RESUMO
Aedes aegypti (Linnaeus) (Diptera: Culicidae) is the main vector of viruses causing dengue, chikungunya, Zika, and yellow fever, worldwide. This report focuses on immuno-blocking four critical proteins in the female mosquito when fed on blood containing antibodies against ferritin, transferrin, one amino acid transporter (NAAT1), and acetylcholinesterase (AchE). Peptides from these proteins were selected, synthetized, conjugated to carrier proteins, and used as antigens to immunize New Zealand rabbits. After rabbits were immunized, a minimum of 20 female mosquitos were fed on each rabbit, per replicate. No effect in their viability was observed after blood-feeding; however, the number of infertile females was 20% higher than the control when fed on AchE-immunized rabbits. The oviposition period was significantly longer in females fed on immunized rabbits than those fed on control (non-immunized) rabbits. Fecundity (eggs/female) of treated mosquitoes was significantly reduced (about 50%) in all four treatments, as compared with the control. Fertility (hatched larvae) was also significantly reduced in all four treatments, as compared with the control, being the effect on AchE and transferrin the highest, by reducing hatching between 70 and 80%. Survival to the adult stage of the hatched larvae showed no significant effect, as more than 95% survival was observed in all treatments, including the control. In conclusion, immuno-blocking of these four proteins caused detrimental effects on the mosquito reproduction, being the effect on AchE the most significant.
Assuntos
Acetilcolinesterase/imunologia , Aedes/fisiologia , Anticorpos/imunologia , Imunização/veterinária , Proteínas de Insetos/imunologia , Sistemas de Transporte de Aminoácidos/imunologia , Animais , Feminino , Ferritinas/imunologia , Fertilidade , Hemolinfa , Imunoglobulina G/imunologia , Mosquitos Vetores/fisiologia , Oviposição , Coelhos , Reprodução , Transferrina/imunologiaRESUMO
The chloroplast 24 kDa RNA binding protein (24RNP) from Spinacea oleracea is a nuclear encoded protein that binds the 3' untranslated region (3'UTR) of some chloroplast mRNAs and seems to be involved in some processes of mRNA metabolism, such as 3'UTR processing, maturation and stabilization. The 24RNP is similar to the 28RNP which is involved in the correct maturation of petD and psbA 3'UTRs, and when phosphorylated, decreases its binding affinity for RNA. In the present work, we determined that the recombinant 24RNP was phosphorylated in vitro either by an animal protein kinase C, a plant Ca(2+)-dependent protein kinase, or a chloroplastic kinase activity present in a protein extract with 3'-end processing activity in which the 24RNP is also present. Phosphorylation of 24RNP increased the binding capacity (B(max)) 0.25 time for petD 3'UTR, and three times for psbA 3'UTR; the affinity for P-24RNP only increased when the interaction with petD was tested. Competition experiments suggested that B(max), not K(d), might be a more important factor in the P-24RNP-3'UTR interaction. The data suggested that the 24RNP role in chloroplast mRNA metabolism may be regulated in vivo by changes in its phosphorylation status carried out by a chloroplastic kinase.
Assuntos
Regiões 3' não Traduzidas/metabolismo , Cloroplastos/metabolismo , Complexo Citocromos b6f/metabolismo , Complexo de Proteína do Fotossistema II/metabolismo , Proteínas de Plantas/metabolismo , Proteínas de Ligação a RNA/metabolismo , Fosforilação , Proteínas de Plantas/genética , Ligação Proteica , Proteínas de Ligação a RNA/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Spinacia oleraceaRESUMO
A 120-kDa glycoprotein was found in beet root (Beta vulgaris L.) plasma membranes. This protein could be phosphorylated in a Ca2+-independent manner. Its carbohydrate moiety was composed of both O-linked galactose-beta(1-3)-N-acetylgalactosamine disaccharides (which bind peanut agglutinin) and N-linked concanavalin A (ConA)-binding oligosaccharides. The phosphorylation of this protein was rapid, half-saturated with 6 microM ATP and higher at alkaline pH values. This protein was phosphorylated more efficiently with Mn-ATP as substrate than with Mg-ATP. This phosphorylation increased when plasma membranes were illuminated with low-fluence blue light, a fact suggesting that the 120-kDa glycoprotein could be similar to phototropin: a blue-light photoreceptor involved in phototropism. This protein was purified using a ConA-Sepharose column. The phosphorylation of the purified protein could be observed, but it was much lower than that of the 120-kDa protein in plasma membranes. In addition, it was not enhanced by light. Some possible explanations for this photosensitivity loss are discussed.
