Cytochrome c: Surfing Off of the Mitochondrial Membrane on the Tops of Complexes III and IV

The proper arrangement of protein components within the respiratory electron transport chain is nowadays a matter of intense debate, since altering it leads to cell aging and other related pathologies. Here, we discuss three current views-the so-called solid, fluid and plasticity models-which describe the organization of the main membrane-embedded mitochondrial protein complexes and the key elements that regulate and/or facilitate supercomplex assembly. The soluble electron carrier cytochrome c has recently emerged as an essential factor in the assembly and function of respiratory supercomplexes. In fact, a 'restricted diffusion pathway' mechanism for electron transfer between complexes III and IV has been proposed based on the secondary, distal binding sites for cytochrome c at its two membrane partners recently discovered. This channeling pathway facilitates the surfing of cytochrome c on both respiratory complexes, thereby tuning the efficiency of oxidative phosphorylation and diminishing the production of reactive oxygen species. The well-documented post-translational modifications of cytochrome c could further contribute to the rapid adjustment of electron flow in response to changing cellular conditions.Spanish Ministry of Economy and Competitiveness (BFU2015-71017/BMC MINECO/FEDER and PGC2018-096049-B-I00 BIO/BMC MICINN/FEDER, EU

Long distance electron transfer through the aqueous solution between redox partner proteins

Despite the importance of electron transfer between redox proteins in photosynthesis and respiration, the inter-protein electron transfer rate between redox partner proteins has never been measured as a function of their separation in aqueous solution. Here, we use electrochemical tunneling spectroscopy to show that the current between two protein partners decays along more than 10 nm in the solution. Molecular dynamics simulations reveal a reduced ionic density and extended electric field in the volume confined between the proteins. The distance-decay factor and the calculated local barrier for electron transfer are regulated by the electrochemical potential applied to the proteins. Redox partners could use electrochemically gated, long distance electron transfer through the solution in order to conciliate high specificity with weak binding, thus keeping high turnover rates in the crowded environment of cells.España, MINECO BFU2015-71017-P/España, Gobierno de Andalucía (BIO198)Comisión de Universidades e Investigación del Departamento de Innovación, Universidades y Empresa de la Generalitat de Catalunya.(2017 SGR 1442, 2014 SGR 1442, and 2017 SGR 1189

La negociación, piedra angular de las investigaciones

The first section of this paper focuses onthe importance of negotiation processesrelated to research and evaluation. Thisimportance is based on five issues: ethicsin production processes, knowledge scope,rigorous inquiry, efficiency of development-oriented findings, and strategy tomaintain contexts in which research andparticipant learning can take place. Thedifferent phases of the negotiation processare also presented: initial, processing andreporting, the last one of special importance.The requirements, types, characteristicsand operation processes of the reportingphase are discussed. The secondsection of this paper provides in the formof an example a specific case of negotiationfrom a qualitative investigation aboutimmigrant families’ participation at school.Its different phases, demands and virtualitiesare discussed. The paper finished withsome final considerations and references.El artículo consta de dos partes complementarias. Una de ellas trata de explicar la importancia de los procesos de negociación en la realización de investigaciones y evaluaciones. Importancia que se basa en cinco razones: ética de los procesos de producción y aplicación del conocimiento,rigor de las indagaciones, eficacia de los hallazgos para la mejora, estrategia para mantener contextos en que se pueda investigar y aprendizaje de los y las participantes. Se plantean las distintas fases de la negociación: inicial, de proceso y de informes, con especial atención a estaúltima. De ella se estudian las exigencias,los tipos, las características y los procedimientos de actuación.La segunda parte muestra, a modo de ejemplo, un caso concreto de negociación de una investigación cualitativa sobre la participación de familias inmigrantes en la escuela, con sus diversas fases, exigencias y virtualidades.Se cierra el artículo con algunas consideraciones finales y con la correspondiente bibliografía

La participación de las familias de alumnos y alumnas inmigrantes en las instituciones educativas

Los fenómenos de la inmigración y de la interculturalidad han cobrado en España un extraordinario protagonismo no sólo por sus dimensiones sino por la variedad de la procedencia de inmigrantes y la complejidad de las situaciones migratorias. El trabajo del grupo consistía en explorar cuál es la respuesta de la escuela a esta situación sobrevenida de forma casi súbita y cuál es la dinámica de participación de las familias en ella.En este artículo se expone el proceso de una investigación etnográfica realizada en seis centros escolares de Málaga (España) durante los años 2007/2009 sobre la participación de las familias de alumnos y alumnas inmigrantes en la escuela. La investigación se negocia con los centros elegidos (4 de Primaria y 2 de Secundaria) en los que se realizan entrevistas, observaciones, grupos de discusión y se analizan documentos relacionados con la temática del trabajo. Se desarrollan los principales resultados, tanto relativos al proceso como al foco de estudio, destacando las buenas prácticas que se están llevando a cabo en los centros y que pueden servir de referencia en otros contextos escolares.Los informes de la investigación han sido devueltos a los centros para su negociación con los participantes. Aquí se plantean las conclusiones de carácter general que se han extraído de los seis centros en los que se realizó la exploración. Hay por consiguiente, cuestiones que se refieren a todos y otras que son peculiares de uno o algunos de ellos

Structural and functional characterization of phosphomimetic mutants of cytochrome c at threonine 28 and serine 47

