Alteration of seed fatty acid composition by an ethyl methanesulfonate-induced mutation in Arabidopsis thaliana affecting diacylglycerol acyltransferase activity.

In characterizing the enzymes involved in the formation of very long-chain fatty acids (VLCFAs) in the Brassicaceae, we have generated a series of mutants of Arabidopsis thaliana that have reduced VLCFA content. Here we report the characterization of a seed lipid mutant, AS11, which, in comparison to wild type (WT), has reduced levels of 20:1 and 18:1 and accumulates 18:3 as the major fatty acid in triacylglycerols. Proportions of 18:2 remain similar to WT. Genetic analyses indicate that the fatty acid phenotype is caused by a semidominant mutation in a single nuclear gene, designated TAG1, located on chromosome 2. Biochemical analyses have shown that the AS11 phenotype is not due to a deficiency in the capacity to elongate 18:1 or to an increase in the relative delta 15 or delta 12 desaturase activities. Indeed, the ratio of desaturase/elongase activities measured in vitro is virtually identical in developing WT and AS11 seed homogenates. Rather, the fatty acid phenotype of AS11 is the result of reduced diacylglycerol acyltransferase activity throughout development, such that triacylglycerol biosynthesis is reduced. This leads to a reduction in 20:1 biosynthesis during seed development, leaving more 18:1 available for desaturation. Thus, we have demonstrated that changes to triacylglycerol biosynthesis can result in dramatic changes in fatty acid composition and, in particular, in the accumulation of VLCFAs in seed storage lipids.     

Adenosine effects upon insulin action on lipolysis and glucose transport in human adipocytes

The dose response effect of a new adenosine analogue, GR 79236 (N-[1S trans-2-hydroxycyclopentyl] adenosine) upon insulin sensitivity was examined in human adipocytes. The influence of adenosine upon insulin sensitivity for suppression of lipolysis and stimulation of glucose transport was examined. Removal of adenosine by use of adenosine deaminase stimulated lipolysis to the same extent as did 10^–9 M noradrenaline. GR79236 brought about dose dependent inhibition of lipolysis with half-maximal effect at 11.3±7.8×10^–9 M. When lipolysis was stimulated by noradrenaline alone the subsequent inhibition of lipolysis brought about by GR79236 was significantly greater than that of insulin. To examine adenosine effects on the insulin signalling pathway separately from those on lipolysis, the insulin sensitivity of glucose transport was examined. Removal of adenosine brought about a small but significant increase in the concentration of insulin required for half-maximal stimulation of glucose transport. Adenosine agonists offer promise as new agents for the modulation of metabolism in diabetes and other states of insulin resistance.     

Characterization of a region of the IncHI2 plasmid R478 which protects Escherichia coli from toxic effects specified by components of the tellurite, phage, and colicin resistance cluster.

The IncHI2 plasmid R478 specifies resistance to potassium tellurite (Te(r)), to some bacteriophages (Phi), and to pore-forming colicins (PacB). The genes encoding the three phenotypes are linked, and an 8.4-kb fragment of R478 DNA encoding them cannot be subcloned unless cocloned with a second section of the plasmid. Subclone pKFW4A contains a 5.9-kb BamHI-EcoRI fragment which caused some toxicity when present in Escherichia coli cells. Bacterial cells containing freshly transformed pKFW4A, examined by light microscopy and electron microscopy, had a filamentous morphology consistent with a block in septation. Insertion of transposon Tn1000 into terZ, -A, -B, and -C genes of pKFW4A resulted in the loss of the filamentation phenotype. Deletion of several regions of the clone confirmed that these latter components are involved in the filamentation phenotype. The region specifying protection from toxicity caused by the larger 8.4-kb fragment (encompassing this cluster and the entire 5.9-kb section of pKFW4A) was sequenced and analyzed by T7 polymerase expression and Tn1000 mutagenesis. Three open reading frames, terW, terY, and terX, were identified in a 2.6-kb region. Two polypeptides with approximate molecular masses of 18 and 28 kDa were expressed in CSRDE3 cells and were consistent with TerW (17.1 kDa; 155 amino acids [aa]) and TerY (26.9 kDa; 248 aa), whereas a protein of 213 aa deduced from terX was not observed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The terX gene product shows strong identity with the previously identified TerE, TerD, and TerZ polypeptides, and there is a conserved motif of 13 residues, GDN(R/L)TG(E/A)GDGDDE, within this group of polypeptides. Complementation analysis indicated that terW, located approximately 6.0 kb upstream of terZ, brings about protection of cells from toxic effects of components of the Te(r), Phi, and PacB cluster.     

