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1.
Adv Exp Med Biol ; 1131: 7-26, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31646505

RESUMO

Measuring free Ca2+ concentration ([Ca2+]) in the cytosol or organelles is routine in many fields of research. The availability of membrane permeant forms of indicators coupled with the relative ease of transfecting cell lines with biological Ca2+ sensors have led to the situation where cellular and subcellular [Ca2+] is examined by many non-specialists. In this chapter, we evaluate the most used Ca2+ indicators and highlight what their major advantages and disadvantages are. We stress the potential pitfalls of non-ratiometric techniques for measuring Ca2+ and the clear advantages of ratiometric methods. Likely improvements and new directions for Ca2+ measurement are discussed.


Assuntos
Cálcio , Citosol , Organelas , Animais , Cálcio/metabolismo , Técnicas Citológicas , Citosol/química , Citosol/metabolismo , Humanos , Organelas/química , Organelas/metabolismo
2.
Adv Exp Med Biol ; 1131: 93-129, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31646508

RESUMO

Plasma membrane Ca2+ transport ATPases (PMCA1-4, ATP2B1-4) are responsible for removing excess Ca2+ from the cell in order to keep the cytosolic Ca2+ ion concentration at the low level essential for normal cell function. While these pumps take care of cellular Ca2+ homeostasis they also change the duration and amplitude of the Ca2+ signal and can create Ca2+ gradients across the cell. This is accomplished by generating more than twenty PMCA variants each having the character - fast or slow response, long or short memory, distinct interaction partners and localization signals - that meets the specific needs of the particular cell-type in which they are expressed. It has become apparent that these pumps are essential to normal tissue development and their malfunctioning can be linked to different pathological conditions such as certain types of neurodegenerative and heart diseases, hearing loss and cancer. In this chapter we summarize the complexity of PMCA regulation and function under normal and pathological conditions with particular attention to recent developments of the field.


Assuntos
Membrana Celular , ATPases Transportadoras de Cálcio da Membrana Plasmática , Animais , Membrana Celular/enzimologia , Membrana Celular/patologia , Citosol/metabolismo , Homeostase/fisiologia , Humanos , ATPases Transportadoras de Cálcio da Membrana Plasmática/genética , ATPases Transportadoras de Cálcio da Membrana Plasmática/metabolismo
3.
Adv Exp Med Biol ; 1131: 321-336, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31646516

RESUMO

Ryanodine receptor calcium release channels (RyRs) play central roles in controlling intracellular calcium concentrations in excitable and non-excitable cells. RyRs are located in the sarcoplasmic or endoplasmic reticulum, intracellular Ca2+ storage compartment, and release Ca2+ during cellular action potentials or in response to other cellular stimuli. Mammalian cells express three structurally related isoforms of RyR. RyR1 and RyR2 are the major RyR isoforms in skeletal and cardiac muscle, respectively, and RyR3 is expressed in various tissues along with the other two isoforms. A prominent feature of RyRs is that the Ca2+ release channel activities of RyRs are regulated by calcium ions; therefore, intracellular Ca2+ release controls positive- and negative-feedback phenomena through the RyRs. RyR channel activities are also regulated by Ca2+ indirectly, i.e. through Ca2+ binding proteins at both cytosolic and sarco/endoplasmic reticulum luminal sides. Here, I summarize Ca2+-dependent feedback regulation of RyRs including recent progress in the structure/function aspects.


Assuntos
Cálcio , Regulação da Expressão Gênica , Canal de Liberação de Cálcio do Receptor de Rianodina , Animais , Cálcio/metabolismo , Citosol/metabolismo , Canal de Liberação de Cálcio do Receptor de Rianodina/química , Canal de Liberação de Cálcio do Receptor de Rianodina/genética , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Relação Estrutura-Atividade
4.
BMC Plant Biol ; 19(1): 368, 2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31429706

