Immunolocalization of plasma-membrane H+-ATPase and tonoplast-type pyrophosphatase in the plasma membrane of the sieve element-companion cell complex in the stem of Ricinus communis L

Plasma-membrane-located primary pumps were investigated in the sieve element (SE)-companion cell complex in the transport phloem of 2-week-old stems of Ricinus communis L. and, for comparison, in stems of Cucurbita pepo L. and in the secondary phloem of Agrobacterium tumefaciens-induced crown galls as a typical sink tissue. The plasma-membrane (PM) H⁺-ATPase and the tonoplast-type pyrophosphatase (PPase) were immunolocalized by epifluorescence and confocal laser scanning microscopy (CLSM) upon single or double labeling with specific monoclonal and polyclonal antibodies. Quantitative fluorescence evaluation by CLSM revealed both pumps in one membrane, the sieve-element PM. Different PM H⁺-ATPase antibody clones, raised against the PM H⁺-ATPase of Zea mays coleoptiles, induced in mouse and produced in mouse hybridoma cells, discriminated between different phloem cell types. Clones 30D5C4 and 44B8A1 labeled sieve elements and clone 46E5B11D5 labeled companion cells, indicating the existence of different phloem PM H⁺-ATPase isoforms. The results are discussed in terms of energization of SE transporters for retrieval of leaking sucrose, K⁺ and amino acids, as one of the unknown roles of ATP found in SEs. The function of the PPase could be related to phloem sucrose metabolism in support of ATP-requiring processes. Received: 3 July 2000 / Accepted: 12 October 2000     

CXCR4–SDF-1 Signalling, Locomotion, Chemotaxis and Adhesion

Chemokines, small pro-inflammatory chemoattractant cytokines, that bind to specific G-protein-coupled seven-span transmembrane receptors present on plasma membranes of target cells are the major regulators of cell trafficking. In addition some chemokines have been reported to modulate cell survival and growth. Moreover, compelling evidence is accumulating that cancer cells may employ several mechanisms involving chemokine-chemokine receptor axes during their metastasis that also regulate the trafficking of normal cells. Of all the chemokines, stromal-derived factor-1 (SDF-1), an alpha-chemokine that binds to G-protein-coupled CXCR4, plays an important and unique role in the regulation of stem/progenitor cell trafficking. First, SDF-1 regulates the trafficking of CXCR4+ haemato/lymphopoietic cells, their homing/retention in major haemato/lymphopoietic organs and accumulation of CXCR4+ immune cells in tissues affected by inflammation. Second, CXCR4 plays an essential role in the trafficking of other tissue/organ specific stem/progenitor cells expressing CXCR4 on their surface, e.g., during embryo/organogenesis and tissue/organ regeneration. Third, since CXCR4 is expressed on several tumour cells, these CXCR4 positive tumour cells may metastasize to the organs that secrete/express SDF-1 (e.g., bones, lymph nodes, lung and liver). SDF-1 exerts pleiotropic effects regulating processes essential to tumour metastasis such as locomotion of malignant cells, their chemoattraction and adhesion, as well as plays an important role in tumour vascularization. This implies that new therapeutic strategies aimed at blocking the SDF-1-CXCR4 axis could have important applications in the clinic by modulating the trafficking of haemato/lymphopoietic cells and inhibiting the metastatic behaviour of tumour cells as well. In this review, we focus on a role of the SDF-1-CXCR4 axis in regulating the metastatic behaviour of tumour cells and discuss the molecular mechanisms that are essential to this process.     

Demographic Characteristics of Female Mottled Sculpin, Cottus Bairdi, in the Coweeta Creek Drainage, North Carolina

We quantified: (1) growth rate, (2) length-mass relationships, (3) size- and age-specific fecundity, (4) egg size-frequencies, and (5) size- and age-specific egg diameter relationships for reproductively active female C. bairdi from one of the southern-most extant populations of this species (Coweeta Creek drainage, North Carolina). Gravid females were collected during February and March in 1993–1995, and 1998. Cottus bairdi reached an age of 7+ and 79mm standard length. The youngest and smallest gravid female collected was a 41mm 1+ individual. Mature 1+ females were not uncommon and we collected 21 during our study. All females older than age 2 were mature. Mean fecundity for C. bairdi at Coweeta was 71 eggs (range 9–166 eggs). We found significant positive relationships between fecundity and female length, weight and age. Female length and weight also significantly affected mean egg diameter, although the relationship was not linear. Neither female size or age significantly affected mean maximum egg diameters. Female C. bairdi from the Coweeta Creek drainage possess a unique suite of reproductive characteristics that may represent adaptations to the local selective regime or ecophenotypic variation.     

