Evolutionary history of Na,K-ATPases and their osmoregulatory role

The Na/K pump, or Na,K-ATPase, is a key enzyme to the homeostasis of osmotic pressure, cell volume, and the maintenance of electrochemical gradients. Its α subunit, which holds most of its functions, belongs to a large family of ATPases known as P-type, and to the subfamily IIC, which also includes H,K-ATPases. In this study, we attempt to describe the evolutionary history of IIC ATPases by doing phylogenetic analysis with most of the currently available protein sequences (over 200), and pay special attention to the relationship between their diversity and their osmoregulatory role. We include proteins derived from many completed or ongoing genome projects, many of whose IIC ATPases have not been phylogenetically analyzed previously. We show that the most likely origin of IIC proteins is prokaryotic, and that many of them are present in non-metazoans, such as algae, protozoans or fungi. We also suggest that the pre-metazoan ancestor, represented by the choanoflagellate Monosiga brevicollis, whose genome has been sequenced, presented at least two IIC-type proteins. One of these proteins would have given rise to most current animal IIC ATPases, whereas the other apparently evolved into a lineage that, so far, has only been found in nematodes. We also propose that early deuterostomes presented a single IIC gene, from which all the extant diversity of vertebrate IIC proteins originated by gene and genome duplications. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Mitochondrial ATPase inactivation by interaction with its substrate

Purified F₁-ATPase is slowly inactivated by interaction, in a preincubation medium, with its substrate MgATP. Interaction with Mg²⁺ before addition of ATP to the preincubation medium is essential to induce the inactivation. This inactivation is different from other Mg²⁺-induced inhibitions previously described. Free ATP concentration is implicated in the inactivation process and a linear relationship can be established between this concentration and the number of turnovers which are necessary for total inactivation. ITP, 2′-dATP, and GTP can also induce inactivation. Although ITP and GTP are hydrolyzed at a lower rate than ATP and 2′-dATP, they induce inactivation after a smaller number of turnovers than the latter. This process closely follows a kinetics of the type described for suicide enzymes. A reaction scheme is suggested and discussed.     

Histochemistry and histoenzymology of the hydatid cyst (Echinococcus granulosus Batsch, 1786). I. The germinal membrane.

Healthy germinal membranes of hydatid cysts from lungs of human and bovine sources were dissected and isolated for histochemical and histoenzymatic research. These techniques were performed in frozen sections and pieces of the whole membrane. Enzymatic research showed that the germinal membrane presents highly differentiated metabolic areas. These areas were topographically related with the origin and insertion of brood capsules, having differentiated structures for metabolic interchange with scolices. Taking our data into account it may be suggested that this functional differentiation could be transitory and variable for all the membrane surface. The accumulation of lipids and enzymes such as simple estarase, lipase, beta-HDH, alpha-GDH and NADPH-reductase in those areas, suggests that lipids are not a simple excretory product. This distribution probably implies that lipid metabolism or its resultant products are important in development and growth of scolices. In that sense other authors' findings and hypothesis about the possible existence of an endocrine system of the parasite, are considered. This idea being demonstrated in further researches, the lipid metabolic pathways shall bring a good pharmacological approach to the interference with parasite development.     

Partial characterization of IR-alpha-MSH peptides found in melanoma tumors.

Our previous work indicated that IR-alpha-MSH (immunoreactive alpha-melanocyte-stimulating hormone) plasma levels are three times as high in melanoma patients with progressing disease than in disease-free patients, and that the melanoma tumor itself may be the source of IR-alpha-MSH. Further identification of the material in tumor extracts has been carried out in this study, and the results presented here show that the immunoreactivity is associated with a major fraction of about 16 kDa and another of 5-9 kDa. Significant amounts of the immunoreactive material were also found in human melanoma cells but not in culture supernatants. The presence of this material may be related to the melanogenic status of the tumor cells. We have estimated the intracellular IR-alpha-MSH to be within a 0.4 to 2.3 nM range in melanoma tumor cells. We have investigated the melanogenic effect of the IR-alpha-MSH material and its relationship to alpha-MSH. Purified extracts both from metastases and cultured cells were found to promote frog skin darkening as well as tyrosinase activity in Cloudman S91 melanoma cells. The IR material could also displace labeled alpha-MSH from its binding sites in human melanoma cells. Our data clearly indicate that melanoma cells engage in an autocrine production of alpha-MSH-like bioactive peptides by melanoma cells, of larger mol.wt., which are able to bind to MSH receptors. These peptides may be involved in the regulation of melanogenesis and possibly in the growth and proliferation of melanoma cells by an autocrine/paracrine mechanism.