RESUMO

BACKGROUND: We previously reported the involvement of nitric oxide (NO) and cyclic nucleotide-gated ion channel 6 (CNGC6) in the responses of plants to heat shock (HS) exposure. To elucidate their relationship with heat tolerance in Arabidopsis thaliana, we examined the effects of HS on several groups of seedlings: wild type, cngc6, and cngc6 complementation and overexpression lines. RESULTS: After HS exposure, the level of NO was lower in cngc6 seedlings than in wild-type seedlings but significantly elevated in the transgenic lines depending on CNGC6 expression level. The treatment of seeds with calcium ions (Ca2+) enhanced the NO level in Arabidopsis seedlings under HS conditions, whereas treatment with EGTA (a Ca2+ chelator) reduced it, implicating that CNGC6 stimulates the accumulation of NO depending on an increase in cytosolic Ca2+ ([Ca2+]cyt). This idea was proved by phenotypic observations and thermotolerance testing of transgenic plants overexpressing NIA2 and NOA1, respectively, in a cngc6 background. Western blotting indicated that CNGC6 stimulated the accumulation of HS proteins via NO. CONCLUSION: These data indicate that CNGC6 acts upstream of NO in the HS pathway, which improves our insufficient knowledge of the initiation of plant responses to high temerature.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Cálcio/metabolismo , Canais de Cátion Regulados por Nucleotídeos Cíclicos/metabolismo , Óxido Nítrico/metabolismo , Termotolerância , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Canais de Cálcio/metabolismo , Canais de Cátion Regulados por Nucleotídeos Cíclicos/genética , Citosol/metabolismo , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Resposta ao Choque Térmico , Mutação , Nitrato Redutase/genética , Nitrato Redutase/metabolismo , Óxido Nítrico Sintase/genética , Óxido Nítrico Sintase/metabolismo , Plântula/genética , Plântula/metabolismo
5.
J Agric Food Chem ; 67(34): 9618-9629, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31381342

RESUMO

Astrocytes provide nutritional support, regulate inflammation, and perform synaptic functions in the human brain. Although butylated hydroxyanisole (BHA) is a well-known antioxidant, several studies in animals have indicated BHA-mediated liver toxicity, retardation in reproductive organ development and learning, and sleep deficit. However, the specific effects of BHA on human astrocytes and the underlying mechanisms are yet unclear. Here, we investigated the antigrowth effects of BHA through cell cycle arrest and downregulation of regulatory protein expression. The typical cell proliferative signaling pathways, phosphoinositide 3-kinase/protein kinase B and extracellular signal-regulated kinase 1/2, were downregulated in astrocytes after BHA treatment. BHA increased the levels of pro-apoptotic proteins, such as BAX, cytochrome c, cleaved caspase 3, and cleaved caspase 9, and decreased the level of anti-apoptotic protein BCL-XL. It also increased the cytosolic calcium level and the expression of endoplasmic reticulum stress proteins. Treatment with BAPTA-AM, a calcium chelator, attenuated the increased levels of ER stress proteins and cleaved members of the caspase family. We further performed an in vivo evaluation of the neurotoxic effect of BHA on zebrafish embryos and glial fibrillary acidic protein, a representative astrocyte biomarker, in a gfap:eGFP zebrafish transgenic model. Our results provide clear evidence of the potent cytotoxic effects of BHA on human astrocytes, which lead to disruption of the brain and nerve development.


Assuntos
Astrócitos/efeitos dos fármacos , Hidroxianisol Butilado/toxicidade , Cálcio/metabolismo , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Neurotoxinas/toxicidade , Animais , Astrócitos/metabolismo , Caspase 3/metabolismo , Caspase 9/metabolismo , Linhagem Celular , Citocromos c/metabolismo , Citosol/efeitos dos fármacos , Citosol/metabolismo , Humanos , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Peixe-Zebra
6.
Results Probl Cell Differ ; 67: 233-250, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31435798

RESUMO

Morphology of Golgi apparatus changes frequently and diversely depending on various cellular conditions and these changes correlate with the balance between membrane inflow and outflow at the Golgi via vesicular transports. In a previous study, we introduced a semi-intact cell system suitable for the reconstitution of morphological changes that organelles undergo as well as the vesicular transport between them. Semi-intact cells are cells that have undergone plasma membrane permeabilization by the cholesterol-dependent pore-forming cytolysin, streptolysin O (SLO). Permeabilization enables the introduction of various molecules into the cells, as well as the substitution of the original cytosol with an exogenously made cytosol prepared from cells in various stages of cell cycle, differentiation, and disease progression. Coupled with a green fluorescent protein(GFP)-visualization technique, this cell-based system enables the analysis of the molecular mechanisms underlying biological processes that are highly dependent on the integrity of the intracellular architecture. In this chapter, we present a variety of reconstitution assays concerning biological reactions pertaining to the Golgi apparatus.


