Identification of Mutations in the Pigment Cell-Specific Gene Located at the Brown Locus in Mouse

The pigment cell-specific gene, located at the brown (b)-locus in mouse, encodes the protein that determines the type of melanin synthesized. This protein is known as tyrosinase-related protein, but here we tentatively term it b-locus protein to avoid confusions with the related sequence cross-hybridizing to the tyrosinase gene. In order to identify the mutation at the b-locus, we have cloned and characterized the b-locus protein gene of BALB/c mouse (b/b, c/c). The gene is about 18 kb long and organized into 8 exons and 7 introns. Sequence analysis of the b-locus protein gene reveals four base changes within the protein-coding regions: two missense mutations and two silent mutations. Two missense mutations result in the Cys to Tyr substitution at position 86 (codon 110) and the Arg to His substitution at position 302 (codon 326) of a b-locus protein molecule. Using allele-specific amplification, we confirmed that these missense mutations are actually present in the genomic DNA of two b-mutant strains examined, BALB/c and DBA/2 (b/b, C/C) mice, suggesting that these mutations are specific for the mutant mice at the b-locus. Moreover, we are able to show that the b-locus protein containing Tyr 86 is not reactive with the anti-b-locus protein monoclonal antibody, TMH-1, in transient expression assays.     

ALLOZYME DIVERSITY AND GENETIC IDENTITY IN SCHIEDEA AND ALSINIDENDRON (CARYOPHYLLACEAE: ALSINOIDEAE) IN THE HAWAIIAN ISLANDS

Genetic diversity of allozymes, genetic identity based on allozyme variability, and phylogenetic relationships were studied with respect to breeding system diversity, population size, and island age in 20 of the 29 species of Schiedea and Alsinidendron (Caryophyllaceae: Alsinoideae), a monophyletic lineage endemic to the Hawaiian Islands. Average levels of genetic variability in Schiedea and Alsinidendron were comparable to or higher than those found in other Hawaiian lineages for which equivalent data are available [Bidens, Tetramolopium, and the silversword alliance (Asteraceae: Madiinae)] and similar to average values for species of dicots. Allozyme variability was strongly dependent on breeding system, which varies widely in the Hawaiian Alsinoideae. Species with autogamous breeding systems showed very low variability, measured as the number of alleles per locus, percent polymorphic loci, and mean heterozygosity per locus. Outcrossing hermaphroditic and dimorphic species (those with gynodioecious, subdioecious, and dioecious breeding systems) showed significantly higher genetic variability. Small population size was associated with lower values for all measures of genetic variability. Nearly half of the species occurring in small populations are also autogamous; thus, both factors may have influenced levels of genetic variability in these species. Founder effect was apparent in one species (Schiedea adamantis), which occurs in a single large population, has a gynodioecious breeding system but a very low genetic variability. Island age appeared to have little effect on genetic variability. Slightly lower values of genetic variability for species occurring on Kaua'i and O'ahu result primarily from the occurrence of autogamous Alsinidendron species on those islands. Values for Nei's genetic identity for different species pairs were 0.201–0.942, a far greater range than in Bidens, the silversword alliance, and Tetramolopium. Using UPGMA clustering, there was only moderate support for relationships detected through cladistic analysis. Nei's unbiased genetic identity (I) was greatest among species with outcrossing breeding systems, which for the most part clustered together. Nei's genetic identities for self-fertilizing species were low, indicating that these species are less similar to one another and to outcrossing species, regardless of their affinities based on cladistic analysis. Parsimony analysis of allele frequency data supported two clades also found in phylogenetic analyses using morphological and molecular data. Clades recognized in parsimony analysis of allele frequencies were those lineages containing selfing species, indicating that conditions favoring fixation of alleles occurred in ancestral species. In contrast, maintenance of high genetic diversity in outcrossing species interferes with recognition of phylogenetic relationships using allozyme variability.     

biparous:A Novel bHLH Gene Expressed in Neuronal and Glial Precursors inDrosophila

Genetic studies have uncovered many genes that are involved in the first steps of neuronal development inDrosophila.Less is known about the intermediate steps during which individual precursor cells follow either the neuronal pathway or the glial pathway. We report the identification of a novel bHLH gene,biparous,expressed in neuronal and glial precursors inDrosophila.Unlike most bHLH genes,biparousexpression continues to the final stages of neurogenesis in the embryo. Expression ofbiparousis not observed in end stage postmitotic neurons and precedes the expression ofrepo,a gene activated in later stages of glial differentiation. The bHLH domain is sufficiently different from previously described bHLH domains to imply a novel function.     

Comparison of muscle force,muscle endurance,and electromyogram activity during an expedition at high altitude

Handgrip force (HF), maximal pinch force (MF), muscle endurance (ME), and the median power frequency (MdPF) of the activity shown in the electromyogram (EMG) were studied at various altitudes in eight normal healthy subjects. MF and ME were measured between the index finger and thumb, and all measurements were obtained at altitudes ranging from 610 to 4860 m during an expedition in the Qinghai Plateau in China. With the change in altitude HF, ME, and MF showed no significant change. Compared to the MdPF at 2260 m on ascent, the MdPF at other altitudes showed a significant decrease (P<0.01). Thus, we conclude that muscle performance (HF, MF, and ME) was not affected by the environment at high altitude. However, MdPF was affected and the mean MdPF at 610 m after the expedition did not recover to initial values of MdPF. We suggest these results may have been affected by fatigue and chronic exposure to the hypobaric hypoxic environment, since the members of the expedition party expressed feelings of sluggishness and fatigue after the expedition.     

