There was a significant decrease in FCD risk with higher intake of omega-3 PUFA, particularly that originating from fish. There was no evidence to indicate that the intake of other types of dietary fat or fat quantity in the previous 12 months was associated with an altered risk of FCD.

No abstract

No abstract

Tnk1/Kos1 is a non-receptor protein tyrosine kinase implicated in negatively regulating cell growth in a mechanism requiring its intrinsic catalytic activity. Tnk1/Kos1 null mice were created by homologous recombination by deleting the catalytic domain. Both Tnk1 +/À and Tnk1 À/À mice develop spontaneous tumors, including lymphomas and carcinomas, at high rates [27% (14 of 52) and 43% (12 of 28), respectively]. Tnk1/Kos1 expression is silenced in tumors that develop in Tnk1 +/À mice but not in adjacent uninvolved tissue, and silencing occurs in association with Tnk1 promoter hypermethylation. Tissues and murine embryonic fibroblasts derived from Tnk1/Kos1-null mice exhibit proportionally higher levels of basal and epidermal growth factor-stimulated Ras activation that results from increased Ras-guanine exchange factor (GEF) activity. Mechanistically, Tnk1/Kos1 can directly tyrosine phosphorylate growth factor receptor binding protein 2 (Grb2), which promotes disruption of the Grb2-Sos1 complex that mediates growth factor-induced Ras activation, providing dynamic regulation of Ras GEF activity with suppression of Ras. Thus, Tnk1/Kos1 is a tumor suppressor that functions to down-regulate Ras activity.

Oxytocin (OT) receptors (OTRs) mediate reproductive functions, including the initiation of labor and milk ejection. OTR messenger RNA levels are highly regulated, reaching the greatest concentration in the uterus at the end of gestation, and in the mammary gland during lactation. Factors directly effecting changes in OTR gene expression in the mammary gland are not known, so the present studies were done to elucidate possible regulators by characterizing the human OTR gene promoter and 5'-flanking sequence. By analyzing expression of promoter-luciferase constructs, we localized a region between -85 and -65 that was required for both basal and serum-induced expression in a mammary tumor cell line (Hs578T) that expresses inducible, endogenous OTRs. This DNA region contains an ets family target sequence (5'-GGA-3'), and a CRE/AP-1-like motif. The specific Ets factor binding to the OTR promoter was identified, by electrophoretic mobility immunoshift assays, to be GABP alpha/beta. Co-transfection of a -85 OTR/luciferase construct with vectors expressing GABP alpha and GABP beta1 had only a modest effect on expression, but cotransfection with GABP alpha/beta- with c-Fos/c-Jun-expressing plasmids resulted in an increase of almost 10-fold in luciferase activity. Mutation of either the GABP- or CRE-like binding sites obliterated the induction. These findings are consistent with the involvement of protein kinase C activity in serum induction of the endogenous gene in Hs578T cells. We showed the requirement for GABP alpha/beta and c-Fos/c-Jun in endogenous OTR gene expression, using oligonucleotide GABP and AP-1 binding decoys to inhibit serum-induced increases in 125I-labeled OT antagonist binding to Hs578T cells. Our work is the first characterization of the proximal promoter region of the human OTR gene, and it sets the stage for studying regulation of OTR expression in breast cells.

, AND S. L. HoARE. Feedback regulation of arginine biosynthesis in blue-green algae and photosynthetic bacteria. J. Bacteriol. 92:375-379. 1966.-A number of blue-green algae and photosynthetic bacteria synthesize arginine from glutamate via acetylated intermediates. Cell-free extracts of these photosynthetic microorganisms contain an N-acetyl glutamate phosphokinase, which is specifically inhibited by arginine. They also contain a transacetylase which forms ornithine from aN-acetyl ornithine and glutamate. The transacetylase appears to be specific for L-glutamate. Arginine synthesis and its regulation by feedback inhibition in photosynthetic microorganisms differ from that in Escherichia coli and other Enterobacteriaceae.

