Regional assignment of the human uroporphyrinogen III synthase (UROS) gene to chromosome 10q25.2→q26.3

Summary Uroporphyrinogen III synthase [UROS; hydroxymethylbilane hydro-lyase (cyclizing), EC 4.2.1.75] is the fourth enzyme in the human heme biosynthetic pathway. The recent isolation of the cDNA encoding human UROS facilitated its chromosomal localization. Human UROS sequences were specifically amplified by the polymerase chain reaction (PCR) from genomic DNA of two independent panels of human-rodent somatic cell hybrids. There was 100% concordance for the presence of the human UROS PCR product and human chromosome 10. For each of the other chromosomes, there was 19%–53% discordance with human UROS. The chromosomal assignment was confirmed by Southern hybridization analysis of DNA from somatic cell hybrids with the full-length UROS cDNA. Using human-rodent hybrids containing different portions of human chromosome 10, we assigned the UROS gene to the region 10q25.2 q26.3.     

Lithium-induced decrease of brain inositol and increase of brain inositol-1-phosphate is transient

The effects of a single does of LiCl (2.5 or 10 mEq/kg) on brain inositol and inositol-1-phosphate (Ins1P), intermediates of brain phosphoinositude (PI) turnover, were determinated in male Han: Wistar rats. There was a remarkable, 36–58 fold elevation of brain Li⁺ as the single does of LiCl was increased 4-fold. Moreover, the accumulation of brain lithium was slow during repeated administration of LiCl. Brain lithium did not correlate with changes in brain PI turnover either after a single or repeated doses. Thus, after a single does of LiCl the increases in brain Ins1P were much less than the decreases in brain inositol. Also, brain inositol was significantly decreased only with the high dose of LiCl whereas brain Ins1P accumulation was more prominent with the lower dose. Moreover, repeated daily doses of LiCl only transiently increased brain Ins1P at 1 and 7 d whereas inositol remained at control levels throughout the 14 d observation period. Lithium probably caused the transient decrease in brain inositol by inhibiting several enzymes, in addition to the inhibition of myo-inositol mono-phosphates, in the PI cycle. Moreover, a slow dampening down of PI turnover by lithium, possible via an inhibitory action on G-protein-coupling, may also explain the present findings.     

Phospholipids chiral at phosphorus: Fourier-transform infrared study on the gel-liquid crystalline transition of chiral thiophosphatidylcholine

Fourier-transform infrared spectroscopy (FT-IR) was used to study the structural properties of Rp, Sp, and Rp + Sp isomers of 1,2-dipalmitoyl-sn-glycero-3-thiophosphocholine (DPPsC), in comparison with those of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC). For the vibrational modes of acyl chains, isomers of DPPsC show similar temperature and phase dependence to DPPC. However, the Rp isomer of DPPsC exhibits several unique properties: the CH2 symmetric stretching band is unusually weak, the CH2 asymmetric stretching band is unusually narrow, and the CH2 wagging bands do not disappear completely at temperatures above the main transition. These differences could imply a tighter packing and be responsible for the unique phase-transition property of (Rp)-DPPsC. For the vibrational modes of the thiophosphodiester group, the frequency of the P-O stretching mode of DPPsC suggests that the POS- triad exists predominantly in the mesomeric form. This is in contrast to the structure of nucleoside phosphorothioates where charge localization at sulfur has been demonstrated [Iyengar, R., Eckstein, F., & Frey, P. A. (1984) J. Am. Chem. Soc. 106, 8309-8310]. This suggests that the different biophysical properties between isomers of DPPsC are not due to different charge distribution in the POS- triad or different geometry of charge distribution on the membrane surface. Instead, factors such as size or hydration property of oxygen and sulfur, as well as the different configuration at phosphorus, could be responsible for the differences in the conformation and packing of acyl chains, as revealed by the different properties in the CH2 stretching and wagging modes of DPPsC.     

Envelope proteins of human T cell leukemia virus type I: characterization by antisera to synthetic peptides and identification of a natural epitope

Three peptides corresponding to selected regions of the env gene products of human T cell leukemia virus type I were synthesized by solid-phase Merrifield techniques. The sequence of peptide designated SP-65 was identical to the predicted C-terminal 12 residues of the transmembrane protein p21env, and peptide SP-74 was inferred from a region shown to be highly conserved among mammalian retroviruses. The third peptide, SP-70, was derived from a C-terminal region of the surface glycoprotein gp46. Antibodies to each peptide were raised in rabbits and were used to identify and further characterize the proteins coded by the env gene. Despite being present at very low levels in purified viral preparations, these proteins were chromatographed by reverse-phase high pressure liquid chromatography and were located by Western blot analysis of the column fractions. Anti-SP-70 recognized the surface glycoprotein (gp46) and also its C-terminal cleavage fragment (gp16). Anti-SP-65 and anti-SP-74 both reacted with the hydrophobic transmembrane protein (p21) and provided evidence that this protein does not undergo apparent C-terminal processing during viral maturation, unlike the trans-membrane protein of murine leukemia virus. As expected, anti-SP-74 also reacted with homologous proteins from other Type C and Type D viruses, confirming that peptide SP-74 corresponds to a broadly conserved region of retroviral transmembrane proteins. SP-70, which is predicted to be quite near the C terminus of the major surface glycoprotein, was also reactive with sera of HTLV-I-positive patients, indicating that this peptide corresponds to, or is part of, a native epitope recognized by the natural host.     

