic oligosaccharides amount to only 14% of the total oligosaccharides, their sialic acid residues occur exclusively as the Sia alpha 2----3Gal group. Removal of the sugar moiety from intact recombinant human interleukin 5 produced a 2.5-fold increase of its activity to induce IgM secretion.     

Fluorescence polarization immunoassay employing immobilized antibody

The use of an antibody immobilized on latex or silver colloid in fluorescence polarization immunoassay (FPI) is assessed. In FPI it is possible to detect antigens of high molecular weight because the molecular weight of the antibody is effectively increased. In the assay for rabbit immunoglobulin G a limit of detection lower by two orders of magnitude and an assay range wider by one order of magnitude can be obtained in comparison with conventional FPI. The detection limit is 10(-10) mol l-1 and the total assay time for one sample is 8 min. This assay combines a low detection limit with a short assay time.     

Comparative studies of the biological activities of human tumor necrosis factor and its derivatives

Biological activities of human tumor necrosis factor (TNF) and its derivatives were compared. In cytotoxicity assay with L929 cells, one derivative, designated as TNF(Asn), showed significantly lower activity than any other TNF examined. In binding assay, this derivative was also shown to have lower affinity for TNF receptors on L929 cells, suggesting that the cytotoxic activity of TNFs on L929 cells correlates with their affinity for receptors. We also found that the cytotoxic activity of TNF on A673 cells and its inhibitory effect on lipoprotein lipase were parallel with the cytotoxic activity on L929 cells, but the growth-enhancing activity on FS-4 cells and the cytotoxic activity on endothelial cells were not. It was also shown that TNF(Asn) had lower affinity than any other TNF for receptors on these target cells tested. These results suggested that there might be at least two types of cellular responses to TNF; one might correlate with the receptor-binding affinity of TNFs and the other not.     

High level expression of a frog α-amidating enzyme,AE-II,in cultured cells and silkworm larvae using aBombyx mori nuclear polyhedrosis virus expression vector

AXenopus laevis peptidyl C-terminal α-amidating enzyme (AE-II) gene, modified by deletion of a region encoding the putative membrane-spanning domain and the putative C-terminal cytosolic tail, was expressed in BoMo-15 AIIc insect cells and silkworm larvae using aBombyx mori baculovirus expression vector system. The expressed enzyme was identified predominantly in the culture medium and the hemolymph of silkworm larvae, indicating successful secretion of the expressed AE-II. The level of recombinant enzyme in the larval hemolymph at 4 days post-infection (40 μg/ml) was more than 100-fold the peak levels found in the culture medium (250 ng/ml). The enzyme activity in the larval hemolymph at 4 days post-infection was 3700 units/ml.     

Neurosecretory cell types in the eyestalk of the freshwater prawn Palaemon paucidens

Summary In the medulla terminalis ganglionic X-organ (MTGX) of the eyestalk of the freshwater prawn, Palaemon paucidens, six peptidergic neuro-secretory cell types (A-, B-, C-, D-, E-, and F-cells) are distinguishable on the basis of the different morphology of their elementary granules and rough endoplasmic reticulum (rER). All of these cell types seem to correspond to Type-IIIa cells or dispersing Type-IV cells, that have previously been differentiated at the light microscopic level (Hisano, 1974), as judged from the dimensions of their cell bodies and nuclei. Two other peptidergic neurosecretory cell types that are apparently comparable to the Type-II and Type-IIIb cells (Hisano, 1974), respectively, are recognized in parts of the optic ganglia other than MTGX, and these are now designated as Gand H-cells, respectively. All the remaining cell types, designated as Type-I, cluster-forming Type-IV, Type-V and Type-VI cells in our previous light microscopic study, have small cored-vesicles in their cytoplasm. It remains undecided whether these, possibly aminergic, neurons are neurosecretory or not.The author wishes to express his sincere appreciation to Prof. T. Aoto for his invaluable advice during the course of this study.     

Systemic activation of macrophages by liposome-entrapped muramyl tripeptide in mice pretreated with the chemotherapeutic agent adriamycin

Summary The purpose of these studies was to determine whether macrophages of mice pretreated with the chemotherapeutic agent adriamycin (ADR) could be systemically activated by IV injection of liposomes containing muramyl tripeptide phosphatidylethanolamine (MTP-PE), a lipophilic derivative of muramyl dipeptide. Lower than normal levels of alveolar macrophages or peritoneal exudate macrophages were found in mice following IV injection of ADR. This decrease was dose-dependent and, in mice given <10 mg ADR/kg, it was transient (14 days). Peritoneal macrophages surviving the administration of 15 mg ADR/kg were tumoricidal.At various times after single or repeated administration of ADR, mice were given IV or IP injections of liposomes containing MTP-PE. One day thereafter, the cytotoxic activity of the in situ-activated macrophages (alveolar or peritoneal exudate) was assessed in culture against syngeneic melanoma cells. Our data demonstrate that under defined conditions the systemic administration of ADR does not interfere with the in situ activation of tumoricidal properties of murine macrophages after IV injection of liposomes containing a macrophage-activating agent.     

Correction to: Rhamphocottus nagaakii (Cottoidea: Rhamphocottidae), a new species of grunt sculpin from the northwestern Pacific,with notes on the phylogeography of the genus Rhamphocottus

A new species of grunt sculpin, Rhamphocottus nagaakii inhabiting the northwestern Pacific, previously identified as Rhamphocottus richardsonii Günther 1874, is described based on genetic evidence and morphological differences. The new species can be distinguished based on morphometric characters related to the head, including head length [45.3–54.6% of standard length (SL)], postorbital head length (18.8–25.5% SL) and the length of pectoral-fin base (15.8–20.7% SL), which are smaller than in R. richardsonii (53.6–60.5% SL, 26.2–31.7% SL, and 19.5–25.2% SL, respectively). Genetic differences between two species markedly exceed levels for intra-specific differences. Rhamphocottus nagaakii is considered to have arisen from a common ancestor of the two species, which probably inhabited somewhere the North Pacific Rim around the Aleutian Archipelago. During a period of cooling in the Pliocene or the Miocene, R. nagaakii and R. richardsonii became separated to the southern regions of the northwestern and northeastern Pacific, and subsequently underwent speciation.

     

Synaptic regulation of paraventricular arginine vasopressin-containing neurons by neuropeptide Y-containing monoaminergic neurons in rats

Summary Synaptic regulation of arginine vasopressin (AVP)-containing neurons by neuropeptide Y (NPY)-containing monoaminergic neurons was demonstrated in the paraventricular nucleus of the rat hypothalamus. NPY and AVP were immunolabeled in the pre- and the post-embedding procedures, respectively, and monoaminergic fibers were marked by incorporating 5-hydroxydopamine (5-OHDA), a false neurotransmitter. The immunoreaction for NPY was expressed by diaminobenzidine (DAB) chromogen, and that for AVP by gold particles. The DAB chromogen was localized on the surface of the membrane structures, such as vesicles or mitochondria, and on the core of large cored vesicles. Gold particles were located on the core of the secretory granules within the AVP cell bodies and processes. The incorporated 5-OHDA was found as dense cores within small or large vesicular structures. From these data, three types of nerve terminals were discernible: NPY-containing monoaminergic, NPY-containing non-aminergic, and monoaminergic fibers. The AVP cell bodies appeared to have synaptic junctions formed by these nerve terminals as well as by the unlabeled nerve terminals which have small clear vesicles and large cored vesicles. These different types of nerve terminals were frequently observed in a closely apposed position on the same AVP cell bodies. The functional relationships of these three types of neuronal terminals are discussed.