Host immunoglobulin G titre and antibody activity in haemolymph of the tick, Ornithodoros moubata

Immunoglobulin G (IgG) in tick haemolymph was analysed immunochemically and biochemically for its antigenicity, antibody activity and relative concentration in a soft tick, Ornithodoros moubata (Murray) sensu Walton 1962 (Acari: Argasidae). Ouchterlony immunodiffusion tests showed that haemolymph from a tick engorged on rabbit IgG (or human IgG) through an artificial membrane, reacted with anti-rabbit IgG (anti-human IgG) but not with anti-human IgG (anti-rabbit IgG). This indicates that haemolymph of the fed tick contains IgG with a similar antigen specificity to host blood IgG. IgG from tick haemolymph was demonstrated by enzyme immunoassay to have the same antibody activity as ingested IgG. The IgG concentration in tick haemolymph was measured by a quantitative single immunodiffusion test. Changes of IgG titre after a bloodmeal were correlated with IgG activity, which was low for 5 days after a bloodmeal and then suddenly increased. The IgG titre reached a maximum 7 days post-engorgement, and remained high for over 4 months during and after oviposition. 125I-labelled IgG was injected into the tick haemocoel to determine the persistence of IgG in the haemolymph. Recovery of labelled IgG was low at 1 and 3 days, and high at 5, 8 and 16 days after engorgement. The data suggest that IgG in haemolymph disappears quickly soon after engorgement possibly by degradation and/or absorption (adhesion to tissues).     

Coordinated mRNA and Protein Expression of Human LAMP-1 in Induction of Melanogenesis After UV-B Exposure and Co-Transfection of Human Tyrosinase and TRP-1 cDNAs

In order to better understand the cascade of melanogenic events in melanocytes, this report has introduced our two recent approaches for the expression of melanogenesis/or melanosome-associated genes and encoded proteins in melanocytes (melanoma cells) after repeated exposure to UV -B and after cotransfection of two human genes, i.e., tyrosinase and tyrosinase-related protein-1 (TRP-1). Repeated exposure of UV B (2.5–5.0 mJ/cm²) caused not only upregulation of tyrosinase and TRP-1 genes but also coordinated increase in the gene and protein synthesis expression of Lamp-1 (lysosome-associated membrane protein-1). When COS-7 kidney cells and amelanotic melanoma (C32 and SKMEL-24) and melanotic melanoma (G361 and SK-MEL-23) cells were exposed to cotransfection of human tyrosinase and TRP-1 cDNAs, there was also an increased expression of Lamp-1 mRNA and protein along with tyrosinase activation and new melanin synthesis. Importantly, single transfectants of human tyrosinase cDNA revealed marked cellular degeneration, whereas this degeneration was not seen in single transfectants of TRP-1 cDNA or cotransfectants of human tyrosinase and TRP-1 cDNAs, indicating that TRP-1 prevented, along with Lamp-1, programmed death of melanocytes after transfection of tyrosinase gene. The coordinated expression of TRP-1 and Lamp-1 was further confirmed by antisense oligodeoxynucleotide hybridization experiment against Lamp-1 gene, showing the decreased expression of TRP-1 as identified by three different types of anti-TRP-1 monoclonal antibodies. We propose therefore that human tyrosinase and TRP-l, when activated or expressed together, will coordinate to upregulate the mRNA expression and protein synthesis of Lamp-1. The Lamp-1 molecules will, in turn, cover the inner surface of melanosomal membrane, together with TRP-1 molecules, thus protecting the melanosomal membrane from toxic melanin intermediates generated during melanogenesis in the presence of active tyrosinase. In contrast, the expression of other lysosome-related proteins, e.g., β-galactosidase and CD63 is not stimulated in new melanogenesis.     