A major action of oxytocin is to stimulate prostaglandin production in reproductive tissues. The two major enzyme systems involved are cytosolic phospholipase A, (#LA,), which catalyses the formation of arachidonic acid from membrane glycerophospholipids, and prostaglandin endoperoxide-H synthases-1 and -2, which allow conversion of arachidonic acid to prostaglandins. During gestation, the concentrations of all three enzymes rise in the rabbit amnion. Agonists, including oxytocin, increase cPLA, activity, in part, by elevating intracellular Ca2+ concentration, which causes cPLA, to be translocated from the cytosol to intracellular membrane binding sites. Cytosolic PLA, is then activated by a mitogen-activated protein kinase (MAPK)-dependent step. Our studies have elucidated signal pathways involved in oxytocin-stimulated prostaglandin output in both rabbit amnion cells and Chinese hamster ovary cells stably transfected with the rat oxytocin receptor. The two cell types are alike with respect to oxytocin-stimulated intracellular Ca2+ transients, mediation via G,, and the specific MAPK that catalyses the phosphorylation of cPLA,. However, they differ with respect to the mechanisms of upregulation of key enzymes involved in prostaglandin E, synthesis. These findings illustrate the tiers of complementary mechanisms involved in oxytocin stimulation of prostaglandin E,, and the extent of the diversity in the cellular signalling pathways involved. Experimental Physiology (2000) 85S, 51S-58S.

As the oxytocin receptor plays a key role in parturition and lactation, there is considerable interest in defining its structure/functional relationships. We previously showed that the rat oxytocin receptor transfected into Chinese hamster ovary cells was coupled to both G q/11 and G i/o , and that oxytocin stimulated ERK-2 phosphorylation and prostaglandin E 2 synthesis via protein kinase C activity. In this study, we show that deletion of 51 amino acid residues from the carboxyl terminus resulted in reduced affinity for oxytocin and a corresponding rightward shift in the dose-response curve for oxytocin-stimulated [Ca 2؉ ] i . However, oxytocin-stimulated ERK-2 phosphorylation and prostaglandin E 2 synthesis did not occur in cells expressing the truncated receptor. Oxytocin also failed to increase phospholipase A activity or activate protein kinase C, indicating that the mutant receptor is uncoupled from G q -mediated pathways. The ⌬51 receptor is coupled to G i , as oxytocin-stimulated Ca 2؉ transients were inhibited by pertussis toxin, and a G␤␥ sequestrant. Preincubation of ⌬51 cells with the tyrosine kinase inhibitor, genistein, also blocked the oxytocin effect. A ⌬39 mutant had all the activities of the wild type oxytocin receptor. These results show that the portion between 39 and 51 residues from the COOH terminus of the rat oxytocin receptor is required for interaction with G q/11 , but not G i/o . Furthermore, an increase in intracellular calcium was generated via a G i ␤␥-tyrosine kinase pathway from intracellular stores that are distinct from G q -mediated inositol trisphosphate-regulated stores. Oxytocin (OT)1 is a nine-amino acid peptide that stimulates uterine smooth muscle and mammary myoepithelial cell contraction, and prostaglandin production by uterine endometrial and amnion cells. Nucleic acid sequencing of cDNA clones of the oxytocin receptor (OTR) indicated that it is a member of the G protein-coupled receptor (GPCR) superfamily (1). As OT plays a pivotal role in parturition and lactation, there is considerable interest in defining the structure of the OTR. It has been shown for a number of GPCRs that several regions in the cytoplasmic domains contribute directly or indirectly to G protein coupling (see Ref. 2 for a review). The juxtamembrane portions of cytoplasmic loop 3 and cytoplasmic loop 2 of several family members have been implicated in receptor-G protein interactions. In addition, the COOH-terminal region of adrenergic receptors (3, 4) and other receptor types (5-7) is also required for G protein interactions, but this domain does not appear to be important for receptor function of all GPCR family members (8 -10).The COOH-terminal domain of some GPCRs plays an important role in G protein isotype selectivity (11,12). At least four isoforms of the prostaglandin EP3 receptor, differing only at their COOH-terminal tails (produced by alternative splicing), couple to different G proteins to activate different second messenger systems (13,14). The COOH terminus of the human parathyroid h...