Internalization of the constitutively active arginine 1152-->glutamine insulin receptor occurs independently of insulin at an accelerated rate.

Signals controlling the insulin receptor endocytotic pathway have been investigated using the R1152Q insulin receptor mutant (M). This mutant receptor exhibits high levels of insulin-independent kinase activity, impaired autophosphorylation, and lack of an insulin stimulatory effect on both auto- and substrate phosphorylation. NIH-3T3 fibroblasts expressing M receptors displayed a 2.5-fold higher 125I-insulin internalization rate than wild type (WT) but lacked insulin-induced receptor internalization and down-regulation. Cell surface recycling of internalized receptors also occurred at a higher rate in M cells and was unaffected by insulin. Cell preincubation with 35 mM Tris, which inhibits the insulin receptor degradative route, elicited no effect on M receptor recycling but inhibited that of WT by 40%. In contrast, the energy depleter 2,4-dinitrophenol, which inhibits normal insulin receptor retroendocytosis, impaired M receptor recycling 4-fold more effectively than that of WT. The release of internalized intact 125I-insulin was 6-fold greater in M than in WT fibroblasts and was almost completely inhibited by dinitrophenol, whereas insulin degradation by M cells was 4-fold decreased as compared with WT. Thus, internalization and recycling of the constitutively active Gln1152 receptor kinase occur in the absence of autophosphorylation. However, tyrosine phosphorylation appears to be required for proper sorting of endocytosed insulin receptors

Mutation in a conserved motif next to the insulin receptor key autophosphorylation sites de-regulates kinase activity and impairs insulin action.

We have recently reported two non-insulin-dependent diabetic patients exhibiting a heterozygous point mutation (R1152-Q) next to the key tyrosine autophosphorylation sites (Y1146, Y1150, Y1151) of the insulin receptor. In the present study, we demonstrate that the Q1152 mutation alters a previously unrecognized consensus sequence in the insulin receptor family of tyrosine kinases. To define the effect of this alteration on insulin receptor function, the mutant insulin receptor (Q1152) was constructed and overexpressed in NIH-3T3 cells. In spite of normal insulin binding, "in vivo" and "in vitro" autophosphorylation as well as transphosphorylation by the wild-type receptor (WT) were deficient in Q1152 as compared with the transfected WT receptors. Insulin-stimulated kinase activity toward poly(Glu, Tyr) 4:1 and the endogenous substrates p120 and p175 were also impaired in Q1152. However, insulin-independent kinase activity of Q1152 was 2-5-fold higher than that of WT. While insulin stimulated 2-deoxyglucose uptake and glycogen synthase activity in WT-transfected cells with a sensitivity proportional to receptor number, no insulin stimulation was observed in Q1152 cells. Similar to the kinase, insulin-independent glycogen synthase activity and 2-deoxyglucose uptake were 2-fold higher in Q1152 than in either WT or parental cells. We conclude that the Q1152 mutation deregulates insulin receptor kinase and generates insulin insensitivity in cells. Alterations in this highly conserved region of the insulin receptor may contribute to non-insulin dependent diabetes mellitin pathogenesis in humans

Individual epidermal growth factor receptor autophosphorylation sites do not stringently define association motifs for several SH2-containing proteins

To determine whether individual autophosphorylation sites in the epidermal growth factor (EGF) receptor define specific interaction sites for the in vivo association of signal transduction proteins that contain src homology 2 (SH2) domains, the capacity of wild-type and mutant EGF receptors to associate with several SH2 domain-containing proteins has been assayed. Mutants included receptors with single autophosphorylation site mutations at each of five autophosphorylation sites and receptors in which multiple autophosphorylation sites were removed by point mutation or deletion of carboxyl-terminal residues. Receptor association, as measured by coimmunoprecipitation, has been determined for phospholipase C-gamma 1, the ras GTPase-activating protein, the p85 subunit of phosphatidylinositol 3-kinase, and the src homology and collagen protein. In contrast to data obtained with single autophosphorylation site mutants of other receptor tyrosine kinases, none of the EGF receptor single site mutants was dramatically impaired in its capacity to associate with any of these SH2-containing proteins. However, association was completely abrogated when all five autophosphorylation sites were mutated or removed by deletion. These results indicate that individual autophosphorylation sites in the EGF receptor are not stringently required for the recognition and association of different SH2-containing substrates. Thus, EGF receptor autophosphorylation sites seem to be flexible and/or compensatory in their capacity to mediate association with these four SH2-containing substrates

La remarquable sélectivité de feuille avant la ponte chez le papillon Cameraria ohridella n’est pas propre à cette espèce dans le genre Cameraria