Assuntos
Permeabilidade da Membrana Celular , Complexo de Golgi/metabolismo , Permeabilidade da Membrana Celular/efeitos dos fármacos , Citosol/química , Citosol/efeitos dos fármacos , Citosol/metabolismo , Complexo de Golgi/efeitos dos fármacos
7.
Science ; 365(6448): 28-29, 2019 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-31273112
8.
Biochemistry (Mosc) ; 84(6): 583-592, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31238858

RESUMO

Actin plays an important role in cellular adhesion, muscle and non-muscle contractility, migration, polarization, mitosis, and meiosis. Investigation of specific mechanisms underlying these processes is essential not only for fundamental research but also for clinical applications, since modulations of actin isoforms are directly or indirectly correlate with severe pathologies. In this review we summarize the isoform-specific functions of actin associated with adhesion structures, motility and division of normal and tumor cells; alterations of the expression and structural organization of actin isoforms in normal and tumor cells. Selective regulation of cytoplasmic ß- or γ-actin expression determines functional diversity between isoforms: ß-actin plays the predominant role in contraction and intercellular adhesion, and γ-actin is responsible for the cellular plasticity and motility. Similar data were obtained in different epithelial and mesenchymal neoplastic cell cultures, as well as in immunomorphological comparison of normal human tissues with tumor analogues. Reorganization of the actin cytoskeleton and cell-cell contacts is essential for proliferation control and acquisition of invasiveness in epithelial tumors.


Assuntos
Actinas/fisiologia , Isoformas de Proteínas/fisiologia , Actinas/química , Animais , Adesão Celular/fisiologia , Movimento Celular/fisiologia , Transformação Celular Neoplásica , Citoplasma/metabolismo , Citosol/metabolismo , Humanos , Mamíferos , Isoformas de Proteínas/química , Relação Estrutura-Atividade
9.
Genes Dev ; 33(15-16): 1031-1047, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31196865

RESUMO

Aneuploidy, a condition characterized by chromosome gains and losses, causes reduced fitness and numerous cellular stresses, including increased protein aggregation. Here, we identify protein complex stoichiometry imbalances as a major cause of protein aggregation in aneuploid cells. Subunits of protein complexes encoded on excess chromosomes aggregate in aneuploid cells, which is suppressed when expression of other subunits is coordinately altered. We further show that excess subunits are either degraded or aggregate and that protein aggregation is nearly as effective as protein degradation at lowering levels of excess proteins. Our study explains why proteotoxic stress is a universal feature of the aneuploid state and reveals protein aggregation as a form of dosage compensation to cope with disproportionate expression of protein complex subunits.


Assuntos
Aneuploidia , Citosol/metabolismo , Compensação de Dosagem (Genética)/fisiologia , Agregados Proteicos/genética , Humanos , Complexos Multiproteicos/química , Complexos Multiproteicos/metabolismo , Agregação Patológica de Proteínas , Subunidades Proteicas/metabolismo , Proteólise , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo
10.
Life Sci ; 232: 116592, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31228515

RESUMO

Acetoacetyl-CoA thiolase also known as acetyl-CoA acetyltransferase (ACAT) corresponds to two enzymes, one cytosolic (ACAT2) and one mitochondrial (ACAT1), which is thought to catalyse reversible formation of acetoacetyl-CoA from two molecules of acetyl-CoA during ketogenesis and ketolysis respectively. In addition to this activity, ACAT1 is also involved in isoleucine degradation pathway. Deficiency of ACAT1 is an inherited metabolic disorder, which results from a defect in mitochondrial acetoacetyl-CoA thiolase activity and is clinically characterized with patients presenting ketoacidosis. In this review I discuss the recent findings, which unexpectedly expand the known functions of ACAT1, indicating a role for ACAT1 well beyond its classical activity. Indeed ACAT1 has recently been shown to possess an acetyltransferase activity capable of specifically acetylating Pyruvate DeHydrogenase (PDH), an enzyme involved in producing acetyl-CoA. ACAT1-dependent acetylation of PDH was shown to negatively regulate this enzyme with a consequence in Warburg effect and tumor growth. Finally, the elevated ACAT1 enzyme activity in diverse human cancer cell lines was recently reported. These important novel findings on ACAT1's function and expression in cancer cell proliferation point to ACAT1 as a potential new anti-cancer target.


