RESUMO
Pollen tubes require functional mitochondria in order to achieve fast and sustained growth. In addition, cell wall expansion requires a calcium gradient in the tube apex formed by a dedicated array of calcium pumps and channels. Most studies have traditionally focused on the molecular aspects of calcium interactions and transport across the pollen tube plasmalemma. However, calcium transients across mitochondrial membranes from pollen tubes are beginning to be studied. Here, we report the presence of a ruthenium red-sensitive mitochondrial calcium uniporter-like activity in tobacco pollen tubes with functional oxidative phosphorylation. The present study provides a framework to measure in situ specifics of mitochondrial transport and respiration in pollen tubes from different plants. The relevance of a mitochondrial calcium uniporter for pollen tube growth is discussed.
Assuntos
Canais de Cálcio/metabolismo , Nicotiana/química , Tubo Polínico/químicaRESUMO
Trypanosoma evansi, which causes surra, is descended from Trypanosoma brucei brucei, which causes nagana. Although both parasites are presumed to be metabolically similar, insufficient knowledge of T. evansi precludes a full comparison. Herein, we provide the first report on the subcellular localisation of the glycolytic enzymes in T. evansi, which is a alike to that of the bloodstream form (BSF) of T. b. brucei: (i) fructose-bisphosphate aldolase, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), hexokinase, phosphofructokinase, glucose-6-phosphate isomerase, phosphoglycerate kinase, triosephosphate isomerase (glycolytic enzymes) and glycerol-3-phosphate dehydrogenase (a glycolysis-auxiliary enzyme) in glycosomes, (ii) enolase, phosphoglycerate mutase, pyruvate kinase (glycolytic enzymes) and a GAPDH isoenzyme in the cytosol, (iii) malate dehydrogenase in cytosol and (iv) glucose-6-phosphate dehydrogenase in both glycosomes and the cytosol. Specific enzymatic activities also suggest that T. evansi is alike to the BSF of T. b. brucei in glycolytic flux, which is much faster than the pentose phosphate pathway flux, and in the involvement of cytosolic GAPDH in the NAD+/NADH balance. These similarities were expected based on the close phylogenetic relationship of both parasites.
Assuntos
Animais , Ratos , Glicólise , Trypanosoma/enzimologia , Tripanossomíase/parasitologia , Modelos Animais de Doenças , Filogenia , Ratos Sprague-Dawley , Especificidade da Espécie , Trypanosoma/classificação , Trypanosoma/genética , UltracentrifugaçãoRESUMO
In Trypanosoma cruzi three isoenzymes of phosphoglycerate kinase (PGK) are found which are simultaneously expressed: the cytosolic isoenzyme PGKB as well as two glycosomal enzymes, PGKA and PGKC. In this paper, we show that PGKA in T. cruzi epimastigotes is associated to the glycosomal membrane; it is responsible for about 23% of the glycosomal PGK activity, the fraction that remains in the pellet after osmotic shock treatment of purified organelles, in contrast to the 77% soluble activity that is mainly attributed to PGKC. Antibodies against the unique 80 amino-acid insertion of PGKA blocked almost completely the glucose consumption by epimastigotes that were partially permeabilized with digitonin. These results indicate that PGKA is the predominant isoenzyme for sustaining glycolysis through the glycosomes of these parasites.
Assuntos
Glucose/metabolismo , Membranas Intracelulares/enzimologia , Microcorpos/enzimologia , Fosfoglicerato Quinase/metabolismo , Proteínas de Protozoários/metabolismo , Trypanosoma cruzi/enzimologia , Transporte Biológico , Doença de Chagas/parasitologia , Citosol/enzimologia , Glicólise , Humanos , Membranas Intracelulares/metabolismo , Isoenzimas/genética , Isoenzimas/metabolismo , Fosfoglicerato Quinase/genética , Proteínas de Protozoários/genética , Trypanosoma cruzi/genética , Trypanosoma cruzi/crescimento & desenvolvimento , Trypanosoma cruzi/metabolismoRESUMO
The branched respiratory chain in mitochondria from the halotolerant yeast Debaryomyces hansenii contains the classical complexes I, II, III and IV plus a cyanide-insensitive, AMP-activated, alternative-oxidase (AOX). Two additional alternative oxidoreductases were found in this organism: an alternative NADH dehydrogenase (NDH2e) and a mitochondrial isoform of glycerol-phosphate dehydrogenase (MitGPDH). These monomeric enzymes lack proton pump activity. They are located on the outer face of the inner mitochondrial membrane. NDH2e oxidizes exogenous NADH in a rotenone-insensitive, flavone-sensitive, process. AOX seems to be constitutive; nonetheless, most electrons are transferred to the cytochromic pathway. Respiratory supercomplexes containing complexes I, III and IV in different stoichiometries were detected. Dimeric complex V was also detected. In-gel activity of NADH dehydrogenase, mass spectrometry, and cytochrome c oxidase and ATPase activities led to determine the composition of the putative supercomplexes. Molecular weights were estimated by comparison with those from the yeast Y. lipolytica and they were IV2, I-IV, III2-IV4, V2, I-III2, I-III2-IV, I-III2-IV2, I-III2-IV3 and I-III2-IV4. Binding of the alternative enzymes to supercomplexes was not detected. This is the first report on the structure and organization of the mitochondrial respiratory chain from D. hansenii.