Protein function is frequently modulated by post-translational modifications of specific residues. Cytochrome c, in particular, is phosphorylated in vivo at threonine 28 and serine 47. However, the effect of such modifications on the physiological functions of cytochrome c – namely, the transfer of electrons in the respiratory electron transport chain and the triggering of programmed cell death – is still unknown. Here we replace each of these two residues by aspartate, in order to mimic phosphorylation, and report the structural and functional changes in the resulting cytochrome c variants. We find that the T28D mutant causes a 30-mV decrease on the midpoint redox potential and lowers the affinity for the distal site of Arabidopsis thaliana cytochrome c1 in complex III. Both the T28D and S47D variants display a higher efficiency as electron donors for the cytochrome c oxidase activity of complex IV. In both protein mutants, the peroxidase activity is significantly higher, which is related to the ability of cytochrome c to leave the mitochondria and reach the cytoplasm. We also find that both mutations at serine 47 (S47D and S47A) impair the ability of cytoplasmic cytochrome c to activate the caspases cascade, which is essential for triggering programmed cell death.Peer reviewe

Oxidative stress is tightly regulated by cytochrome c phosphorylation and respirasome factors in mitochondria

Respiratory cytochrome c has been found to be phosphorylated at tyrosine 97 in the postischemic brain upon neuroprotective insulin treatment, but how such posttranslational modification affects mitochondrial metabolism is unclear. Here, we report the structural features and functional behavior of a phosphomimetic cytochrome c mutant, which was generated by site-specific incorporation at position 97 of p-carboxymethyl-l-phenylalanine using the evolved tRNA synthetase method. We found that the point mutation does not alter the overall folding and heme environment of cytochrome c, but significantly affects the entire oxidative phosphorylation process. In fact, the electron donation rate of the mutant heme protein to cytochrome c oxidase, or complex IV, within respiratory supercomplexes was higher than that of the wild-type species, in agreement with the observed decrease in reactive oxygen species production. Direct contact of cytochrome c with the respiratory supercomplex factor HIGD1A (hypoxia-inducible domain family member 1A) is reported here, with the mutant heme protein exhibiting a lower affinity than the wild-type species. Interestingly, phosphomimetic cytochrome c also exhibited a lower caspase-3 activation activity. Altogether, these findings yield a better understanding of the molecular basis for mitochondrial metabolism in acute diseases, such as brain ischemia, and thus could allow the use of phosphomimetic cytochrome c as a neuroprotector with therapeutic applications.España, Junta de Andalucía BIO-198España MINECO BFU2015-71017/BM

Glucose 6-P Dehydrogenase Overexpression Improves Aging-Induced Endothelial Dysfunction in Aorta from Mice: Role of Arginase II

The increase of vascular arginase activity during aging causes endothelial dysfunction. This enzyme competes with the endothelial nitric oxide synthase (eNOS) for L-arginine substrate. Our hypothesis is that glucose 6-P dehydrogenase (G6PD) overexpression could improve the endothelial function modulating the arginase pathway in aorta from mice. For this study, three groups of male mice were used: young wild type (WT) (6-9 months), old WT (21-22 months) and old G6PD-Tg (21-22 months) mice. Vascular reactivity results showed a reduced acetylcholine-dependent relaxation in the old WT but not old G6PD-Tg group. Endothelial dysfunction was reverted by nor-NOHA, an arginase inhibitor. Mice overexpressing G6PD underexpressed arginase II and also displayed a lower activity of this enzyme. Moreover, histological analyses demonstrated that age causes a thickness of aortic walls, but this did not occur in G6PD-Tg mice. We conclude that the overexpressing G6PD mouse is a model to improve vascular health via the arginase pathway

Grape pomace in ewes diet: effects on meat quality and the fatty acid profile of their suckling lambs

Producción CientíficaGrape pomace (GP) is an inexpensive natural antioxidant promising as animal feed supplement due to its high content of phenolic compounds. In order to evaluate its effect in lactating ewe rations on meat quality and fat composition of their suckling lambs, 48 Churra ewes were divided into 4 treatments. All animals were fed a ration containing linseed oil (Control) supplemented with Vitamin E or two levels of GP. Lambs were nourished exclusively by suckling until they were slaughtered. Dietary GP did not generate adverse effects on carcasses or lambs meat quality when compared with Control or Vit-E diets. GP improved the water holding capacity of the meat. In addition, lambs meat FA profile was not nutritionally affected with the diets assayed. Hence, the use of GP as a dietary supplement in ewe rations would not have negative effects on meat from suckling lambs.INIA (RTA2010-0068-C02)Consejería de Educación de la Junta de Castilla y León (VA196A11-2

The role of PKD1 in aging-related neurodegeneration: Structural and functional imaging studies

Trabajo presentado en el Second Spanish Molecular Imaging Network (SMIN) Meeting, celebrado en Madrid (España) el 26 de febrero de 2018

Respiratory complexes III and IV can each bind two molecules of cytochrome c at low ionic strength

The transient interactions of respiratory cytochrome c with complexes III and IV is herein investigated by using heterologous proteins, namely human cytochrome c, the soluble domain of plant cytochrome c1 and bovine cytochrome c oxidase. The binding molecular mechanisms of the resulting cross-complexes have been analyzed by Nuclear Magnetic Resonance and Isothermal Titration Calorimetry. Our data reveal that the two cytochrome c-involving adducts possess a 2:1 stoichiometry – that is, two cytochrome c molecules per adduct – at low ionic strength. We conclude that such extra binding sites at the surfaces of complexes III and IV can facilitate the turnover and sliding of cytochrome c molecules and, therefore, the electron transfer within respiratory supercomplexes.España, MINECO Grant Nos. BFU2010-19451/BMC and BFU2012-31670/BMCJunta de Andalucía Grant PAI, BIO198España Ministerio de Educación, y European Social Fund-ERDF AP2009-409