Assuntos
Acetil-CoA C-Acetiltransferase/metabolismo , Neoplasias/enzimologia , Acetil-CoA C-Acetiltransferase/antagonistas & inibidores , Citosol/enzimologia , Humanos , Mitocôndrias/enzimologia , Terapia de Alvo Molecular , Neoplasias/tratamento farmacológico , Processamento de Proteína Pós-Traducional , Piruvato Desidrogenase (Lipoamida)/metabolismo , Esterol O-Aciltransferase/metabolismo
11.
Chem Commun (Camb) ; 55(52): 7474-7477, 2019 Jul 04.
Artigo em Inglês | MEDLINE | ID: mdl-31184664

RESUMO

Inspired by clinical studies on alcohol abuse induced endoplasmic reticulum disruption, we designed a N-hydroxylethyl peptide assembly to regulate the ER stress response in cancer cells. Upon coupling with a coumarin derivative via an ester linkage, a prodrug was synthesized to promote esterase-facilitated self-delivery of N-hydroxylethyl peptide assemblies around the ER, inducing ER dilation. Following this, ER-specific apoptosis was effectively and efficiently activated in various types of cancer cells including drug resistant and metastatic ones.


Assuntos
Citosol/metabolismo , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Peptídeos/farmacologia , Apoptose/efeitos dos fármacos , Carboxilesterase/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cumarínicos/química , Humanos , Microscopia de Fluorescência , Peptídeos/metabolismo , Pró-Fármacos/química , Pró-Fármacos/metabolismo , Pró-Fármacos/farmacologia
12.
Biochemistry (Mosc) ; 84(4): 358-369, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31228927

RESUMO

Cytoplasmic actin structures are essential components of the eukaryotic cytoskeleton. According to the classic concepts, actin structures perform contractile and motor functions, ensuring the possibility of cell shape changes during cell spreading, polarization, and movement both in vitro and in vivo, from the early embryogenesis stages and throughout the life of a multicellular organism. Intracellular organization of actin structures, their biochemical composition, and dynamic properties play a key role in the realization of specific cellular and tissue functions and vary in different cell types. This paper is a review of recent studies on the organization and properties of actin structures in endotheliocytes, interaction of these structures with other cytoskeletal components and elements involved in cell adhesion, as well as their role in the functional activity of endothelial cells.


Assuntos
Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Citoesqueleto de Actina/química , Actinas/química , Actinas/genética , Caderinas/química , Caderinas/metabolismo , Citosol/metabolismo , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Humanos , Proteínas Associadas aos Microtúbulos/metabolismo , Microtúbulos/química , Microtúbulos/metabolismo
13.
Plant Physiol Biochem ; 141: 353-369, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31207496

RESUMO

Reactive oxygen species (ROS) - the byproducts of aerobic metabolism - influence numerous aspects of the plant life cycle and environmental response mechanisms. In plants, ROS act like a double-edged sword; they play multiple beneficial roles at low concentrations, whereas at high concentrations ROS and related redox-active compounds cause cellular damage through oxidative stress. To examine the dual role of ROS as harmful oxidants and/or crucial cellular signals, this review elaborates that (i) how plants sense and respond to ROS in various subcellular organelles and (ii) the dynamics of subsequent ROS-induced signaling processes. The recent understanding of crosstalk between various cellular compartments in mediating their redox state spatially and temporally is discussed. Emphasis on the beneficial effects of ROS in maintaining cellular energy homeostasis, regulating diverse cellular functions, and activating acclimation responses in plants exposed to abiotic and biotic stresses are described. The comprehensive view of cellular ROS dynamics covering the breadth and versatility of ROS will contribute to understanding the complexity of apparently contradictory ROS roles in plant physiological responses in less than optimum environments.