Ultrastructural changes in Blastomyces dermatitidis after in vitro exposure to lidocaine

Electron microscopic examination of yeasts of Blastomyces dermatitidis, exposed in vitro to concentrations of lidocaine that occur when the drug is used for topical anesthesia, showed that lidocaine rapidly damaged intracellular structures. The extent of damage was dependent on the concentration of drug and length of exposure. The observed ultrastructural changes were very similar to those reported for other drugs that directly damage membranes. This relationship suggests that the antifungal effect of lidocaine is the result of direct membrane damage.     

Effect of stimulation of non-adrenergic inhibitory nerves on cyclic nucleotide levels in bullfrog lung

Carbachol in the presence of atropine and propranolol was employed to stimulate a non-adrenergic neural inhibitory system in the hemilung of the bullfrog (Rana catesbeiana). Tissue levels of cGMP were elevated 95% by carbachol whilst cAMP levels were unchanged. The phosphodiesterase inhibitor papaverine did not affect either cAMP or cGMP levels, but did selectively increase the carbachol-induced increase in lung cGMP to 220% of control levels. Papaverine did not potentiate the relaxant effects of carbachol. The results suggest that cyclic nucleotides may not be directly involved in the relaxation produced by stimulation of the non-adrenergic neural inhibitory system in this preparation.     

Evidence for a new type of endosymbiotic organization in a population of the ciliate Mesodinium rubrum from British Columbia

The ciliate Mesodinium rubrum has excited considerable interest because it has been reported to contain cryptophycean endoxymbionts which lack nuclei (Taylor et al., 1969, 1971). Hibberd (1977), however, has recently shown that in Mesodinium from the United Kingdom the cryptophycean chloroplasts and mitochondria share a common cytoplasm with an apparent cryptophycean nucleus. This cytoplasm is separated from the ciliate cytoplasm by a single membrane. Here we confirm, through extensive observation including serial sectioning, the presence of multiple, discrete, anucleate chloroplast-mitochondrial complexes in a population of Mesodinium from British Columbia. In addition to the normal ciliate macro- and micronuclei, however, these cells contain a third type of nucleus, possibly of cryptophycean origin, which is apparently in its own membrane-bound cytoplasm. This suggests the possibility of a new type of endosymbiotic organization in which endosymbiont organelles have become separated into discrete cytoplasmic compartments.     

The pineal body: Site of extraocular perception of celestial cues for orientation in the tiger salamander (Ambystoma tigrinum)

Summary Tiger salamanders (Ambystoma tigrinum) trained to orient in a particular compass direction under the sun fail to orient in the trained direction if they are (i) eyeless and simultaneously have the brain covered with opaque plastic or are (ii) eyeless and pinealectomized (Fig. 1–2, Table 1). Salamanders with either the eyes or the pineal intact and unobstructed continue to orient in the trained direction. These data strongly support the hypothesis that the pineal body is an effective extraocular photoreceptor (EOP) for compass orientation in tiger salamanders.We thank M.P. Farrell for developing computer programs for data analysis. B. Bailey and R. Walton helped conduct tests. Financial support was provided for separate phases of this research by a Biomedical Science Support Grant (NIH FR 07033-05) and NSFgrants GB-30647 and BMS 75-18693 to K. Adler and an Indiana Academy of Science Research Grant, a postdoctoral fellowship (NSF GU-2058), a Miami University Research Grant and an NSF grant (GB-41102) to D.H. Taylor.