Assuntos
Beta vulgaris/metabolismo , Glicoproteínas de Membrana/isolamento & purificação , Fosfotransferases/isolamento & purificação , Raízes de Plantas/metabolismo , Trifosfato de Adenosina/farmacologia , Beta vulgaris/efeitos da radiação , Cálcio/farmacologia , Metabolismo dos Carboidratos , Membrana Celular/metabolismo , Concentração de Íons de Hidrogênio , Luz , Glicoproteínas de Membrana/metabolismo , Glicoproteínas de Membrana/efeitos da radiação , Fosforilação/efeitos dos fármacos , Fosforilação/efeitos da radiação , Fosfotransferases/metabolismo , Fosfotransferases/efeitos da radiação , Raízes de Plantas/efeitos da radiaçãoRESUMO
Several plasma-membrane proteins from beet root (Beta vulgaris L.) have been functionally incorporated into reconstituted proteoliposomes. These showed H(+)-ATPase activity, measured both as ATP hydrolysis and H+ transport. The proton-transport specific activity was 10 times higher than in plasma membranes, and was greatly stimulated by potassium and valinomycin. These proteoliposomes also showed calcium-regulated protein kinase activity. This kinase activity is probably due to a calmodulin-like domain protein kinase (CDPK), since two protein bands were recognized by antibodies against soybean and Arabidopsis CDPK. This kinase phosphorylated histone and syntide-2 in a Ca(2+)-dependent manner. Among the plasma-membrane proteins phosphorylated by this kinase, was the H(+)-ATPase. When the H(+)-ATPase was either prephosphorylated or assayed in the presence of Ca2+, both the ATP-hydrolysis and the proton-transport activities were slower. This inhibition was reversed by an alkaline-phosphatase treatment. A trypsin treatment (that has been reported to remove the C-terminal autoinhibitory domain from the H(+)-ATPase) also reversed the inhibition caused by phosphorylation. These results indicate that a Ca(2+)-dependent phosphorylation, probably caused by a CDPK, inhibits the H(+)-ATPase activities. The substrate of this regulatory phosphorylation could be the H(+)-ATPase itself, or a different protein influencing the ATPase activities.
Assuntos
Cálcio/metabolismo , Chenopodiaceae/enzimologia , Raízes de Plantas/enzimologia , ATPases Translocadoras de Prótons/metabolismo , Hidrólise , Transporte de Íons , Fosforilação , Proteínas Quinases/metabolismo , Proteolipídeos , ATPases Translocadoras de Prótons/antagonistas & inibidores , Tripsina/farmacologiaRESUMO
Obese children 8-12 years old from 61 families were randomized to treatment groups that targeted increased exercise, decreased sedentary behaviors, or both (combined group) to test the influence of reinforcing children to be more active or less sedentary on child weight change. Significant decreases in percentage overweight were observed after 4 months between the sedentary and the exercise groups (-19.9 vs. -13.2). At 1 year, the sedentary group had a greater decrease in percentage overweight than did the combined and the exercise groups (-18.7 vs. -10.3 and -8.7) and greater decrease in percentage of body fat (-4.7 vs. -1.3). All groups improved fitness during treatment and follow-up. Children in the sedentary group increased their liking for high-intensity activity and reported lower caloric intake than did children in the exercise group. These results support the goal of reducing time spent in sedentary activities to improve weight loss.
