RESUMO
Cu+ -chaperones are a diverse group of proteins that allocate Cu+ ions to specific copper proteins, creating different copper pools targeted to specific physiological processes. Symbiotic nitrogen fixation carried out in legume root nodules indirectly requires relatively large amounts of copper, for example for energy delivery via respiration, for which targeted copper deliver systems would be required. MtNCC1 is a nodule-specific Cu+ -chaperone encoded in the Medicago truncatula genome, with a N-terminus Atx1-like domain that can bind Cu+ with picomolar affinities. MtNCC1 is able to interact with nodule-specific Cu+ -importer MtCOPT1. MtNCC1 is expressed primarily from the late infection zone to the early fixation zone and is located in the cytosol, associated with plasma and symbiosome membranes, and within nuclei. Consistent with its key role in nitrogen fixation, ncc1 mutants have a severe reduction in nitrogenase activity and a 50% reduction in copper-dependent cytochrome c oxidase activity. A subset of the copper proteome is also affected in the ncc1 mutant nodules. Many of these proteins can be pulled down when using a Cu+ -loaded N-terminal MtNCC1 moiety as a bait, indicating a role in nodule copper homeostasis and in copper-dependent physiological processes. Overall, these data suggest a pleiotropic role of MtNCC1 in copper delivery for symbiotic nitrogen fixation.
Assuntos
Medicago truncatula , Fixação de Nitrogênio , Fixação de Nitrogênio/genética , Medicago truncatula/genética , Medicago truncatula/metabolismo , Cobre/metabolismo , Nódulos Radiculares de Plantas/metabolismo , Simbiose/fisiologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismoRESUMO
Symbiotic nitrogen fixation in legume root nodules requires a steady supply of molybdenum for synthesis of the iron-molybdenum cofactor of nitrogenase. This nutrient has to be provided by the host plant from the soil, crossing several symplastically disconnected compartments through molybdate transporters, including members of the MOT1 family. Medicago truncatula Molybdate Transporter (MtMOT) 1.2 is a Medicago truncatula MOT1 family member located in the endodermal cells in roots and nodules. Immunolocalization of a tagged MtMOT1.2 indicates that it is associated to the plasma membrane and to intracellular membrane systems, where it would be transporting molybdate towards the cytosol, as indicated in yeast transport assays. Loss-of-function mot1.2-1 mutant showed reduced growth compared with wild-type plants when nitrogen fixation was required but not when nitrogen was provided as nitrate. While no effect on molybdenum-dependent nitrate reductase activity was observed, nitrogenase activity was severely affected, explaining the observed difference of growth depending on nitrogen source. This phenotype was the result of molybdate not reaching the nitrogen-fixing nodules, since genetic complementation with a wild-type MtMOT1.2 gene or molybdate-fortification of the nutrient solution, both restored wild-type levels of growth and nitrogenase activity. These results support a model in which MtMOT1.2 would mediate molybdate delivery by the vasculature into the nodules.
Assuntos
Proteínas de Transporte de Ânions/fisiologia , Medicago truncatula/metabolismo , Molibdênio/metabolismo , Proteínas de Plantas/fisiologia , Nódulos Radiculares de Plantas/metabolismo , Proteínas de Transporte de Ânions/metabolismo , Medicago truncatula/ultraestrutura , Microscopia Confocal , Microscopia Eletrônica , Proteínas de Plantas/metabolismo , Nódulos Radiculares de Plantas/ultraestruturaRESUMO
Zinc (Zn) is an essential nutrient for plants that is involved in almost every biological process. This includes symbiotic nitrogen fixation, a process carried out by endosymbiotic bacteria (rhizobia) living within differentiated plant cells of legume root nodules. Zn transport in nodules involves delivery from the root, via the vasculature, release into the apoplast and uptake into nodule cells. Once in the cytosol, Zn can be used directly by cytosolic proteins or delivered into organelles, including symbiosomes of infected cells, by Zn efflux transporters. Medicago truncatula MtMTP2 (Medtr4g064893) is a nodule-induced Zn-efflux protein that was localized to an intracellular compartment in root epidermal and endodermal cells, as well as in nodule cells. Although the MtMTP2 gene is expressed in roots, shoots, and nodules, mtp2 mutants exhibited growth defects only under symbiotic, nitrogen-fixing conditions. Loss of MtMTP2 function resulted in altered nodule development, defects in bacteroid differentiation, and severe reduction of nitrogenase activity. The results presented here support a role of MtMTP2 in intracellular compartmentation of Zn, which is required for effective symbiotic nitrogen fixation in M. truncatula.