The involvement of glutamate dehydrogenase in the adaptation of mitochondria to oxidize glutamate in sucrose starved pea embryos

Embryos of pea (Pisum sativum L. cv Sol) deprived of cotyledons were cultured for 3 days in medium with or without sucrose. Respiratory activity of embryos (intact) as well as the ability to oxidize glutamate by mitochondria isolated from embryos were studied. Respiration of intact embryos grown in sucrose supplemented medium was more intensive than in the starved ones. Transfer of the starved embryos to the sucrose-containing medium induced the increase in the intensity of O₂ consumption. Mitochondria isolated from both starved and control embryos exhibited respiratory control. Mitochondria isolated from embryos cultured in the absence of sucrose showed higher (about 60 %) ability to oxidize glutamate and α-ketoglutarate than mitochondria from embryos grown in sucrose containing medium. The absence of sucrose in the medium led to a rapid increase in the specific activity of glutamate dehydrogenase (NADH-GDH and NAD-GDH) and it was accompanied by changes in izoenzymatic pattern of enzyme. These results suggest that in the conditions of sucrose starvation glutamate dehydrogenase may be responsible for the increase of glutamate oxidation by mitochondria of pea embryos. Electrophoretic separation of glutamate dehydrogenase isolated from embryos cultured in medium without sucrose showed the presence of ca. 17 isoenzymes while in non-starved embryos only 7 isoenzymes were identified. However, the addition of sucrose to starved embryos after 24 hours of cultivation led to a decrease in glutamate dehydrogenase activity (up to 40 %) but it did not cause the changes in isoenzymatic pattern. These results suggest that in the conditions of sucrose starvation glutamate dehydrogenase maybe responsible for the increase of glutamate oxidation by mitochondria of pea embryos. The posibility of glutamate dehydrogenase regulation by sucrose is discussed.     

Metabolic responses of Lemna minor to lead ions II. Induction of antioxidant enzymes in roots

Duckweed Lemna minor L. was grown on Wang culture medium supplemented with lead ions for 24 hours. Metal was tested at 1.5, 3 and 6 mg·dm⁻³ concentrations. The response of antioxidant enzymes, such as superoxide dismutase, catalase and peroxidase in lead-treated roots of duckweed was investigated. Lead ions had no effect on the spectrum of catalase and peroxidase isoenzymes while a new isoform of superoxide dismutase appeared on the Pb treated roots. A lead-depended increase in activities of superoxide dismutase and peroxidase was observed, whereas catalase activity was maintained at relatively constant values at lower lead concentrations and then decreased markedly below control level.     

Progranulin directly binds to the CRD2 and CRD3 of TNFR extracellular domains

We previously reported that PGRN directly bound to TNF receptors (TNFR) in vitro and in chondrocytes (Tang, et al., Science, 2011). Here we report that PGRN also associated with TNFR in splenocytes, and inhibited the binding of TNFα to immune cells. Proper folding of PGRN is essential for its binding to TNFR, as DTT treatment abolished its binding to TNFR. In contrast, the binding of PGRN to Sortilin was enhanced by DTT. Protein interaction assays with mutants of the TNFR extracellular domain demonstrated that CRD2 and CRD3 of TNFR are important for the interaction with PGRN, similar to the binding to TNFα. Taken together, these findings provide the molecular basis underlying PGRN/TNFR interaction and PGRN-mediated anti-inflammatory activity in various autoimmune diseases and conditions.     

The Interaction Between Selenium Status, Sex Hormones, and Thyroid Metabolism in Adolescent Girls in the Luteal Phase of their Menstrual Cycle

The objective of the present work was to study all physiological relationships among selenium status (SeS), sex hormones secretion (SH), and thyroid metabolism (ThM) in healthy adolescent girls, at one time. Forty-four girls aged 13.4–16.6 years (mean age, 14.5 ± 0.5 years) entered the statistical model. Parameters reflecting SeS: plasma selenium concentration (Se) and plasma glutathione peroxidase activity (GPX3); SH: serum estradiol (E₂) and progesterone (P₄); age of menarche (AoM); and ThM: thyroid stimulating hormone (TSH), free thyroxine (fT4), free triiodothyronine (fT3), antithyroid peroxidase antibodies (anti-TPO) in serum, and thyroid volume (ThV), were determined, and the interactions between them were evaluated by means of the partial least squares method (PLS). PLS method was, for the first time, successfully applied to the problem of selenium and hormone interactions and revealed that selenium status and female reproductive system are interrelated and affect thyroid physiology in adolescent girls in the luteal phase. The strongest associations were revealed for the pairs of parameters, Se and fT4/fT3, Se and P₄, the modest ones for the pairs, Se and ThV, P₄ and fT4/fT3, Se and AoM, and P₄ and AoM. There was no correlation between E₂, GPX3, and TSH, and any other considered parameter. Se and P₄ had the greatest influence on ThM parameters.     

The pivotal role of glutamate dehydrogenase (GDH) in the mobilization of N and C from storage material to asparagine in germinating seeds of yellow lupine

In germinating seeds of legumes, amino acids liberated during mobilization of storage proteins are partially used for synthesis of storage proteins of the developing axis, but some of them are respired. The amino acids are catabolized by both glutamate dehydrogenase (GDH) and transaminases. Ammonium is reassimilated by glutamine synthetase (GS) and, through the action of asparagine synthetase (AS), is stored in asparagine (Asn).This review presents the ways in which amino acids are converted into Asn and their regulation, mostly in germinating seeds of yellow lupine, where Asn can make up to 30% of dry matter. The energy balance of the synthesis of Asn from glutamate, the most common amino acid in lupine storage proteins, also shows an adaptation of lupine for oxidation of amino acids in early stages of germination.Regulation of the pathway of Asn synthesis is described with regard to the role of GDH and AS, as well as compartmentation of particular metabolites. The regulatory effect of sugar on major links of the pathway (mobilization of storage proteins, induction of genes and activity of GDH and AS) is discussed with respect to recent genetic and molecular studies. Moreover, the effect of glutamate and phytohormones is presented at various stages of Asn biosynthesis.