Assuntos
Exercício Físico , Estilo de Vida , Obesidade/terapia , Redução de Peso , Criança , Estudos de Coortes , Feminino , Seguimentos , Humanos , Masculino , TelevisãoRESUMO
A novel human mRNA whose expression is induced over 200-fold in B lymphocytes by latent Epstein-Barr virus (EBV) infection was reverse transcribed, cloned, and sequenced. The mRNA is predicted to encode a protein containing four peptides which precisely match amino acid sequences from a previously identified 55-kDa actin-bundling protein, p55. In vitro translation of the cDNA results in a 55-kDa protein which binds to actin filaments in the presence of purified p55 from HeLa cells. The p55 mRNA is undetectable in non-EBV-infected B- and T-cell lines or in a myelomonocytic cell line (U937). Newly infected primary human B lymphocytes, EBV-transformed B-cell lines, latently infected Burkitt tumor cells expressing EBNA2 and LMP1, a chronic myelogenous leukemia cell line (K562), and an osteosarcoma cell line (TK143) contain high levels of p55 mRNA or protein. In EBV-transformed B cells, p55 localizes to perinuclear cytoplasm and to cell surface processes that resemble filopodia. The p55 mRNA is detected at high levels in spleen and brain tissues, at moderate levels in lung and placenta tissues, and at low levels in skeletal muscle, liver, and tonsil tissues and is undetectable in heart, kidney, pancreas, and bone marrow tissues. Immunohistochemical staining of human brain tissue demonstrates p55 localization to the perinuclear cytoplasm and dendritic processes of many, but not all, types of cortical or cerebellar neurons, to glial cells, and to capillary endothelial cells. In cultured primary rat neurons, p55 is distributed throughout the perinuclear cytoplasm and in subcortical filamentous structures of dendrites and growth cones. p55 is highly evolutionarily conserved since it shows 40% amino acid sequence identity to the Drosophila singed gene product and 37% identity to fascin, an echinoderm actin-bundling protein. The evolutionary conservation of p55 and its lack of extensive homology to other actin-binding proteins suggest that p55 has specific microfilament-associated functions in cells in which it is differentially expressed, including neural cells and EBV-transformed B lymphocytes.
Assuntos
Linfócitos B/metabolismo , Regulação da Expressão Gênica , Herpesvirus Humano 4/genética , Peptídeos e Proteínas de Sinalização Intracelular , Proteínas de Membrana , Proteínas dos Microfilamentos/genética , Sequência de Aminoácidos , Linfócitos B/virologia , Sequência de Bases , Evolução Biológica , Transformação Celular Viral , DNA Complementar/isolamento & purificação , Proteína GAP-43 , Humanos , Glicoproteínas de Membrana/análise , Proteínas dos Microfilamentos/análise , Proteínas dos Microfilamentos/química , Dados de Sequência Molecular , Peso Molecular , Substrato Quinase C Rico em Alanina Miristoilada , Proteínas do Tecido Nervoso/análise , Especificidade de Órgãos , Proteínas/análiseRESUMO
Two studies on resting metabolic rate (RMR) in bulimia nervosa were conducted. The first study compared RMR before treatment in 25 normal-weight women with bulimia nervosa and 20 control subjects of similar height, weight, body composition, age, and activity level. No significant difference in RMR adjusted for fat-free weight was observed. The second study sought to determine whether RMR in women with bulimia nervosa changed if they ceased vomiting and resumed eating in a more normal fashion after cognitive-behavioral treatment. There was no differential change in RMR from pre- to posttreatment for the "improved" bulimics (9 of 12 subjects who received treatment) relative to 13 control subjects who were also tested twice at the same time intervals as the treated bulimia nervosa subjects. These findings do not support the hypothesis that normal-weight women with bulimia nervosa have a suppressed RMR, nor is it altered with treatment compared with matched control subjects.