RESUMO
Molybdenum, as a component of the iron-molybdenum cofactor of nitrogenase, is essential for symbiotic nitrogen fixation. This nutrient has to be provided by the host plant through molybdate transporters. Members of the molybdate transporter family Molybdate Transporter type 1 (MOT1) were identified in the model legume Medicago truncatula and their expression in nodules was determined. Yeast toxicity assays, confocal microscopy, and phenotypical characterization of a Transposable Element from Nicotiana tabacum (Tnt1) insertional mutant line were carried out in the one M. truncatula MOT1 family member specifically expressed in nodules. Among the five MOT1 members present in the M. truncatula genome, MtMOT1.3 is the only one uniquely expressed in nodules. MtMOT1.3 shows molybdate transport capabilities when expressed in yeast. Immunolocalization studies revealed that MtMOT1.3 is located in the plasma membrane of nodule cells. A mot1.3-1 knockout mutant showed impaired growth concomitant with a reduction of nitrogenase activity. This phenotype was rescued by increasing molybdate concentrations in the nutritive solution, or upon addition of an assimilable nitrogen source. Furthermore, mot1.3-1 plants transformed with a functional copy of MtMOT1.3 showed a wild-type-like phenotype. These data are consistent with a model in which MtMOT1.3 is responsible for introducing molybdate into nodule cells, which is later used to synthesize functional nitrogenase.
Assuntos
Proteínas de Transporte de Ânions/metabolismo , Medicago truncatula/metabolismo , Molibdênio/metabolismo , Nitrogenase/metabolismo , Nódulos Radiculares de Plantas/metabolismo , Proteínas de Plantas/metabolismoRESUMO
OBJECTIVES: Define the impact and causes of non-adherent type-2 diabetes mellitus (DM2) patients, possible solutions and the role of the different health care professionals involved in the treatment. DESIGN: Structured questionnaire rating by a two-round Delphi method. LOCATION: The study was conducted in the Primary Care settings. PARTICIPANTS: The expert panel consisted of renowned medical professionals with extensive experience in diabetes. MAIN MEASUREMENTS: Assessment through a 9-point Likert scale, of the degree of agreement or disagreement on 131 items grouped into 4 blocks: impact; causes of nonadherence; diagnosis of non-adherence, and possible causes, solutions and role of the different professionals involved in adherence. RESULTS: The participation rate was 76.31%. The primary care health professionals agreed on 110 of the 131 proposals statements (84%), showing agreement on 102 items (77.9%) and disagreement in 8 (6.1%). Consensus was not reached on 21 items. CONCLUSIONS: The lack of adherence of DM2 patients makes the achievement of therapeutic control difficult. The medical practice needs to have specific training and enough resources to reduce the impact of the lack of therapeutic compliance.
Assuntos
Diabetes Mellitus Tipo 2/terapia , Cooperação do Paciente , Técnica Delphi , Humanos , Inquéritos e QuestionáriosRESUMO
OBJECTIVE: Only a small percentage of patients with acute stroke receive thrombolytic therapy, mainly due to late hospital arrival. Factors excluding those who arrive within 3h after stroke onset are less well known. PATIENTS AND METHODS: During the first year after implementing a protocol for stroke thrombolysis, we prospectively evaluated all patients with stroke admitted to our center within 3h from onset. Within-hospital time intervals were calculated and the reasons for exclusion from thrombolysis were analyzed. RESULTS: Ninety-six patients (representing 16% of all stroke patients admitted) arrived in less than 3h, and 25 of them (representing 7.5% of all patients with ischemic stroke) received thrombolytic therapy, with a door-to-needle interval of 51 min (range, 33-121). The reasons that accounted for 75% of therapy exclusions were non-modifiable (a too mild or improving deficit, and intracranial hemorrhage), except for a time window exceeded, which would probably require increasing public awareness about stroke. CONCLUSIONS: Most reasons for not applying thrombolysis to patients who arrive early enough are non-modifiable. Minimizing the door-to-needle time could compensate for late hospital arrival, which continues to be the main reason for not applying this therapy to stroke patients throughout the world.