Acute toxoplasma infection of mice induces spleen NK cells that are cytotoxic for T. gondii in vitro

Murine spleen natural killer (NK) cells from normal and Toxoplasma-infected BALB/c mice were examined for their reactivity against RH strain tachyzoites in vitro. First, the effect of suspending medium on survival of extracellular RH tachyzoites was determined. Optimal parasite viability (by ethidium bromide-acridine orange staining) was observed when tachyzoites were incubated in phosphate-buffered saline (PBS) containing 10% horse serum (HS) for as long as 5 hr. In addition, parasite viability in PBS-HS correlated with subsequent infectivity, because freshly harvested and PBS-HS-incubated tachyzoites were equivalent in their ability to cause lethal infections in normal mice and to survive within normal mouse macrophages. Furthermore, viability and tumoricidal capacity of murine spleen NK cells incubated in PBS-HS was comparable to that of NK cells incubated in a standard cytotoxicity medium. Incubation of effector NK cells and target tachyzoites in PBS-HS in vitro revealed that spleen NK cells from 3-day Toxoplasma-infected mice had significantly greater cytotoxicity for extracellular RH tachyzoites than did control cells from uninfected mice. Moreover, Toxoplasma gondii-induced spleen NK cell toxoplasmacidal activity was significant at all effector to target cell ratios tested, and appeared to be mediated by direct contact between the host cell and the parasite. These in vitro results suggest that NK cells may be important in host defense against T. gondii.     

Retroperitoneal fibromatosis and acquired immunodeficiency syndrome in macaques: clinical and immunologic studies

A simian acquired immunodeficiency syndrome (SAIDS) associated with retroperitoneal fibromatosis (RF) has been observed in several species of macaque at the Washington Regional Primate Research Center. Clinical signs were recurrent diarrhea, weight loss, mesenteric lymphadenopathy, and opportunistic infections. Most affected macaques in the later stages of illness showed marked immunodeficiency. Response of peripheral blood mononuclear cells to mitogens was impaired significantly. There was sharply depressed primary and secondary antibody response to the T-cell dependent antigen, bacteriophage phi X174. Affected monkeys did not switch from IgM to IgG antibody following a secondary immunization, as did normal macaques. Twenty-four (67%) of 36 affected animals with progressive RF or deteriorated stages of illness had hypoproteinemia and hypoalbuminemia. Quantitative serum immunoglobulins of 23 cases showed that eight (35%) had hypogammaglobulinemia, six (26%) had hypergammaglobulinemia, and the remainder (39%) were within the normal range. Opportunistic infections were predominantly bacterial pathogens. Type D retrovirus appeared to be closely associated with RF-affected macaques (12/12 or 100%). The case fatality rate (including animals sacrificed after prolonged illness) was 98%. The leading cause of death was due directly to RF lesions in 43%, to enterocolitis in 36%, septicemia in 12%, amyloidosis in 5%, and malignant lymphoma (2%). Clinical, immunologic and pathologic changes reveal an acquired immunodeficiency syndrome that has many similarities to human AIDS. SAIDS and RF may be a useful model for studying human AIDS.     

Transformation of Tobacco, Tomato, Potato, and Arabidopsis thaliana Using a Binary Ti Vector System

Using a binary tumor-inducing (Ti) plasmid vector system, several plant species were transformed with a kanamycin resistance marker (neomycin phosphotransferase gene). Four Nicotiana species, seven tomato cultivars, two potato cultivars, and Arabidopsis thaliana were transformed by the binary vector transformation method. In this method, various plant organ pieces were co-cultivated with Agrobacterium tumefaciens cells carrying the binary vector, pGA472, and a helper Ti plasmid. We have also demonstrated that a wild type Ti plasmid can be used as a helper to obtain a transformed plant.     

Expression and secretion of biologically active human atrial natriuretic peptide in Saccharomyces cerevisiae

A hybrid gene was constructed containing a fusion between the DNA sequences encoding the secretory precursor of the yeast mating pheromone alpha-factor and a synthetic sequence encoding a biologically active 24-amino acid carboxyl-terminal portion of the human atrial natriuretic peptide (hANP) precursor. Transformation of Saccharomyces cerevisiae with the hybrid gene resulted in the yeast cells secreting biologically active hANP into the extracellular medium. The secreted hANP was purified and found to be accurately processed at the junction in the chimeric alpha-factor/hANP protein, producing the desired mature hANP amino terminus. The secreted product was also folded correctly with respect to the single disulfide bond. However, the carboxyl terminus of the secreted hANP material was heterogeneous such that the major form lacked the last two amino acids of the peptide while the minor form was the full length material. The observed processing at the carboxyl terminus of the secreted hANP may reflect a normal processing event involved in alpha-factor peptide maturation.