Assuntos
Metabolismo Basal , Bulimia/metabolismo , Ingestão de Energia , Vômito/metabolismo , Adulto , Terapia Comportamental , Glicemia , Composição Corporal , Bulimia/terapia , Estudos Transversais , Ingestão de Alimentos , Feminino , Humanos , Norepinefrina/sangue , Hormônios Tireóideos/sangueRESUMO
CaM kinase-Gr is a multifunctional Ca2+/calmodulin-dependent protein kinase which is enriched in neurons and T lymphocytes. The kinase is absent from primary human B lymphocytes but is expressed in Epstein-Barr virus (EBV)-transformed B-lymphoblastoid cell lines, suggesting that expression of the kinase can be upregulated by an EBV gene product(s). We investigated the basis of CaM kinase-Gr expression in EBV-transformed cells and the mechanisms that regulate its activity therein by using an EBV-negative Burkitt lymphoma cell line, BJAB, and BJAB cells converted to expression of individual EBV proteins by single-gene transfer. CaM kinase-Gr expression was upregulated in BJAB cells by EBV latent-infection membrane protein 1 (LMP1) but not by LMP2A or by nuclear proteins EBNA1, EBNA2, EBNA3A, and EBNA3C. In LMP1-converted BJAB cells, the kinase was functional and was dramatically activated upon cross-linking of surface immunoglobulin M. Overlapping cDNA clones that encode human CaM kinase-Gr were sequenced, revealing 81% amino acid identity between the rat and human proteins. Transfection of BJAB cells with an expression construct for the human enzyme resulted in a functional kinase which was shown by epitope tagging to localize primarily to cytoplasmic and perinuclear structures. Induction of CaM kinase-Gr expression by LMP1 provides the first example of a Ca2+/calmodulin-dependent protein kinase upregulated by a viral protein. In view of the key role played by LMP1 in B-lymphocyte immortalization by EBV, these findings implicate CaM kinase-Gr as a potential mediator of B-lymphocyte growth transformation.
Assuntos
Antígenos Virais/farmacologia , Linfócitos B , Proteínas Quinases Dependentes de Cálcio-Calmodulina/biossíntese , Transformação Celular Viral , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Herpesvirus Humano 4 , Proteínas da Matriz Viral/farmacologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteína Quinase Tipo 4 Dependente de Cálcio-Calmodulina , Proteínas Quinases Dependentes de Cálcio-Calmodulina/genética , Proteínas Quinases Dependentes de Cálcio-Calmodulina/isolamento & purificação , Compartimento Celular , Núcleo Celular/química , Células Cultivadas , Clonagem Molecular , Reagentes de Ligações Cruzadas , Citoplasma/química , Ativação Enzimática , Indução Enzimática , Imunofluorescência , Humanos , Imunoglobulina M/metabolismo , Dados de Sequência Molecular , Receptores de Antígenos de Linfócitos B/metabolismo , Proteínas Recombinantes/biossíntese , Homologia de Sequência de AminoácidosRESUMO
A comparison of cognitive-behavior therapy alone, desipramine alone, and cognitive-behavior therapy combined with desipramine was made in the treatment of bulimia nervosa. The study was terminated early with an N of only 7 subjects per condition because of a high drop-out rate and lack of positive response in the desipramine alone group compared to the other two groups. By this time it was also apparent that at posttreatment and at 6 months follow-up no benefit was being realized from combining cognitive-behavior therapy with desipramine.
Assuntos
Bulimia/tratamento farmacológico , Bulimia/terapia , Terapia Cognitivo-Comportamental/métodos , Desipramina/uso terapêutico , Adolescente , Adulto , Bulimia/diagnóstico , Terapia Combinada , Feminino , Humanos , Pessoa de Meia-Idade , Escalas de Graduação Psiquiátrica , Inquéritos e QuestionáriosRESUMO
Evaluated children's choice for sedentary versus vigorous activity. Experiment 1 assessed the influence of percent overweight (less than 20%, 20% to 80%, greater than 80%) on choice of a moderately liked vigorous activity at a constant variable ratio (VR) 2 reinforcement schedule versus a highly liked sedentary activity with the schedule varied from VR2 to VR32. All children chose the sedentary choice when the schedules were VR2. As the cost for sedentary activity increased, lean and moderately obese children switched to the vigorous activity, but the very obese children still chose the sedentary activity. Experiment 2 compared moderately obese children's choice between easily accessible, highly or least liked vigorous activities (VR2) and highly liked sedentary activity with the schedule varied from VR2 through VR16. When the reinforcement schedules were equal, and sedentary activity and vigorous activity were rated as equally linked, children chose the sedentary activity. All subjects switched from the sedentary to the vigorous activity, but there were no differences in choice as a function of liking for vigorous activities.