Assuntos
Serviço Hospitalar de Emergência , Fibrinolíticos/administração & dosagem , Acidente Vascular Cerebral/tratamento farmacológico , Ativador de Plasminogênio Tecidual/administração & dosagem , Adulto , Idoso , Idoso de 80 Anos ou mais , Diagnóstico Precoce , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Seleção de Pacientes , Estudos Prospectivos , Acidente Vascular Cerebral/diagnóstico , Fatores de Tempo , Índices de Gravidade do TraumaRESUMO
The prevalence and significance of microalbuminuria in hypertensive patients with impaired fasting glucose (IFG) has received very little attention. A total of 10,320 hypertensive patients who attended primary care centers were enrolled in this study, and the final analysis was done in 7625 patients: 1459 without IFG (plasma glucose <100 mg/dl), 3010 with IFG (plasma glucose > or =100 mg/dl and <126 mg/dl), and 3156 with type 2 diabetes (plasma glucose >126 mg/dl). Microalbuminuria was determined using the Micro Albustix reactive strip from Bayer (high urinary albumin excretion [UAE]: Albumin/creatinine ratio > or =3.4 mg/mmol). The proportion of patients with high UAE was 39.4, 48.3, and 65.6%, respectively, in the three groups (P < 0.01 for the trend). The differences in UAE between the group with IFG and the group with normal fasting glucose persisted after adjustment for age, gender, systolic BP, fasting plasma glucose, and cardiovascular comorbidity (odds ratio 1.74; 95% confidence interval 1.08 to 2.80). Hypertensive patients with IFG and high UAE showed a higher prevalence of ischemic heart disease, cardiac insufficiency, left ventricular hypertrophy, atrial fibrillation, and renal insufficiency than the group with normal UAE. Global prevalence of cardiovascular conditions was 30.4% in the group with high UAE compared with 21.4% in the group with normal UAE (odds ratio 1.60; 95% confidence interval 1.31 to 1.95). It is concluded that almost half of hypertensive patients with IFG have high UAE and a higher prevalence of associated cardiovascular involvement and renal insufficiency.
Assuntos
Albuminúria/complicações , Doenças Cardiovasculares/epidemiologia , Hiperglicemia/complicações , Hipertensão/complicações , Idoso , Albuminúria/fisiopatologia , Pressão Sanguínea/fisiologia , Doenças Cardiovasculares/complicações , Doenças Cardiovasculares/fisiopatologia , Estudos Transversais , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/fisiopatologia , Feminino , Humanos , Hiperglicemia/fisiopatologia , Hipertensão/fisiopatologia , Masculino , Pessoa de Meia-Idade , Prevalência , Fatores de Risco , EspanhaRESUMO
The objective of this study was to assess the relationship between urinary albumin excretion (UAE) and GF across the spectrum of the glucose metabolism abnormalities in a large population of patients with hypertension. The Microaluminuria en Pacientes con Glucemia Basal Alterada (MAGAL) is a multicenter, cross-sectional study that was carried out by 1723 primary care physicians. A total of 6227 patients with essential hypertension (in three groups: [1] normal fasting glucose <100 mg/dl, [2] impaired fasting glucose > or =100 to 126 mg/dl, and [3] type 2 diabetes) were analyzed in this substudy. GFR was estimated by using the Modification of Diet in Renal Disease (MDRD) abbreviated equation. A single first-morning urine albumin/creatinine ratio was measured using Bayer reagent strip Microalbustix, a semiquantitative method. Abnormal UAE was defined as an albumin/creatinine ratio > or =3.4 mg/mmol (equivalent to > or =30 mg/g). The prevalence of abnormal UAE, > or =3.4 mg/mmol, increased across the spectrum of glucose abnormalities: 39.7, 46.2, 48.6, and 65.6% for normoglycemic, low-range, and high-range impaired fasting glucose and diabetes, respectively. UAE was positively related to SBP (P = 0.003) and inversely to GFR (P < 0.001). Renal insufficiency (GFR <60 ml/min per 1.73 m2) was present in 21.8% of the patients, more frequently older patients, women, and those with diabetes. The factors that were related to renal insufficiency were UAE > or =3.4 mg/mmol (odds ratio 1.86; 95% confidence interval 1.60 to 2.17) and diabetes (odds ratio 1.62; 95% confidence interval 1.29 to 2.04). There is a close relationship between abnormal UAE and renal insufficiency in essential hypertension. This is more marked in patients with diabetes and moderate in patients with high-range impaired fasting glucose.