Cameraria ohridella, a recent leaf-mining invader of the horse-chestnut tree Aesculus hippocastanum, likely persists as a strong pest with unusually high levels of incidence. For example, among Cameraria moths, C. ohridella ordinarily features more than hundred times denser upon Aesculus leaflets than C. gr. guttifinitella is upon leaves of Quercus emoryi. There is general agreement to consider that the singular success of C. ohridella in mining Aesculus leaves mainly results from deficient top-down control, especially extremely low level of parasitoids efficiency. Another remarkable aspect of C. ohridella outbreaks however is the very high level of leaves acceptance (low selectivity) of females prior to egg-laying, which may often rise up near to 100 %. Would this low bottom-up control also feature (or not) as a specificity of this species within the genus Cameraria? Using an appropriate indirect method, it is shown that the proportions of "acceptable" leaves by ovipositing females are substantially similar (and high) in both Cameraria species (96 % and 78 % for Cameraria ohridella and C. gr. guttifinitella respectively), in spite of the dramatically different proportions of "accepted" (= actually oviposited) leaves (89 % and 2 % respectively). In turn, this would contribute to make still more relevant the currently oriented focus upon top-down regulation as one of the major lever for a better control of Cameraria ohridellaCameraria ohridella, microlépidoptère dont les chenilles minent les feuilles du Marronnier Aesculus hippocastanum, se révèle un envahisseur persistant et tenace depuis son apparition en Europe, il y a une trentaine d'années. Ses niveaux d'incidence, ordinairement très élevés, contrastent avec ceux, bien plus modérés, rencontrés chez d'autres espèces du genre Cameraria. Ainsi, par exemple, C. gr. guttifinitella est cité avec une incidence plus faible de près de deux ordres de grandeurs sur les feuilles de son hôte, Quercus emoryi aux Etats-Unis. La performance numérique exceptionnelle de C. ohridella est généralement attribuée principalement à une remarquable déficience de régulation des populations par les parasitoïdes, lesquels d'ordinaire n'affectent guère plus de quelques pour cent des populations de C. ohridella. Ceci étant, un autre aspect remarquable chez C. ohridella est la très faible sélectivité dont font montre les femelles pondeuses parmi les feuilles de Marronnier qui s'offrent comme support de ponte; la proportion de feuilles acceptées étant souvent voisine des 100 %. Cette faible sélectivité est évidemment de nature à apporter une contribution supplémentaire au succès numérique de C. ohridella. Cette faible sélectivité pour les supports de ponte est-elle également une caractéristique distinctive de l'espèce au sein du genre Cameraria? Ou bien au contraire, d'autres espèces du genre, telle C. gr. guttifinitella, présentent-elles aussi des exigences sélectives faibles vis-à-vis de leur support de ponte? L'estimation du degré de sélectivité ("ratio d'acceptabilité des feuilles α") n'étant, en général, pratiquement pas réalisable à partir des données brutes de terrain, on fait par conséquent usage ici d'une méthode d'inférence indirecte du ratio d'acceptabilité (résumée en Annexe). On montre alors que les proportions de feuilles (respectivement de Aesculus et Quercus) que C. ohridella et C. gr. guttifinitella considèrent comme potentiellement acceptables sont relativement voisines et élevées (96 % et 78 %) alors que les incidences (proportions de feuilles effectivement acceptées c'est-à-dire minées) diffèrent, comme on l'a dit, de près de deux ordres de grandeur (89 % et 2 % respectivement) entre les deux espèces de Cameraria. Un faible niveau de sélectivité parmi les feuilles, sites potentiels de ponte, ne semble donc pas spécifique à l'espèce C. ohridella. Ce résultat ne fait dès lors que renforcer le bien-fondé des études visant à restaurer le niveau de régulation des populations envahissantes de C. ohridella par une meilleure efficience de la prédation, notamment via parasitoïdes

Insulin and insulin-like growth factor I (IGF I) stimulate phosphorylation of a Mr 175,000 cytoskeleton-associated protein in intact FRTL5 cells.

