Modest cholinergic deafferentation fails to alter hippocampal G-proteins.

The integrity of hippocampal G-protein mediated signalling following ibotenate induced lesion of the medial septum was examined. The lesion was confined histologically to the septum and induced a 23% reduction in hippocampal choline acetyltransferase (ChAT) activity and G-proteins levels and related enzyme activities were measured in the hippocampus following a 21 day survival period. The relative levels of five G-protein subunits (Gbeta, G(alpha)o, G(alpha)i1, G(alpha)i2, and G(alpha)s-L), basal GTPase, the degree of carbachol- or baclofen-stimulated GTPase activities, and the basal and fluoroaluminate-stimulated adenylate cyclase activities were apparently unaffected. To determine if our assay methodology was sensitive to changes in pre-synaptic signalling, we compared G-protein density in synaptosomes with total hippocampal homogenates. The concentration of G(alpha)q/11, G(alpha)i1, and G(alpha)i2. were significantly lower in synaptosomes, while G(alpha)o, was only marginally reduced. Thus, modest lesions of the medial-septal nucleus fail to alter G-protein signalling. However, our findings that G-protein density is lower in synaptosomal membranes than in total homogenates, indicates that the analysis of signalling events in synaptosomes following deafferentation could clarify adaptive changes which may occur at the presynaptic level.     

Acceptor specificity of the human leukocyte alpha3 fucosyltransferase: role of FucT-VII in the generation of selectin ligands

The alpha3 fucosyltransferase, FucT-VII, is one of the key glycosyltransferases involved in the biosynthesis of the sialyl Lewis X (sLex) antigen on human leukocytes. The sialyl Lewis X antigen (NeuAcalpha(2-3)Galbeta(1-4)[Fucalpha(1-3)]GlcNAc-R) is an essential component of the recruitment of leukocytes to sites of inflammation, mediating the primary interaction between circulating leukocytes and activated endothelium. In order to characterize the enzymatic properties of the leukocyte alpha3 fucosyltransferase FucT-VII, the enzyme has been expressed in Trichoplusia ni insect cells. The enzyme is capable of synthesizing both sLexand sialyl-dimeric-Lexstructures in vitro , from 3'-sialyl-lacNAc and VIM-2 structures, respectively, with only low levels of fucose transfer observed to neutral or 3'-sulfated acceptors. Studies using fucosylated NeuAcalpha(2-3)-(Galbeta(1- 4)GlcNAc)3-Me acceptors demonstrate that FucT-VII is able to synthesize both di-fucosylated and tri-fucosylated structures from mono- fucosylated precursors, but preferentially fucosylates the distal GlcNAc within a polylactosamine chain. Furthermore, the rate of fucosylation of the internal GlcNAc residues is reduced once fucose has been added to the distal GlcNAc. These results indicate that FucT-VII is capable of generating complex selectin ligands, in vitro , however the order of fucose addition to the lactosamine chain affects the rate of selectin ligand synthesis.      

The ERI-6/7 helicase acts at the first stage of an siRNA amplification pathway that targets recent gene duplications

Endogenous small interfering RNAs (siRNAs) are a class of naturally occuring regulatory RNAs found in fungi, plants, and animals. Some endogenous siRNAs are required to silence transposons or function in chromosome segregation; however, the specific roles of most endogenous siRNAs are unclear. The helicase gene eri-6/7 was identified in the nematode Caenorhabditis elegans by the enhanced response to exogenous double-stranded RNAs (dsRNAs) of the null mutant. eri-6/7 encodes a helicase homologous to small RNA factors Armitage in Drosophila, SDE3 in Arabidopsis, and Mov10 in humans. Here we show that eri-6/7 mutations cause the loss of 26-nucleotide (nt) endogenous siRNAs derived from genes and pseudogenes in oocytes and embryos, as well as deficiencies in somatic 22-nucleotide secondary siRNAs corresponding to the same loci. About 80 genes are eri-6/7 targets that generate the embryonic endogenous siRNAs that silence the corresponding mRNAs. These 80 genes share extensive nucleotide sequence homology and are poorly conserved, suggesting a role for these endogenous siRNAs in silencing of and thereby directing the fate of recently acquired, duplicated genes. Unlike most endogenous siRNAs in C. elegans, eri-6/7-dependent siRNAs require Dicer. We identify that the eri-6/7-dependent siRNAs have a passenger strand that is ～19 nt and is inset by ～3-4 nts from both ends of the 26 nt guide siRNA, suggesting non-canonical Dicer processing. Mutations in the Argonaute ERGO-1, which associates with eri-6/7-dependent 26 nt siRNAs, cause passenger strand stabilization, indicating that ERGO-1 is required to separate the siRNA duplex, presumably through endonucleolytic cleavage of the passenger strand. Thus, like several other siRNA-associated Argonautes with a conserved RNaseH motif, ERGO-1 appears to be required for siRNA maturation.     

Effects of juvenile non-indigenous Carcinus maenas on the growth and condition of juvenile Cancer irroratus

The Atlantic rock crab, Cancer irroratus, is a commercially fished species and a critical prey item for the American lobster, Homarus americanus, in Atlantic Canada. The recent invasion of European green crab, Carcinus maenas, may have significant effects on the growth and condition of native C. irroratus, because both species overlap spatially and temporally and have similar habitat and dietary requirements. To examine such potential effects, we measured the growth of juvenile C. irroratus in the presence of juvenile C. maenas over a period of 4 months (growing season), under the following species combinations: (1) one C. irroratus (10-25 mm CW); (2) two C. irroratus (10-25 mm CW); (3) one C. irroratus (10-25 mm CW) and one C. maenas (10-15 mm CW). Morphological measurements included pre- and post-molt carapace width, chela height, abdomen width (mm), weight (g), and estimates of molt increment (%) and intermolt duration (days). Analysis of the hepatopancreas for % lipid content at the end of the experiment provided an estimate of physiological condition. The effect of the presence of C. maenas on the growth of C. irroratus shifted from negative to positive, when C. irroratus reached CW of 19-22 mm and gained a presumably significant size advantage over C. maenas. The positive effect resulted from increased energy intake through crab consumption. In the absence of crab consumption, the presence of a second crab (conspecific or C. maenas) had no effect on growth. C. irroratus consumed crabs more frequently when the second individual was a green crab than a conspecific. Consumption of C. maenas had a pronounced effect on the growth rate of C. irroratus, resulting in shorter intermolt periods and larger percent molt increments than in the presence of a conspecific. Therefore, the presence of juvenile C. maenas does not appear to have a prolonged negative effect on the growth of C. irroratus; rather, it may provide an additional food item as rock crabs grow, as long as encounters between the two species occur at high enough rates.