Ouabain-insensitive Na+-stimulated ATPase activity of basolateral plasma membranes from guinea-pig kidney cortex cells: II. Effect of Ca2+

The ouabain-insensitive, Mg²⁺-dependent, Na⁺-stimulated ATPase activity present in fresh basolateral plasma membranes from guinea-pig kidney cortex cells (prepared at pH 7.2) can be increased by the addition of micromolar concentrations of Ca²⁺ to the assay medium. The Ca²⁺ involved in this effect seems to be associated with the membranes in two different ways: as a labile component, which can be quickly and easily ‘deactivated’ by reducing the free Ca²⁺ concentration of the assay medium to values lower than 1 μM; and as a stable component, which can be ‘deactivated’ by preincubating the membranes for periods of 3–4 h with 2 mM EDTA or EGTA. Both components are easily activated by micromolar concentrations of Ca²⁺. The $\text{K}{}_{\mathrm{<mtext>a</mtext>}}$ of the system for Na⁺ is the same, 8 mM, whether only the stable component or both components, stable and labile, are working. In other words, the activating effect of Ca²⁺ on the Na⁺-stimulated ATPase is on the $\text{V}{}_{\mathrm{<mtext>max</mtext>}}$, and not on the $\text{K}{}_{\mathrm{<mtext>a</mtext>}}$ of the system for Na⁺. The activating effect of Ca²⁺ may be related to some conformational change produced by the interaction of this ion with the membranes, since it can also be obtained by resuspending the membranes at pH 7.8 or by ageing the preparations. Changes in the Ca²⁺ concentration may modulate the ouabain-insensitive, Na⁺-stimulated ATPase activity. This modulation could regulate the magnitude of the extrusion of Na⁺ accompanied by Cl^? and water that these cells show, and to which the Na⁺-ATPase has been associated as being responsible for the energy supply of this mode of Na⁺ extrusion.     

Translational regulation of Arabidopsis XIPOTL1 is modulated by phosphocholine levels via the phylogenetically conserved upstream open reading frame 30

In Arabidopsis thaliana, XIPOTL1 encodes a phosphoethanolamine N-methyltransferase with a central role in phosphatidylcholine biosynthesis via the methylation pathway. To gain further insights into the mechanisms that regulate XIPOTL1 expression, the effect of upstream open reading frame 30 (uORF30) on the translation of the major ORF (mORF) in the presence or absence of endogenous choline (Cho) or phosphocholine (PCho) was analysed in Arabidopsis seedlings. Dose-response assays with Cho or PCho revealed that both metabolites at physiological concentrations are able to induce the translational repression of a mORF located downstream of the intact uORF30, without significantly altering its mRNA levels. PCho profiles showed a correlation between increased endogenous PCho levels and translation efficiency of a uORF30-containing mORF, while no correlation was detectable with Cho levels. Enhanced expression of a uORF30-containing mORF and decreased PCho levels were observed in the xipotl1 mutant background relative to wild type, suggesting that PCho is the true mediator of uORF30-driven translational repression. In Arabidopsis, endogenous PCho content increases during plant development and affects root meristem size, cell division, and cell elongation. Because XIPOTL1 is preferentially expressed in Arabidopsis root tips, higher PCho levels are found in roots than shoots, and there is a higher sensitivity of this tissue to translational uORF30-mediated control, it is proposed that root tips are the main site for PCho biosynthesis in Arabidopsis.     

Electrophoretic Separation of Different Phophosproteins Associated with Ca-ATPase and Na,K-ATPase in Human Red Cell Ghosts

Ca has been found to increase the quantity of ³²P incorporated into red cell ghosts from [γ-³²P]ATP over the levels obtained by incubation with Mg alone or with Mg + Na, in correlation with the effect of Ca on the associated ATPase activities. When the ³²P-labeled ghosts were solubilized in sodium dodecyl sulfate (SDS) and electrophoresed on acrylamide gels only two bands could be detected either by autoradiography or by counting the sliced gels. The faster moving band (P-2) had the same mobility and the same molecular weight (103,000) as the phosphoprotein found either with Mg alone or with Mg + Na. The slower moving band (P-1) was not found in extensively washed ghosts labeled in the absence of Ca. The molecular weight of P-1 is approximately 150,000. P-1 like P-2 was not affected by pretreatment of intact cells with Pronase before labeling indicating that neither the phosphorylating mechanism nor the phosphoprotein are accessible to externally applied Pronase. The demonstration that a Ca-phosphoprotein is separable from the Na-stimulated phosphoprotein suggests that the Ca-ATPase is distinct from and independent of the Na,K-ATPase. The fact that Ca blocks the dephosphorylation by K of the Na-phosphoprotein indicates that caution is required in interpreting results when the activities of the different phosphoproteins have not been separately determined.     