Intradermal administration of lipopolysaccharide in treatment of human cancer

 Lipopolysaccharide (LPS) has been recognized as a potent antitumor agent in animal tumor models; however, its use in human cancer therapy has been limited to only one trial, in which LPS from Salmonella was given intravenously. It was not very successful because of poor tumor response and was also toxic. We originally developed LPS prepared from Pantoea agglomerans (LPSp), and this was a well-purified, small-molecular-mass (5 kDa) agent. We chose intradermal rather than intravenous administration in the hope that the former would release LPS slowly into the bloodstream, and thus be less toxic while preserving antitumor activity. In our animal tumor models, intradermal administration was indeed less toxic and more beneficial for tumor regression than intravenous administration. We made a pilot study with intradermal administration of LPSp on the treatment of ten advanced cancer patients. Five of them had evaluable tumor, which had failed earlier to respond to conventional chemotherapy. Cyclophosphamide was also administered in this trial, in anticipation of its synergistic effect with LPSp. In this study LPSp was injected intradermally into each patient twice a week, starting with an initial dose of 0.4 ng/kg, and raising it to 600 or 1800 ng/kg. A 400-mg/m² dose of cyclophosphamide was given intravenously every 2 weeks. After completion of the dose escalation, the treatment was continued for at least 4 months, and it was found that 1800 ng/kg LPSp was well tolerated. A significant level of cytokines was observed in the sera for at least 8 h. These results indicate higher tolerable doses and remarkably more continuous induction of the cytokines than were reported in a previous study by others using intravenous administration. Three of the five evaluable tumors showed a significant response to our combined therapy. Intradermally administered, LPS was less toxic and elicited a tumor response in combination with cyclophosphamide; it can thus can be applied to cancer treatment even in humans. Received: 3 August 1995 / Accepted: 2 April 1996     

Complete1H NMR assignments of synthetic glycopeptides from the carbohydrate-protein linkage region of serglycins

We present complete¹H NMR assignments for two synthetic glycopeptides representative of the carbohydrateprotein linkage region of serglycin proteoglycans. The peptides are: Ser(Galp-Xylp)-Gly-Ser-Gly-Ser(Galp-Xylp)-Gly and, Ser(Galp-Xylp)-Gly-Ser(Galp-Xylp)-Gly-Ser(Galp-Xylp)-Gly. A number of 2D NMR spectra together with a 3D NOESY-TOCSY spectrum were acquired at 600 MHz to complete the assignments of the glycopeptides dissolved in water with 40% trifluoroethanol. Preliminary analysis of the NMR data suggests folded structures for the glycopeptides.A preliminary account of this work was presented at an International Symposium held at the University of Alabama at Birmingham in November, 1994 on the occasion of the 65th birthday of Professor Lennart Rodén.     

Expression of NADH-Dependent Glutamate Synthase in Response to the Supply of Nitrogen in Rice Cells in Suspension Culture

The effects of inorganic and organic nitrogen on the levelsof mRNA for NADH-dependent glutamate synthase (GOGAT) and theprotein were examined in rice cells in suspension culture. Asupply of NH⁺₄, NO^-₃, glutamine, or asparagine induced the accumulationof the protein and mRNA, but levels of mRNA for ferredoxin-GOGATwere hardly affected. ¹Present address: P.C. Center Wakuya-cho, Toda-gun, Miyagi,Japan.     

Bilateral giant myelolipoma in the adrenal of a cotton-top tamarin (Saguinus oedipus)

Abstract: A rare case of bilateral adrenal myelolipomas in a female cotton-top tamarin is reported. Large bilateral masses in the adrenal glands were composed of mature adipose cells containing varying amounts of hematopoietic cells of the myeloid, erythroid, and megakaryocyte series. The gross and histologic features of this case closely resemble human “giant” adrenal myelolipomas.     

Bioconversion of 7-Aminocephalosporanic Acid by Intact Rhodotorula glutinis Cells

When 7-aminocephalosporanic acid (7-ACA) was used as a single carbon source in the enrichment culture medium for screening 7-ACA-degrading microorganisms, pink yeast colonies appeared frequently, and these were identified as Rhodotorula glutinis. These intact R. glutinis cells converted (i) 7-ACA to deacetyl-7-ACA (7-ADACA) and (ii) monochloroacetyl-7-ACA to monochloroacetyl-7-ADACA at sufficiently high levels to be of commercial interest. Acetylation of 7-ADACA to 7-ACA, the reverse reaction of hydrolysis in an organic medium with methyl acetate as an acetyl donor, was also demonstrated.     

Biochemical systematics of Japanese fireflies of the subfamily Luciolinae and their flash communication systems

Japanese fireflies of the subfamily Luciolinae are biochemically analyzed using 13 allozymes, and the phylogenetic relationships obtained from this analysis are compared with their flash communication systems. As a result, the Japanese Luciolinae can be divided into three groups.Hotaria parvula andH. tsushimana together withLuciola yayeyamana andL. kuroiwae from the first group, and they use the same communication system.L. lateralis, Curtos okinawana, andC. costipennis make up the second group, and their communication systems are also the same.L. cruciata makes up the last one, and its communication system is different from the other fireflies of Luciolinae. Therefore, their taxonomical arrangement and communication systems are not congruent. However, the genetic similarity deduced by allozymic analysis of the members of the Japanese Luciolinae is highly consistent with their flash communication systems.