High-trhoughput screening of cytochrome C novel partners chlamydomonas ReindhartII

Comunicaciones a congreso

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

Molecular characterization of autophagic and apoptotic signaling induced by sorafenib in liver cancer cells

Sorafenib is the unique accepted molecular targeted drug for the treatment of patients in advanced stage of hepatocellular carcinoma. The current study evaluated cell signaling regulation of endoplasmic reticulum (ER) stress, c-Jun-N-terminal kinase (JNK), Akt, and 5′AMP-activated protein kinase (AMPK) leading to autophagy and apoptosis induced by sorafenib. Sorafenib induced early (3–12 hr) ER stress characterized by an increase of Ser51P-eIF2α/eIF2α, C/EBP homologous protein (CHOP), IRE1α, and sXBP1, but a decrease of activating transcription factor 6 expression, overall temporally associated with the increase of Thr183,Tyr185P-JNK1/2/JNK1/2, Thr172P-AMPKα, Ser413P-Foxo3a, Thr308P-AKt/AKt and Thr32P-Foxo3a/Foxo3a ratios, and reduction of Ser2481P-mammalian target of rapamycin (mTOR)/mTOR and protein translation. This pattern was related to a transient increase of tBid, Bim EL, Beclin-1, Bcl-xL, Bcl-2, autophagy markers, and reduction of myeloid cell leukemia-1 (Mcl-1) expression. The progressive increase of CHOP expression, and reduction of Thr308P-AKt/AKt and Ser473P-AKt/AKt ratios were associated with the reduction of autophagic flux and an additional upregulation of Bim EL expression and caspase-3 activity (24 hr). Small interfering-RNA (si-RNA) assays showed that Bim, but not Bak and Bax, was involved in the induction of caspase-3 in sorafenib-treated HepG2 cells. Sorafenib increased autophagic and apoptotic markers in tumor-derived xenograft model. In conclusion, the early sorafenib-induced ER stress and regulation of JNK and AMPK-dependent signaling were related to the induction of survival autophagic process. The sustained drug treatment induced a progressive increase of ER stress and PERK-CHOP-dependent rise of Bim EL, which was associated with the shift from autophagy to apoptosis. The kinetic of Bim EL expression profile might also be related to the tight balance between AKt- and AMPK-related signaling leading to Foxo3a-dependent BIM EL upregulation.Ministerio de Economía y Competitividad BFU2016‐75352‐PInstituto de Salud Carlos III PI15/00034, PI13/ 00021, PI16/00090, PI14/01349Ministerio de Educación FPU16/05127, FPU12/01433, FPU13/01237Junta de Andalucía CTS-6264, PI-00025-2013, PI-0127-2013, PI-0198-201

Histone chaperone activity of Arabidopsis thaliana NRP1 is blocked by cytochrome c

Higher-order plants and mammals use similar mechanisms to repair and tolerate oxidative DNA damage. Most studies on the DNA repair process have focused on yeast and mammals, in which histone chaperone-mediated nucleosome disassembly/reassembly is essential for DNA to be accessible to repair machinery. However, little is known about the specific role and modulation of histone chaperones in the context of DNA damage in plants. Here, the histone chaperone NRP1, which is closely related to human SET/TAF-I, was found to exhibit nucleosome assembly activity in vitro and to accumulate in the chromatin of Arabidopsis thaliana after DNA breaks. In addition, this work establishes that NRP1 binds to cytochrome c, thereby preventing the former from binding to histones. Since NRP1 interacts with cytochrome c at its earmuff domain, that is, its histone-binding domain, cytochrome c thus competes with core histones and hampers the activity of NRP1 as a histone chaperone. Altogether, the results obtained indicate that the underlying molecular mechanisms in nucleosome disassembly/reassembly are highly conserved throughout evolution, as inferred from the similar inhibition of plant NRP1 and human SET/TAF-I by cytochrome c during DNA damage response

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

Petrologic and geochemical constraints on the origin of Astaneh pluton, Zagros orogenic belt, Iran

The Astaneh plutonic complex consists of a series of granitoid rocks ranging in composition from quartzdiorites to monzogranites and evolving from metaluminous to weakly peraluminous compositions. They belong to the high-K calc-alkaline series, having features of typical Andean-type cordilleran granitoids. Trace and rare-earth elements distribution patterns for the Astaneh rocks indicate a distinctive depletion in Nb, Sr, Ba, P and Ti relative to other trace elements and a greater enrichment in LILE compared to HFSE. These geochemical characteristics suggest the participation of an important recycled (sedimentary?) component in the source region of the granitoids. They have Sr initial isotopic ratios in the range 0.7078–0.7084 and negative eNd values of 5.39 to 6.13 for a time of generation of 170 Ma. There is a genetic link between quartz-diorites and granodiorites, the dominant rock types of the Astabeh intrusion. Direct melting or fractionation from a diorite source is very unlike. It is proposed that the Astaneh parental Qtd-diorite magmas were produced by the partial melting of a mixed source, dominantly composed of amphibolites and sediments, that was formed during subduction of Neo-Tethyan oceanic crust below the Iranian microcontinent during Middle Jurassic times

Source apportionment of ultrafine and fine particles in Huelva industrial city

Comunicación presentada en: 2012 European Aerosol Conference (EAC-2012), B-WG01S2P30, celebrada del 2 al 7 de septiembre de 2012 en Granada

Reinforced silica-carbon nanotube monolithic aerogels synthesised by rapid controlled gelation

This work introduces a new synthesis procedure for obtaining homogeneous silica hybrid aerogels with carbon nanotube contents up to 2.50 wt.%. The inclusion of nanotubes in the highly porous silica matrix was performed by a two-step sol–gel process, resulting in samples with densities below 80 mg/cm3. The structural analyses (N2 physisorption and SEM) revealed the hierarchical structure of the porous matrix formed by nanoparticles arranged in clusters of 100 and 300 nm in size, specific surface areas around 600 m2/g and porous volumes above 4.0 cm3/g. In addition, a relevant increase on the mechanical performance was found, and an increment of 50% for the compressive strength and 90% for the maximum deformation were measured by uniaxial compression. This reinforcement was possible thanks to the outstanding dispersion of the CNT within the silica matrix and the formation of Si–O–C bridges between nanotubes and silica matrix, as suggested by FTIR. Therefore, the original synthesis procedure introduced in this work allows the fabrication of highly porous hybrid materials loaded with carbon nanotubes homogeneously distributed in the space, which remain available for a variety of technological applications

Influence of Saharan dust in deposition fluxes of nutrients in Spain

Comunicación presentada en: 2012 European Aerosol Conference (EAC-2012), B-WG01S2P30, celebrada del 2 al 7 de septiembre de 2012 en Granada

The cytochrome f–plastocyanin complex as a model to study transient interactions between redox proteins

AbstractTransient complexes, with a lifetime ranging between microseconds and seconds, are essential for biochemical reactions requiring a fast turnover. That is the case of the interactions between proteins engaged in electron transfer reactions, which are involved in relevant physiological processes such as respiration and photosynthesis. In the latter, the copper protein plastocyanin acts as a soluble carrier transferring electrons between the two membrane-embedded complexes cytochrome b6f and photosystem I. Here we review the combination of experimental efforts in the literature to unveil the functional and structural features of the complex between cytochrome f and plastocyanin, which have widely been used as a suitable model for analyzing transient redox interactions