Laura Vela

University of Zaragoza, Zaragoza, Aragon, Spain

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Publications (7)23.54 Total impact

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    ABSTRACT: The key event in the mitochondrial pathway of apoptosis is the activation of Bax and Bak by BH3-only proteins, through a molecular mechanism that is still a matter of debate. We have studied here interactions among anti- and pro-apoptotic proteins of the Bcl-2 family in living cells by using Bimolecular Fluorescence Complementation (BiFC) analysis. Our results indicate that the antiapoptotic proteins Mcl-1 and Bcl-xL preferably bind to the BH3-only proteins Bim, PUMA and Noxa, but can also bind to Bak and Bax. We have also found a direct interaction between Bim, PUMA or Noxa with either Bax or Bak during apoptosis induction. In HeLa cells, interaction of Bim with Bax occurs in cytosol and then Bim/Bax complexes translocate to mitochondria. Complexes of either PUMA or Noxa with Bax or Bak were always detected at mitochondria. Overexpression of Bcl-xL or Mcl-1 delayed Bim/Bax translocation to mitochondria. These results reveal the ability of main BH3-only proteins to directly activate Bax and Bak in living cells and suggest that a complex network of interactions regulate the function of Bcl-2 family members during apoptosis.
    Journal of Biological Chemistry 01/2013; · 4.65 Impact Factor
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    ABSTRACT: Microtubule poisons and other anti-mitotic drugs induce tumor death but the molecular events linking mitotic arrest to cell death are still not fully understood. We have analyzed cell fate after mitotic arrest produced by the microtubule-destabilizing drug vincristine in a panel of human tumor cell lines showing different response to vincristine. In Jurkat, RPMI 8226 and HeLa cells, apoptosis was triggered shortly after vincristine-induced mitotic arrest. However, A549 cells, which express a great amount of Bcl-x(L) and undetectable amounts of Bak, underwent mitotic slippage prior to cell death. However, when Bcl-x(L) gene was silenced in A549 cells, vincristine induced apoptosis during mitotic arrest. Another different behavior was found in MiaPaca2 cells, where vincristine caused death by mitotic catastrophe that switched to apoptosis when cyclin B1 degradation was prevented by proteasome inhibition. Overexpression of Bcl-x(L) or silencing Bax and Bak expression delayed the onset of apoptosis in Jurkat and RPMI 8226 cells, enabling mitotic slippage and endoreduplication. In HeLa cells, overexpression of Bcl-x(L) switched cell death from apoptosis to mitotic catastrophe. Mcl-1 offered limited protection to vincristine-induced cell death and Mcl-1 degradation was not essential for vincristine-induced death. All these results, taken together, indicate that the Bcl-x(L)/Bak ratio and the ability to degrade cyclin B1 determine cell fate after mitotic arrest in the different tumor cell types.
    Toxicology and Applied Pharmacology 12/2011; 258(3):384-93. · 3.98 Impact Factor
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    ABSTRACT: Gold compounds are being investigated as potential antitumor drugs. Some gold(III) derivatives have been shown to induce cell death in solid tumors but their mechanism of action differs from that of cisplatin, since most of these compounds do not bind to DNA. We have explored cellular events triggered by three different iminophosphorane-organogold(III) compounds in leukemia cells (a neutral compound with two chloride ligands [Au{κ(2)-C,N-C(6)H(4)(PPh(2)=N(C(6)H(5))-2}Cl(2)] 1, and two cationic compounds with either a dithiocarbamate ligand [Au{κ(2)-C,N-C(6)H(4)(PPh(2)=N(C(6)H(5))-2}(S(2)CN-Me(2))]PF(6)2, or a water-soluble phosphine and a chloride ligand [Au{κ(2)-C,N-C(6)H(4)(PPh(2)=N(C(6)H(5))-2}(P{Cp(m-C(6)H(4)-SO(3)Na)(2)}(3)) Cl]PF(6)3). All three compounds showed higher toxicity against leukemia cells when compared to normal T-lymphocytes. Compounds 1 and 2 induced both necrosis and apoptosis, while 3 was mainly apoptotic. Necrotic cell death induced by 1 and 2 was Bax/Bak- and caspase-independent, while apoptosis induced by 3 was Bax/Bak-dependent. Reactive oxygen species (ROS) production at the mitochondrial level was a critical step in the antitumor effect of these compounds.
    Journal of inorganic biochemistry 06/2011; 105(10):1306-13. · 3.25 Impact Factor
