Genetic background modifies inner ear and eye phenotypes of jag1 heterozygous mice

Mice heterozygous for missense mutations of the Notch ligand Jagged1 (Jag1) exhibit head-shaking behavior indicative of an inner ear vestibular defect. In contrast, mice heterozygous for a targeted deletion of the Jag1 gene (Jag1del1) do not demonstrate obvious head-shaking behavior. To determine whether the differences in inner ear phenotypes were due to the types of Jag1 mutations or to differences in genetic background, we crossed Jag1del1 heterozygous mice onto the same genetic background as the missense mutants. This analysis revealed that variation of the Jag1 mutant inner ear phenotype is caused by genetic background differences and is not due to the type of Jag1 mutation. Genome scans of N2 backcross mice identified a significant modifier locus on chromosome 7, as well as a suggestive locus on chromosome 14. We also analyzed modifiers of an eye defect in Jag1del1 heterozygous mice from this same cross.     

Peroxisomal metabolism of propionic acid and isobutyric acid in plants

The subcellular sites of branched-chain amino acid metabolism in plants have been controversial, particularly with respect to valine catabolism. Potential enzymes for some steps in the valine catabolic pathway are clearly present in both mitochondria and peroxisomes, but the metabolic functions of these isoforms are not clear. The present study examined the possible function of these enzymes in metabolism of isobutyryl-CoA and propionyl-CoA, intermediates in the metabolism of valine and of odd-chain and branched-chain fatty acids. Using (13)C NMR, accumulation of beta-hydroxypropionate from [2-(13)C]propionate was observed in seedlings of Arabidopsis thaliana and a range of other plants, including both monocots and dicots. Examination of coding sequences and subcellular targeting elements indicated that the completed genome of A. thaliana likely codes for all the enzymes necessary to convert valine to propionyl-CoA in mitochondria. However, Arabidopsis mitochondria may lack some of the key enzymes for metabolism of propionyl-CoA. Known peroxisomal enzymes may convert propionyl-CoA to beta-hydroxypropionate by a modified beta-oxidation pathway. The chy1-3 mutation, creating a defect in a peroxisomal hydroxyacyl-CoA hydrolase, abolished the accumulation of beta-hydroxyisobutyrate from exogenous isobutyrate, but not the accumulation of beta-hydroxypropionate from exogenous propionate. The chy1-3 mutant also displayed a dramatically increased sensitivity to the toxic effects of excess propionate and isobutyrate but not of valine. (13)C NMR analysis of Arabidopsis seedlings exposed to [U-(13)C]valine did not show an accumulation of beta-hydroxypropionate. No evidence was observed for a modified beta-oxidation of valine. (13)C NMR analysis showed that valine was converted to leucine through the production of alpha-ketoisovalerate and isopropylmalate. These data suggest that peroxisomal enzymes for a modified beta-oxidation of isobutyryl-CoA and propionyl-CoA could function for metabolism of substrates other than valine.     

Ecological role of <Emphasis Type="Italic">Phyllophora antarctica</Emphasis> drift accumulations in coastal soft-sediment communities of McMurdo Sound,Antarctica

At Cape Evans on Ross Island, Antarctica, the rhodophyte Phyllophora antarctica is the dominant primary producer in terms of biomass from 10 to >30 m depth. The vast majority of Phyllophora occurs as accumulations of unattached plants. Whilst decomposition and incorporation of macroalgal drift material into the food web is rapid in temperate ecosystems, we predicted these processes to be slow in Antarctica. We address the functional role of macroalgal detritus in fuelling the biodiversity of benthic communities at Cape Evans during the summers of 2001 and 2002. Specifically we (a) describe the distribution and biomass of attached and drift algae, (b) assess the photosynthetic capacity and degradation of drift accumulations using in situ fluorometry, (c) assess the effect of patches of drift Phyllophora on underlying macrofaunal communities, and, (d) use stable isotopes to investigate the possible uptake of Phyllophora by macrofauna. We found drift Phyllophora accumulations throughout the depth range investigated (3–31 m), with peak biomasses of 140±30 g dwt m^–2 in the 15–25 m depth strata. At this depth stratum Phyllophora was a conspicuous habitat element with the % cover on the seafloor averaging 30%. While initially the drift algal accumulations appeared in good health we measured significant declines in photosynthetic capacity between years suggesting ongoing, albeit slow, degradation of the drift algal accumulations. Our results demonstrate that Phyllophora drift accumulations have a structuring role on soft-sediment communities, which increases in strength with the gradual degradation of the algae. The longevity of Phyllophora is enhanced by secondary metabolites, which serve as protection against grazers, and their extreme shade adaptation. However, our carbon and nitrogen stable isotope data of polychaetes and amphipods associated with Phyllophora suggest that macroalgal detritus enters the food web, and although this process is slow, Phyllophora accumulations might serve to dampen the seasonality in food supply providing higher trophic levels with a more constant food source.     