Spectrophotometric Characterization of Eumelanin and Pheomelanin in Hair

Mammalian melanins exist in two chemically distinct forms: the brown to black eumelanins and the yellow to reddish-brown pheomelanins. They can be quantified by HPLC analysis of pyrrole-2,3,5-tricarboxylic acid (PTCA) and aminohydroxyphenylalanine (AHP). We recently developed a spectrophotometric method for assaying the total amount of eu- and pheomelanins by dissolving melanins in Soluene-350 plus water. In this study, we examined whether absorbance at 500 nm (A₅₀₀) of the Soluene-350 solution reflects the total amount of melanins obtained by the HPLC methods, and whether the ratio of absorbances between 650 and 500 nm reflects the eumelanin/total melanin ratio in mouse hair, sheep wool, and human hair. Our findings were as follows: (1) Total melanin levels calculated from A₅₀₀ values correlate well with those obtained from PTCA and AHP values by multiplying with the following factors: for mice, PTCA × 45 + AHP × 2.5; for sheep, PTCA × 40 + AHP × 15; and for humans, PTCA × 160 + AHP × 10. (2) The A₆₅₀/A₅₀₀ ratios were higher (0.25–0.33) in black to brown hair while they were significantly lower (0.10–0.14) in yellow to red hair. These results indicate that (1) the A₅₀₀ value can be used to quantify the total combined amount of eu- and pheomelanins, and (2) the A₆₅₀/A₅₀₀ ratio can serve as a parameter to estimate the eumelanin/total melanin ratio. The present method provides a convenient way to qualitatively characterize eu- and pheomelanins in melanins produced in follicular melanocytes.     

Biochemical Studies on the Acrosome Reaction of the Starfish, Asterias Amurensis I. Factors Participating in the Acrosome Reaction

In contrast with the case in sea urchin sperm, in starfish the acrosome reaction is not spontaneously induced by simply increasing the extracellular Ca²⁺ concentration or pH. At higher pHs, starfish sperm undergo morphological changes accompanied by exocytosis of the acrosomal vacuole, but they do not form acrosomal filaments. Nomarski-microscopic observation confirmed that spermatozoa undergo the acrosome reaction within the jelly coat. Acrosome reaction-inducing substance, a glycoprotein from the egg jelly, required a diffusible cofactor(s) present in the egg jelly for full activity. Several lines of evidence showed that this diffusible factor(s) is not merely Ca²⁺.     

S100 protein-immunoreactive trigeminal neurons innervating the rat molar tooth pulp

S100-immunoreactivity (ir) was examined in tooth pulp primary neurons of the rat. An immunofluorescence method demonstrated that the molar tooth pulp contained S100-immunoreactive (ir) nerve fibers. In the root pulp, pulp horn and roof of the pulp chamber, S100-ir smooth and varicose fibers ramified and formed subodontoblastic nerve plexuses. All the fibers became varicose at the base of the odontoblastic layer and extended to the odontoblastic layer. Some varicose endings could be traced into the dentin. The trigeminal neurons retrogradely labeled with fluorogold (FG) from the first and second maxillary molar tooth pulps exhibited S100- and parvalbumin-ir. Approximately 60% and 24% of the labeled cells were ir for S100 and parvalbumin, respectively. Virtually all parvalbumin-ir FG-labeled cells showed S100-ir, while 40% of S100-ir ones coexpressed parvalbumin-ir. An immunoelectron microscopic method revealed that all myelinated axons and half of the unmyelinated axons in the root pulp contained S100-ir. In the odontoblastic layer, predentin and dentin, S100-ir neurites lost the Schwann cell ensheathment and made close contact with cell bodies and processes of odontoblasts. The odontoblastic layer also contained parvalbumin-ir neurites. These neurites were devoid of the Schwann cell ensheathment and in close apposition to cell bodies and processes of odontoblasts. S100-ir pulpal axons seemed to be insensitive to repeated neonatal capsaicin treatment. This study suggests that S100-ir tooth pulp primary neurons are mostly myelinated and that S100-ir unmyelinated axons in the root pulp are preterminal segments of myelinated stem axons.     

Quantitative trait loci controlling aluminium tolerance in two accessions of Arabidopsis thaliana (Landsberg erecta and Cape Verde Islands)

Quantitative trait loci (QTL) analysis of aluminium (Al) tolerance was performed using Ler/Cvi recombinant inbred (RI) lines of Arabidopsis thaliana. Relative root length (RRL) (root length with 4 µm Al/root length with no Al at pH 5.0) on day 5 was used as the Al tolerance index for QTL analysis. Al tolerance judged by RRL was well correlated to tolerance judged by other indexes, including accumulation of callose, reactive oxygen species in the root apex and growth performance on acid soil containing a large amount of exchangeable Al. Using data sets with an h_b² of 0.91, two QTLs were detected at the top of chromosome 1 and bottom of chromosome 3. These QTLs explained 40 and 16% of the phenotypic variation of Al tolerance, respectively, and the positive effect of the Cvi allele. The QTL on chromosome 1 overlapped with a major QTL in another recombinant inbred population, and is possibly related to malate excretion. A complete pair-wise search revealed 11 sets of epistatic interacting loci pairs, which accounted for the transgressive segregation among the RI population. Several epistatic interactions shared the same chromosomal region, indicating the possible involvement of regulatory proteins in Al tolerance in Arabidopsis.     

