Naphthalene oxygenase fromCorynebacterium renale: Characterisation and mechanism of oxygenation

The formation ofcis-l,2,-dihydroxy-l,2,-dihydronaphthalene from naphthalene by naphthalene oxygenase, purified fromCorynebacterium renale ATCC 15075, was demonstrated to involve oxidation of a mol NADH and consumption of one mol oxygen. The enzyme contains one g-atom Fe²⁺ and one FAD. Catalase inhibited product formation and H2O2 could substitute for NADH in the reaction. Superoxide dismutase inhibited enzyme activity when either NADH or H₂O₂ was present; the generation of superoxide anion on addition of NADH to the enzyme, in the absence of naphthalene, was detected by the nitro blue tetrazolium reduction method. Hydroxyl radical scavengers, ethanol, mannitol and sodium benzoate, inhibited product formation when either NADH or H₂O₂ was present. Electron spin resonance studies, under aerobic conditions, indicated that iron of the enzyme underwent valence changes during the course of the reaction     

Inactivation by chloroquine of alpha-galactosidase in cultured human skin fibroblasts

When human skin fibroblasts are cultured in the presence of chloroquine or NH₄Cl there is a decrease in the intracellular level of lysosomal hydrolases and a concomitant increase in the extracellular activity as compared with cells grown in the absence of a base (cf [18]). In a medium with 25 μM chloroquine or 5 mM NH₄Cl, the decrease in the intracellular activity of β-hexosaminidase, arylsulphatase and β-glucuronidase is 10–40% after 1 day. A similar decrease in α-galactosidase activity is observed in cells grown in the presence of 5 mM NH₄Cl. However, in the presence of 25 μM chloroquine, the intracellular activity of α-galactosidase decreases by 80–90% within 6 h. The inactivation is irreversible. After removal of the chloroquine and further culture of the cells in chloroquine-free medium, α-galactosidase activity gradually increases due to de novo synthesis. The turnover time of α-galactosidase was calculated to be 1.9 days. Inactivation of α-galactosidase also occurs when homogenates are incubated with chloroquine, but the concentration of the base required for maximum inactivation is at least three orders of magnitude higher than that which must be present in the medium of intact cells to obtain the same effect.     

Human α-globin maps to pter-p13.3 in chromosome 16 distal to PGP

Summary Fibroblasts from a fetus with an unbalanced karyotype 46(XY),-16,+(16qter-16p13.3::4q31.1-4qter) were found to possess only one allele at the 3 hypervariable region (3HVR) close to the -globin locus and two alleles at the PGP locus. This places the -globin locus at the very tip of 16p, distal to PGP.     

A 4-base-pair deletion polymorphism in man

Partial nucleotide sequencing of C11p11, a probe mapping close to the gene determining familial adenomatous polyposis (FAP) on human chromosome 5, in 4 unrelated persons has revealed a 4-base-pair deletion variant designated DELI at an arbitrary DNA locus D5S71. For screening the deletion variants that may frequently occur in the non-coding DNA sequences, we set up a non-invasive procedure which involves DNA amplification by PCR, simple polyacrylamide gel electrophoresis and direct visualisation of alleles under long wave ultraviolet light by ethidium bromide staining.     

Assignment of human transcobalamin II (TC2) to chromosome 22 using somatic cell hybrids and monosomic meningioma cells

Summary Human transcobalamin II (TC2), a vitamin B12 binding serum protein, is synthesized and secreted into the medium by cells growing in vitro. Mouse-man somatic cell hybrids were analyzed in order to map the locus of TC2. The presence of human TC2 in the culture media was correlated with the results of genetic marker and chromosome analysis of the hybrid cells. Chromosome 22 showed 100% concordancy. However, chromosome 6 (90% concordancy) and chromosome 7 (96% concordancy) were not completely excluded. Meningioma cells obtained from patients heterozygous for TC2 showed a concomitant loss of one chromosome 22 and one of the TC2 alleles, strongly supporting the assignment to chromosome 22.     

Goblet Cell Derived RELM-β Recruits CD4+ T Cells during Infectious Colitis to Promote Protective Intestinal Epithelial Cell Proliferation

Enterohemorrhagic Escherichia coli and related food and waterborne pathogens pose significant threats to human health. These attaching/effacing microbes infect the apical surface of intestinal epithelial cells (IEC), causing severe diarrheal disease. Colonizing the intestinal luminal surface helps segregate these microbes from most host inflammatory responses. Based on studies using Citrobacter rodentium, a related mouse pathogen, we speculate that hosts rely on immune-mediated changes in IEC, including goblet cells to defend against these pathogens. These changes include a CD4⁺ T cell-dependent increase in IEC proliferation to replace infected IEC, as well as altered production of the goblet cell-derived mucin Muc2. Another goblet cell mediator, REsistin-Like Molecule (RELM)-β is strongly induced within goblet cells during C. rodentium infection, and was detected in the stool as well as serum. Despite its dramatic induction, RELM-β’s role in host defense is unclear. Thus, wildtype and RELM-β gene deficient mice (Retnlb ^-/-) were orally infected with C. rodentium. While their C. rodentium burdens were only modestly elevated, infected Retnlb ^-/- mice suffered increased mortality and mucosal ulceration due to deep pathogen penetration of colonic crypts. Immunostaining for Ki67 and BrDU revealed Retnlb ^-/- mice were significantly impaired in infection-induced IEC hyper-proliferation. Interestingly, exposure to RELM-β did not directly increase IEC proliferation, rather RELM-β acted as a CD4⁺ T cell chemoattractant. Correspondingly, Retnlb ^-/- mice showed impaired CD4⁺ T cell recruitment to their infected colons, along with reduced production of interleukin (IL)-22, a multifunctional cytokine that directly increased IEC proliferation. Enema delivery of RELM-β to Retnlb ^-/- mice restored CD4⁺ T cell recruitment, concurrently increasing IL-22 levels and IEC proliferation, while reducing mucosal pathology. These findings demonstrate that RELM-β and goblet cells play an unexpected, yet critical role in recruiting CD4⁺ T cells to the colon to protect against an enteric pathogen, in part via the induction of increased IEC proliferation.     

Diversity and Distribution of Hantaviruses in South America

Hantaviruses are important contributors to disease burden in the New World, yet many aspects of their distribution and dynamics remain uncharacterized. To examine the patterns and processes that influence the diversity and geographic distribution of hantaviruses in South America, we performed genetic and phylogeographic analyses of all available South American hantavirus sequences. We sequenced multiple novel and previously described viruses (Anajatuba, Laguna Negra-like, two genotypes of Castelo dos Sonhos, and two genotypes of Rio Mamore) from Brazilian Oligoryzomys rodents and hantavirus pulmonary syndrome cases and identified a previously uncharacterized species of Oligoryzomys associated with a new genotype of Rio Mamore virus. Our analysis indicates that the majority of South American hantaviruses fall into three phylogenetic clades, corresponding to Andes and Andes-like viruses, Laguna Negra and Laguna Negra-like viruses, and Rio Mamore and Rio Mamore-like viruses. In addition, the dynamics and distribution of these viruses appear to be shaped by both the geographic proximity and phylogenetic relatedness of their rodent hosts. The current system of nomenclature used in the hantavirus community is a significant impediment to understanding the ecology and evolutionary history of hantaviruses; here, we suggest strict adherence to a modified taxonomic system, with species and strain designations resembling the numerical system of the enterovirus genus.