Assuntos
Exercício Físico , Comportamentos Relacionados com a Saúde , Obesidade/psicologia , Criança , Comportamento de Escolha , Feminino , Humanos , Estilo de Vida , Masculino , Meio SocialRESUMO
The phosphorylation of plasma membrane proteins from red beet (Beta vulgaris L.) by radioactive inorganic phosphate was studied. Only few proteins were phosphorylated, among them was one polypeptide with an apparent molecular weight of about 100,000. The phosphorylation of this protein was decreased when orthovanadate was present in the reaction mixture, or when the phosphorylated protein was treated with hydroxylamine. These facts suggest that this protein is a transport ATPase which is phosphorylated in a carboxyl group during the catalytic cycle. This protein was identified immunologically as the plasma membrane H(+)-ATPase. The phosphorylation level of this enzyme was enhanced by dimethyl sulfoxide, whereas potassium ions did not have a significant effect on this level unless ATP was present. ATP stimulated the phosphorylation by inorganic phosphate. This stimulation was more apparent in the presence of potassium ions.
RESUMO
In this study the relationship between caloric intake and activity levels was examined in forty-three 18-month-old children. Caloric consumption was measured during a laboratory meal, and activity levels were assessed using an ambulatory monitor over a 24-hour period. Caloric intake was found to be significantly correlated with activity levels, such that children with a high caloric intake tended to have lower activity levels. This finding suggests that two risk factors influencing the development of obesity, caloric intake and physical activity levels, are likely to occur together in a subset of young children. A possible mechanism underlying this association is discussed.
Assuntos
Comportamento Infantil/fisiologia , Ingestão de Energia/fisiologia , Esforço Físico , Feminino , Humanos , Lactente , Masculino , Obesidade/etiologiaRESUMO
The effects of inorganic phosphate on the plasma membrane H(+)-ATPase of red beet (Beta vulgaris L.) were studied. ATPase activity was inhibited weakly and noncompetitively by phosphate. This anion also relieved the inhibition caused by vanadate by displacing it from the enzyme. From this effect, a dissociation constant for phosphate of 25 millimolar and an extrapolated activity at infinite phosphate concentration of 84% of the activity without inhibitors were calculated. The partial inhibition by phosphate indicates the existence of a catalytically active enzyme-phosphate complex. In the presence of 24% dimethylsulfoxide, the inhibition of ATPase activity by phosphate is much greater than in its absence. This suggests that the active enzyme-phosphate complex could be converted into a covalent phosphoenzyme through a dehydration promoted by the low water activity of the medium. The inhibitory ability of phosphate in 24% dimethylsulfoxide was dependent on the presence of potassium. Potassium ions increased both the affinity for phosphate and the inhibition caused by an infinite phosphate concentration, suggesting that potassium stimulates both phosphate binding and phosphoenzyme formation.
Assuntos
Pletismografia , Varizes/cirurgia , Adulto , Feminino , Seguimentos , Hemodinâmica , Humanos , Ligadura , Masculino , Pessoa de Meia-Idade , Período Pós-Operatório , Varizes/fisiopatologiaAssuntos
Peso Corporal , Ajustamento Social , Adolescente , Sintomas Afetivos/psicologia , Imagem Corporal , Feminino , Humanos , Masculino , Autoimagem , Fatores SexuaisRESUMO
Dark and light oxidation of NADPH was measured in Spirulina maxima thylakoid membranes. The dark reaction was more cyanide sensitive than the light reaction. In light, 83% of the electrons from NADPH produced H2O2 on reducing oxygen, whereas in the dark this number was only 36%. These results are explained by assuming the presence of an electron transport segment common to the photosynthetic and the respiratory chains, so that electrons flowing through the cyanide sensitive oxidase in the dark are diverted to the photosytem (PS) I reaction center (P700). In addition, cytochrome (cyt) c 553 was found to be an electron donor for both cyt oxidase and P700. Half maximum reduction rates were obtained with 7 µM cyt c 553. The intrathylakoidal concentration of cyt c 553 was determined to be 83 µM. About 60% of the respiratory NADPH oxidation activity was lost by extracting the membranes with pentane and was restored by adding plastoquinone (the main photosythetic quinone). NADPH oxidation activity was also inhibited upon washing the membranes with a low salt buffer. This activity was restored by adding partially purified ferredoxin-NADP(+) oxido-reductase (FNR). A model for the electron transport in thylakoids, in which cyt c 553, plastoquinone and FNR participate in both photosynthesis and respiration is proposed.