Endoplasmic reticulum export sites and Golgi bodies behave as single mobile secretory units in plant cells

In contrast with animals, plant cells contain multiple mobile Golgi stacks distributed over the entire cytoplasm. However, the distribution and dynamics of protein export sites on the plant endoplasmic reticulum (ER) surface have yet to be characterized. A widely accepted model for ER-to-Golgi transport is based on the sequential action of COPII and COPI coat complexes. The COPII complex assembles by the ordered recruitment of cytosolic components on the ER membrane. Here, we have visualized two early components of the COPII machinery, the small GTPase Sar1p and its GTP exchanging factor Sec12p in live tobacco (Nicotiana tabacum) leaf epidermal cells. By in vivo confocal laser scanning microscopy and fluorescence recovery after photobleaching experiments, we show that Sar1p cycles on mobile punctate structures that track with the Golgi bodies in close proximity but contain regions that are physically separated from the Golgi bodies. By contrast, Sec12p is uniformly distributed along the ER network and does not accumulate in these structures, consistent with the fact that Sec12p does not become part of a COPII vesicle. We propose that punctate accumulation of Sar1p represents ER export sites (ERES). The sites may represent a combination of Sar1p-coated ER membranes, nascent COPII membranes, and COPII vectors in transit, which have yet to lose their coats. ERES can be induced by overproducing Golgi membrane proteins but not soluble bulk-flow cargos. Few punctate Sar1p loci were observed that are independent of Golgi bodies, and these may be nascent ERES. The vast majority of ERES form secretory units that move along the surface of the ER together with the Golgi bodies, but movement does not influence the rate of cargo transport between these two organelles. Moreover, we could demonstrate using the drug brefeldin A that formation of ERES is strictly dependent on a functional retrograde transport route from the Golgi apparatus.     

N2-Fixation in Cyanobacterial Mats from Ponds on the McMurdo Ice Shelf,Antarctica

We have investigated the ecological importance of N2-fixation in cyanobacterial mats, dominated by oscillatorean species, in ponds of the Bratina Island area of the McMurdo Ice Shelf, Antarctica (78°S, 166°E). Nitrogenase activity, estimated as acetylene reducing activity (ARA), was found in all the mats investigated (n = 16). The average ARA was 75.9 mmol ethylene m-2 h-1, ranging from 6 to 201 mmol ethylene m-2 h-1. Nitrogenase activity was positively correlated with dissolved reactive phosphorus concentration in pondwater and the C/N ratio of the mat, and was negatively correlated with pondwater NH4+-N concentrations and natural abundance of 15N in the mats. ARA was restricted to the upper, oxic layer of the mats. Experiments conducted to ascribe ARA to different groups of prokaryotes suggested that ARA was mainly conducted by heterocystous cyanobacteria, since no activity was found in the dark and the activity was inhibited by the photosystem II inhibitor DCMU (3-[3,4-dichlorophenyl]-1,1-dimethyl urea). In spite of 24 h of daylight, nitrogenase activity showed a diel cycle with maximum activity at midday (10-18 h) and minimal activity at early morning (6-10 h) when pond temperatures were at their minima. Light dependency of nitrogenase activity for three cyanobacterial communities showed that the irradiance required for saturating ARA was low, in every case lower than 100 mmol photon m-2s-1. Irradiance rarely fell below 100 mmol photon m-2s-1 during Antarctic summer days and ARA was likely to be light saturated for much of the time. We estimate that N2 fixation represented on average a N input into the ponds of over 1 g m-2y-1. This value appears to be the highest N input to this Antarctic ecosystem.     

The Mouse GalR2 Galanin Receptor: Genomic Organization, cDNA Cloning, and Functional Characterization

Abstract: The diverse physiological actions of galanin are thought to be mediated through activation of galanin receptors (GalRs). We report the genomic and cDNA cloning of a mouse GalR that possesses a genomic structure distinct from that of GalR1 and encodes a functional galanin receptor. The mouse GalR gene consists of two exons separated by a single intron within the protein-coding region. The splicing site for the intron is located at the junction between the third transmembrane domain and the second intracellular loop. The cDNA encodes a 370-amino acid putative G protein-coupled receptor that is markedly different from human GalR1 and rat GalR3 (38 and 57%) but shares high homology with rat GalR2 (94%). In binding studies utilizing membranes from COS-7 cells transfected with mouse GalR2 cDNA, the receptor displayed high affinity ( K _D = 0.47 n M ) and saturable binding with ¹²⁵I-galanin ( B _max = 670 fmol/mg). The radioligand binding can be displaced by galanin and its analogues in a rank order: galanin ⋍ M40 ⋍ M15 ⋍ M35 ⋍ C7 ⋍ galanin (2–29) ⋍ galanin (1–16) ≫ galanin (10–29) ⋍ galanin (3–29), which resembles the pharmacological profile of the rat GalR2. Receptor activation by galanin in COS-7 cells stimulated phosphoinositide metabolism, which was not reversed by pertussis toxin. Thus, the galanin receptor encoded in the cloned mouse GalR gene is the type 2 galanin receptor and is active in both ligand binding and signaling assays.