Degradation of aromatic amino acids by Rhodococcus erythropolis

A Gram-positive Rhodococcus erythropolis strain S1 was shown to assimilate aromatic amino acids such as L-phenylalanine, L-tyrosine, L-tryptophan, D-phenylalanine, D-tyrosine and D-tryptophan, which were utilized not only as the sole carbon source but also as a suitable nitrogen source. The highest growth on these aromatic amino acids occurred at a temperature of 30°C. L-Phenylalanine, L-tyrosine and L-tryptophan degradative pathways would appear to be independent, and to be induced alternatively. The strain S1 also showed the ability to assimilate peptides which consisted of only L-phenylalanine and L-tyrosine.     

Macrophage catabolism of lipid A is regulated by endotoxin stimulation

Lipopolysaccharide (LPS) is a Gram-negative bacterial glycolipid that is believed to cause, by virtue of its stimulatory actions on macrophages and other eukaryotic cells, the life-threatening symptoms associated with Gram-negative infections. Macrophages both respond to and catabolically deactivate LPS. The lipid A moiety of LPS is responsible for the stimulatory actions of LPS on macrophages. We have previously developed methods employing a radiolabeled bioactive lipid A precursor, 4'-32P-lipid IVA, to study the interaction of this class of lipids with animal cells (Hampton, R. Y., Golenbock, D. T., and Raetz, C. R. H. (1988). J. Biol. Chem. 263, 14802-14807). In the current work, we have examined the uptake and catabolism of 4'-32P-lipid IVA by the RAW 264.7 cell line in serum-containing medium at physiological temperatures and have studied the effect of LPS stimulation on the ability of these cells to catabolize lipid IVA. RAW 264.7 macrophage-like cells avidly take up 4'-32P-lipid IVA under cell culture conditions at nanomolar concentrations. Uptake of lipid IVA was accompanied by lysosomal dephosphorylation of a fraction of the lipid to yield 4'-monophosphoryl lipid IVA. Chemically generated 4'-monophosphoryl lipid IVA was found to be substantially less bioactive than lipid IVA in the RAW cell, indicating that this catabolic dephosphorylation results in detoxification. In uptake experiments of 3-4 h duration, all metabolism of lipid IVA is blocked by ligands of the macrophage scavenger receptor. In longer experiments (24 h), both scavenger receptor-dependent and -independent uptake are responsible for the lysosomal catabolism of lipid IVA. Preincubation of RAW 264.7 cells with LPS caused dose-dependent inhibition of lipid IVA dephosphorylation. Sufficient LPS stimulation resulted in essentially complete inhibition of lipid IVA catabolism in both short- and long-term uptake experiments. This effect occurred at physiologically relevant concentrations of LPS (IC50 less than 1 ng/ml), and our data indicate that LPS-induced blockade of lipid IVA catabolism was due to the resultant physiological stimulation of the cells, and not inhibition of dephosphorylation by competition for uptake or enzymatic sites or by simple sequestration of labeled lipid IVA by LPS aggregates. We suggest that in the macrophage, LPS can modulate its own catabolism by virtue of its pharmacological properties. This effect of LPS could play a role in LPS pathophysiology as well as in macrophage biology.     

Chromosome studies in the red howler monkey, Alouatta seniculus stramineus (Platyrrhini, Primates): description of an X1X2Y1Y2/X1X1X2X2 sex-chromosome system and karyological comparisons with other subspecies

In the red howler monkey, Alouatta seniculus stramineus (2n = 47, 48, or 49), variations in diploid chromosome number are due to different numbers of microchromosomes. Males exhibit a Y;autosome translocation involving the short arm of an individual biarmed autosome. Consequently, the sex-chromosome constitution in the male is X1X2Y1Y2, with X1 representing the original X chromosome, X2 the biarmed autosome (No. 7), Y1 the Y;7p translocation product, and Y2 the acrocentric homolog of 7q. In the first meiotic division, a quadrivalent with a chain configuration can be observed in spermatocytes. Females have an X1X1X2X2 sex-chromosome constitution. Chromosome heteromorphisms were observed in pair 13, due to a pericentric inversion, and pair 19, due to the presence of constitutive heterochromatin. Microchromosomes, which varied in number between individuals, were also heterochromatic. NOR-staining was observed at two separate sites on a single chromosome pair (No. 10). A comparison of A.s. stramineus with A.s. macconnelli shows that these two subspecies have identical diploid chromosome numbers (47, 48, or 49), again due to a varying number of microchromosomes, and that they share a similar sex-chromosome constitution. Their karyotypes, however, are not identical, but can be derived from each other by a reciprocal translocation. Further comparisons with other A. seniculus subspecies reported in the literature indicate that this taxon is not karyologically uniform and that substantial chromosome shuffling has occurred between populations that have been considered to be subspecies by taxonomic criteria based on their morphometric attributes.     

The effects of thiamine inhibition on ruminal fermentation: a preliminary study

Inhibition of methanogenesis in ruminal cultures was attempted by hindering thiamine availability through its degradation by ‘polyphenols’ and competition for active sites on enzymes and transporters using thiamine structural analogs. Effects on fermentation were small and not consistently reversed by adding thiamine. Lack of major effects of the compounds evaluated could be due to intracellular synthesis of thiamine covering most requirements.     

Release of arachidonic acid from human endometrial cells in culture mediated by calcium ionophore (A23187) or fluoride.

Primary cultures of endometrial glands and stromal cells were labelled with [14C]-arachidonic acid for 4 h before exposure to either the calcium ionophore, A23187 (which activates phospholipase A2 (PLA2) by increasing intracellular calcium concentrations) or sodium fluoride (which activates a G-protein). Calcium ionophore (0.5-50 mumol/l) stimulated a dose- and time-dependent release of arachidonic acid from endometrial glands. Incubation with ionophore (10 mumol/l) for 1 h released 22% of the incorporated arachidonic acid. There was a corresponding decrease in phospholipids and no loss from triglycerides. Stromal cells were unresponsive to ionophore. Fluoride (10 mmol/l) stimulated a release of arachidonic acid from stromal cells and endometrial glands (6.5% of the total arachidonic acid incorporated). In stromal cells, arachidonic acid was released from triglycerides in Day-1 cultures and from phospholipids in Day-2 cultures. In both Day-1 and Day-2 cultures of endometrial glands, arachidonic acid was released from phospholipids, but not from triglycerides. Among the phospholipids, phosphatidylcholine was always the major source of arachidonic acid. Arachidonic acid release from endometrial glands and stromal cells may be mediated by activation of PLA2 (or phospholipase C) via a G-protein, but in glands calcium ionophore may have a direct effect on PLA2. The response to calcium ionophore may reflect the differences in calcium requirements of the two endometrial PLA2 isoenzymes.