Chemical Characterization and Pronase Susceptibility of the Na:K Pump-Associated Phosphoprotein of Human Red Blood Cells

The phosphoproteins formed by incubation of red cell ghosts with [γ-³²P]ATP in the presence of Mg and Na + Mg have been characterized by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. The ³²P-labeled phosphoprotein was seen as a single peak confined to the region of the diffuse 90,000 dalton polypeptide band; labeling with Na + Mg considerably increased the quantity of ³²P-phosphoprotein contained in this band relative to labeling with Mg alone. Treatment of intact cells with Pronase known to partially hydrolyze the glycoproteins and the 90,000 daltons polypeptide did not change either the amount or the position of the ³²P-phosphoprotein present in the gels. The molecular weight of the ³²P-phosphoprotein is estimated to be 103,000. Pronase treatment of intact cells also did not significantly alter any of the transport parameters of the membrane such as the K pump flux, ouabain binding, or Na,K-ATPase. In contrast, treatment of ghosts with Pronase not only resulted in drastic alteration of the transport parameters but also inhibited the formation of the phosphoprotein under all conditions. Thus, while the Na:K pump is not intrinsically resistant to Pronase, those elements of the pump which are susceptible are not accessible from the outside of the cell. Further, SDS-polyacrylamide gel electrophoresis after Pronase treatment of intact cells results in a substantial increase in the purification of the phosphoprotein relative to that which was previously possible in ghosts.     

Low effectiveness of the Natura 2000 network in preventing land-use change in bat hotspots

One of the most important issues in biodiversity conservation is an exploration of the relationships among protected areas, land-use changes and biodiversity, so we aimed to assess the performance of the Natura 2000 network (N2000) in representing the bat conservation hotspots in peninsular Spain and the Balearic Islands and to compare the rates of land-use changes within these hotspots with those observed throughout the rest of the study area. First, we applied a Combined Index that integrates various biodiversity metrics (species richness, rarity and vulnerability) to identify hotspots, and once they were identified, we used null models to assess the performance of N2000 in representing them. Finally, also using null models, we tested whether the changes in land use (“anthropization” or “naturalization”) within the hotspots occurred at a significantly higher or lower frequency than in the rest of the study area; for this, we considered two temporal windows (1980–2006 and 2006–2012) corresponding with periods before and after the official designation of the N2000 sites. Our results show that bat hotspots are effectively represented in the Iberian N2000, but although land-use changes were generally higher in Spain before 2006, hotspots have not experienced lower rates of change compared to the remainder of the territory (regardless of the period under consideration). This suggests low effectiveness of the Iberian N2000 in preventing land-use change in bat hotspots, so to preserve the Iberian bat fauna, we encourage the urgent implementation of management plans to avoid intensive changes in land use both inside and around bat hotspots.     

Na+-stimulated ATPase activities in kidney basal-lateral plasma membranes

In the present work we demonstrate the existence of a Na⁺-stimulateo, Mg²⁺-dependent ATPase activity, which is insensitive to ouabain, and which is 100% inhibited by 1.5 mM ethacrynic acid in freshly prepared, basal-lateral enriched, plasma-membrane fractions obtained from guinea-pig kidney cortex slices (which are mainly made up by proximal tubules). This ATPase activity has characteristics similar to those described previously in a microsomal fraction of the guinea-pig kidney cortex by a procedure which required ageing of the preparation for more than two weeks (Proverbio, F., Condrescu-Guidi, M. and Whittembury, G. (1975) Biochim. Biophys. Acta 394, 281–292), it does not seem to be due to some partially modified activity of the Mg²⁺-ATPase, the (Na⁺ + K⁺)-or the Ca²⁺-ATPase activities present in this tissue. It seems to be due to the activity of another system, which is located on the basal-lateral membrane of the kidney tubular cells.     

Sexual size dimorphism in greater mouse‐eared bat Myotis myotis (Chiroptera: Vespertilionidae) from a Mediterranean region

Although sexual size dimorphism (SSD) is common among mammals, there is no clear explanation for its maintenance in nature. Bats are one of the few groups of mammals where reverse SSD appears. In this group, the size of individuals may have very important ecological consequences related with flight. In this study, we examine sexual dimorphism in the wing measurements of 195 adult individuals (141 males and 54 females) of the greater mouse‐eared bat Myotis myotis in the south‐east of the Iberian Peninsula. We also investigated size differences between paired and single males in a swarming roost. The results indicate that there are significant differences in the wing measurements between sexes, females being bigger than males in this respect. While no significant differences in the wing measurements of paired and single males were observed, significant differences were found in their relative weight and fitness, single males being significantly heavier and having a better physical condition. We discuss the implications of SSD for the female of M. myotis in terms of reproductive advantages, trophic niche segregation and a greater ability to move, which may favour gene flow between populations.