DNA synthesis as related to the reappearance of "light interruption rhythm" in a long-day duckweed, Lemna gibba G3

DNA synthesis in the light perturbation period and its relationto the reappearance, due to light perturbation, of once faded-out"light interruption rhythm" in a long-day duckweed, Lemna gibbaG 3, were studied. After long continuous darkness, the duckweedincorporated ³H-thymidine into both nuclear and satellite DNA_sunder a light condition, but into satellite DNA alone undera dark condition. The number of dividing cells in frond epidermisincreased in proportion to the length of the light perturbationperiod. This increase was inhibited by 5-fluorodeoxyuridine.From these and previous results we conclude that nuclear DNAnewly synthesized in the light is intimately related with thereappearance of the rhythm. (Received June 15, 1970; )     

Ommochrome Deficiency in an Albino Strain of a Terrestrial Isopod,Armadillidium vulgare

In order to clarify the cause of ommochrome deficiency in an albino strain of the terrestrial isopod, Armadillidium vulgare, levels of xanthom-matin, 3-hydroxykynurenine, 3-hydroxyanthranilic acid and tryptophan in whole body extracts of the albino and the wild type individuals were determined together with enzyme activities of kynurenine-3-hydroxylase, kynureninase and tryptophan-2,3-dioxygenase. Xanthommatin could not be detected in the albinos. The levels of 3-hydroxykynurenine and 3-hydroxyanthranilic acid were determined by high-performance liquid chro-matography (HPLC) with electrochemical detection and were markedly low in the albinos compared with the wild type individuals. In contrast to those, the tryptophan levels determined by HPLC with fluorescence detection did not differ significantly between the two phenotypes. In the albino A. vulgare, kynurenine-3-hydroxylase activity was lower and kynureninase activity was higher than in the wild type, although the differences were not statistically significant. Tryptophan-2,3-dioxygenase activity in the albinos was less than 10% that in the wild type. Thus, ommochrome deficiency in the albino A. vulgare is considered to be caused by the extremely low activity of tryptophan-2,3-dioxygenase.     

基于Rag1基因序列变异的鲤形目鱼类系统发育重建:采样策略对生命之树的影响

The phylogenetic relationships of species are fundamental to any biological investigation, including all evolutionary studies. Accurate inferences of sister group relationships provide the researcher with an historical framework within which the attributes or geographic origin of species (or supraspecific groups) evolved. Taken out of this phylogenetic context, interpretations of evolutionary processes or origins, geographic distributions, or speciation rates and mechanisms, are subject to nothing less than a biological experiment without controls. Cypriniformes is the most diverse clade of freshwater fishes with estimates of diversity of nearly 3,500 species. These fishes display an amazing array of morphological, ecological, behavioral, and geographic diversity and offer a tremendous opportunity to enhance our understanding of the biotic and abiotic factors associated with diversification and adaptation to environments. Given the nearly global distribution of these fishes, they serve as an important model group for a plethora of biological investigations, including indicator species for future climatic changes. The occurrence of the zebrafish, Danio rerio, in this order makes this clade a critical component in understanding and predicting the relationship between mutagenesis and phenotypic expressions in vertebrates, including humans. With the tremendous diversity in Cypriniformes, our understanding of their phylogenetic relationships has not proceeded at an acceptable rate, despite a plethora of morphological and more recent molecular studies. Most studies are pre-Hennigian in origin or include relatively small numbers of taxa. Given that analyses of small numbers of taxa for molecular characters can be compromised by peculiarities of long-branch attraction and nodal-density effect, it is critical that significant progress in our understanding of the relationships of these important fishes occurs with increasing sampling of species to mitigate these potential problems. The recent Cypriniformes Tree of Life initiative is an effort to achieve this goal with morphological and molecular (mitochondrial and nuclear) data. In this early synthesis of our understanding of the phylogenetic relationships of these fishes, all types of data have contributed historically to improving our understanding, but not all analyses are complementary in taxon sampling, thus precluding direct understanding of the impact of taxon sampling on achieving accurate phylogenetic inferences. However, recent molecular studies do provide some insight and in some instances taxon sampling can be implicated as a variable that can influence sister group relationships. Other instances may also exist but without inclusion of more taxa for both mitochondrial and nuclear genes, one cannot distinguish between inferences being dictated by taxon sampling or the origins of the molecular data.