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    ABSTRACT: The influence of environmental factors on microcystin production by toxic cyanobacteria has been extensively studied. However, the effect of nitrogen on the synthesis of this toxin remains unclear because of the literature contradictory data. The aim of this work was to determine how nitrate affects the transcriptional response of mcyD gene and the microcystin-LR synthesis in Microcystis aeruginosa PCC 7806. For first time real time RT-PCR has been used to investigate the effect of nitrogen availability. Our results show that, under laboratory conditions, an excess of nitrate triggers Microcystis aeruginosa growth without increasing the synthesis of microcystin-LR per cell. The concentration of microcystin in the cultures correlates with mcyD gene expression, being both parameters independent of nitrate availability. Analysis of the bidirectional promoter mcy unravels that the transcription start points of mcyA and mcyD genes did not change under different nitrate regimes. The effect of nitrate inputs in the development of toxic blooms is primarily due to the increased growth rate and population, not to the induction of the mcy operon.
    Ecotoxicology 10/2010; 19(7):1167-73. · 2.77 Impact Factor
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    ABSTRACT: Microcystins are toxins produced by cyanobacteria that entail serious health and environmental problems. They are cyclic heptapeptides synthesized via a mixed polyketide synthase/non-ribosomal peptide synthetase system called microcystin synthetase. Environmental and nutritional factors that trigger microcystin synthesis are still debated and this work deals with the study of the influence of iron nutritional status on the microcystin synthesis. The results indicate that iron deficiency could be one of the inducing factors of the microcystin synthesis. For the first time, increased transcription of an essential mcy gene and correlative microcystin synthesis has been established. Real-time PCR analysis of mcyD, and microcystin-LR synthesis were studied on Microcystis aeruginosa PCC7806 grown in iron-replete and iron-deplete media. Iron starvation causes an increase of mcyD transcription, correlative to the increase of microcystin-LR levels. Four transcription start points were identified for mcyD and two for mcyA, and they are not changed as a consequence of iron deficiency.
    Environmental Microbiology 10/2008; 10(10):2476-83. · 6.24 Impact Factor
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    ABSTRACT: The physiological role of microcystin-LR is still under discussion, and since binding of microcystin-LR to proteins different from their main cellular targets was described, we have performed experiments in order to explore this interaction. A non-specific interaction of microcystin-LR with a variety of soluble proteins in vitro is disrupted when using organic solvents such as methanol. The isoelectric point of proteins is not affected by their interaction with microcystin-LR, even though the presence of microcystin-LR alters the pool of peptides obtained by tryptic digestions. Under the conditions tested, microcystin-LR does not exhibit affinity for DNA. Although it is unlikely that the non-specific binding of microcystin-LR to proteins has a physiological meaning, one must be aware of the fact that determinations of the toxin extracted from any biological sample may be affected by the presence of proteins in the extracts. Consequently, we strongly recommend use organic solvents and to lyophilise the tissue samples to guarantee the accessibility of these organic solvents to microcystin-LR when performing experiments with tissue or cell extracts.
    Toxicology in Vitro 08/2008; 22(7):1714-8. · 2.65 Impact Factor
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    ABSTRACT: The increasing eutrophication of the fresh water reservoirs has lead to the frequent occurrence of uncontrolled proliferations of phytoplankton, mainly of cyanobacteria, which can have toxic character. One of the most virulent strains is the genera Microcystis which produces the most prevalent cyanotoxin, the microcystin. The microcystins pose a serious health and environmental risk, and the Spanish current legislation already controls the level of the toxin in potable waters. There are two main open questions about microcystins that have been long discussed and still remain unresolved. Firstly, what the factors that trigger microcystin synthesis are, and secondly, what the physiological role of these molecules could be in the cyanobacteria, since they are mainly found as endotoxins. La creciente eutrofización de los acuíferos ha provocado que frecuentemente se produzcan proliferaciones incontroladas de fitoplancton, principalmente de cianobacterias, que pueden tener carácter tóxico. Una de las cepas más virulentas es del género Microcystis y produce la cianotoxina más ubicua, la microcistina. Las microcistinas producen graves problemas sanitarios y medioambientales, y la legislación Española contempla ya su control en aguas emergentes de depuradoras. Hay dos grandes problemas planteados en torno a la microcistina. En primer lugar se desconoce qu¿e factores son los que desencadenan su síntesis, y en segundo lugar, qué papel fisiológico juegan en la cianobacteria, ya que se trata mayoritariamente de una endotoxina.
    Revista de la Academia de Ciencias Exactas, Físicas, Químicas y Naturales de Zaragoza, ISSN 0370-3207, Nº. 62, 2007, pags. 135-146.