Abstract FRTL5 rat thyroid cells possess separate high affinity receptors for insulin and insulin-like growth factor I (IGF I) that undergo beta-subunit phosphorylation upon interaction with the specific ligand. Phosphorylation is rapid and dose-dependent and occurs primarily on tyrosine residues. Within 2 min, both insulin and IGF I also give rise to a Mr 175,000 phosphoprotein (pp175) that can be immunoprecipitated by anti-phosphotyrosine antibody (alpha-Tyr(P]. Phosphorylation of pp175 occurs on serine and threonine as well as tyrosine residues. When FRTL5 cells are solubilized with 1% Triton X-100, alpha-Tyr(P) immunoprecipitates phosphorylated insulin and IGF I receptors but little pp175 from the Triton-soluble fraction. After treatment of the Triton-insoluble portion with 1% sodium dodecyl sulfate at 100 degrees C, pp175 can be identified by immunoprecipitation with alpha-Tyr(P). The fraction of FRTL5 cells that remains after extraction of an attached monolayer with 1% Triton for 5 min at 22 degrees C contains most of the cytoskeleton and also nuclei. Extraction of this 32P-labeled cytoskeleton preparation with sodium dodecyl sulfate followed by alpha-Tyr(P) immunoprecipitation results in almost complete recovery of the pp175 content of the cells. When a nuclear fraction was prepared from FRTL5 cells by differential centrifugation, pp175 was not found in the nuclear pellet from labeled cells, but greater than 80% of pp175 was recovered in the supernatant. We conclude that pp175 is a common substrate for insulin and IGF I receptor kinases which, in FRTL5 cells, is associated with the cytoskeleton. It is suggested that phosphorylation of proteins associated with cytoskeletal elements could be involved in insulin and IGF I action in cells

Consumer Perceptions of Cigarette Design in France: A Comparison of Regular, Slim, Pink and Plain Cigarettes

Introduction The cigarette, like the cigarette pack, is used by tobacco companies as a promotional tool. We explore how the cigarette could potentially be used as a dissuasive tool. Methods An online survey was conducted with 15-30 year old smokers and non-smokers (N=998) in France to explore their perceptions of a plain cigarette (grey with no brand name) and three branded cigarettes (regular, slim, pink). Participants were randomly assigned to view the plain cigarette and either the regular, slim or pink cigarette. They were asked to rate the cigarettes by Appeal (tastiest, highest quality, most expensive), Harm (most dangerous, most effective for motivating people to talk about tobacco dangers), and Perceived behavioral impact (most effective to convince teenagers not to start, to motivate smokers to reduce consumption and quit). Results In comparison to the grey cigarette, each of the branded cigarettes were considered more appealing, less harmful, and more likely to motivate teenagers to start and less likely to motivate smokers to reduce consumption or quit. Conclusions The study suggests that altering the appearance of the cigarette may reduce cigarette appeal, increase harm perceptions, and deter both young people and smokers. Implications Very little research has focused on dissuasive cigarettes whereas the cigarette stick has become very important for tobacco companies for communication purposes. This is the first study to compare the effect of various branded cigarettes (regular, slim, pink) with a plain grey cigarette on young adult smokers and non-smokers. The findings suggest that a plain grey cigarette can reduce cigarette appeal, increase perceptions of harm, and may deter use among both smokers and non-smokers

Lysosomal Targeting of Epidermal Growth Factor Receptors via a Kinase-dependent Pathway Is Mediated by the Receptor Carboxyl-terminal Residues 1022-1123

Binding of epidermal growth factor (EGF) to its receptor induces rapid internalization and degradation of both ligand and receptor via the lysosomal pathway. To study the mechanism of intracellular sorting of EGF-EGF receptor complexes to lysosomes, NIH 3T3 cells transfected with wild-type and mutant EGF receptors were employed. The kinetics of 125I-EGF trafficking was analyzed using low concentrations of the ligand to avoid saturation of the specific sorting system. The relative size of the pool of internalized 125I-EGF-receptor complexes that were capable of recycling decreased as receptors traversed the endosomal system. The rate of 125I-EGF sequestration from the recycling pathway correlated with the rate of 125I-EGF transition from early to late endosomes as measured by Percoll gradient fractionation. Deletion of the last 63 amino acids of the EGF receptor cytoplasmic tail did not inhibit the process of sequestration and targeting to the late endosomes and lysosomes. Truncation of the 123 residues, however, resulted in impaired lysosomal targeting and increased recycling of EGF. Receptor mutant in which 165 residues were deleted displayed maximal ability to recycle and a minimal extent of sorting to the late endosomes. The data suggest that two regions of the EGF receptor molecule, residues 1022-1063 and to a lesser extent residues 1063-1123, contribute in the regulation of routing of EGF receptors to the degradation pathway. The kinase-negative receptor mutant recycled EGF more intensively compared with the wild-type receptor, and the transport of this mutant to late endosomes was inhibited. These results support the view that the receptor kinase activity is important for ligand-induced sorting of EGF receptors to the pathway of lysosomal degradation

Contributo alla flora della Libia in base a piante raccolte dall'ottobre 1911 al luglio 1912 / di Augusto Beguinot e Antonio Vaccari

Contributo alla flora della Libia in base a piante raccolte dall'ottobre 1911 al luglio 1912 / di Augusto Beguinot e Antonio Vaccari Roma : Tip. Nazionale di G. Bertero e C., 1912 72 p., 10 c. di tav. : ill. ; 25 cm In testa al front.: Ministero degli Affari esteri, Direzione centrale degli affari coloniali, Ufficio di studi colonial