Assuntos
Oxirredução , Estresse Oxidativo , Fenômenos Fisiológicos Vegetais , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Estresse Fisiológico , Aclimatação , Antioxidantes/metabolismo , Arabidopsis/metabolismo , Núcleo Celular/metabolismo , Cloroplastos/metabolismo , Citosol/metabolismo , Regulação da Expressão Gênica , Genes de Plantas , Mitocôndrias/metabolismo , Oryza/metabolismo , Oxigênio/metabolismo , Peroxissomos/metabolismo , Fotossíntese , Populus/metabolismo
14.
Biophys Chem ; 252: 106195, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31195340

RESUMO

Astrocytes, the most common type of glial cell, are critical to the health of the central nervous system. Evidence implies that changes in the astrocyte's cytosolic calcium concentration is part of a central mechanism by which information is passed and processed in the cell, and it is linked to both external stimuli impacting the cell as well as downstream events such as metabolism and neurotransmitter release. This work proposes a novel chemical model to further the understanding of how extracellular signals could affect intracellular calcium dynamics and metabolic processes within the cell.


Assuntos
Astrócitos/metabolismo , Cálcio/metabolismo , Glucose/metabolismo , Algoritmos , Animais , Astrócitos/química , Citosol/química , Citosol/metabolismo , Humanos , Modelos Biológicos
15.
Nat Commun ; 10(1): 2752, 2019 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-31227693

RESUMO

Various studies have demonstrated that the two leaflets of cellular membranes interact, potentially through so-called interdigitation between the fatty acyl groups. While the molecular mechanism underlying interleaflet coupling remains to be fully understood, recent results suggest interactions between the very-long-chain sphingolipids in the outer leaflet, and phosphatidylserine PS18:0/18:1 in the inner leaflet, and an important role for cholesterol for these interactions. Here we review the evidence that cross-linking of sphingolipids may result in clustering of phosphatidylserine and transfer of signals to the cytosol. Although much remains to be uncovered, the molecular properties and abundance of PS 18:0/18:1 suggest a unique role for this lipid.


Assuntos
Membrana Celular/fisiologia , Bicamadas Lipídicas/metabolismo , Fosfatidilserinas/metabolismo , Esfingolipídeos/metabolismo , Animais , Linhagem Celular , Colesterol/metabolismo , Reagentes para Ligações Cruzadas/metabolismo , Citosol/fisiologia , Humanos
16.
Biosci Rep ; 39(6)2019 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-31160481

RESUMO

An explosion of sequence information in the genomics era has thrown up thousands of protein sequences without functional assignment. Though our ability to predict function based on sequence alone is improving steadily, we still have a long way to go. Proteins with common evolutionary origins carry telling sequence signatures, which ought to reveal their biological roles. These sequence signatures have allowed us to classify proteins into families with similar structures, and possibly, functions. Yet, evolution is a perpetual tinkerer, and hence, sequence signatures alone have proved inadequate in understanding the physiological activities of proteins. One such enigmatic family of enzymes is the NUDIX ( nu cleoside di phosphate linked to a moiety X ) hydrolase family that has over 80000 members from all branches of the tree of life. Though MutT, the founding member of this family, was identified in 1954, we are only now beginning to understand the diversity of substrates and biological roles that these enzymes demonstrate. In a recent article by Cordeiro et al. in Bioscience Reports [Biosci. Rep. (2019)], two members of this protein family from the human pathogen Trypanosoma brucei were deorphanized as being polyphosphate hydrolases. The authors show that of the five NUDIX hydrolases coded by the T. brucei genomes, TbNH2 and TbNH4, show in vitro hydrolytic activity against inorganic polyphosphate. Through classical biochemistry and immunostaining microscopy, differences in their substrate specificities and sub-cellular localization were revealed. These new data provide a compelling direction to the study of Trypanosome stress biology as well as our understanding of the NUDIX enzyme family.


Assuntos
Trypanosoma brucei brucei , Hidrolases Anidrido Ácido , Citosol , Humanos , Microcorpos , Polifosfatos , Pirofosfatases , Especificidade por Substrato
17.
MBio ; 10(3)2019 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-31064825

RESUMO

The mitochondrial Ca2+ uptake in trypanosomatids, which belong to the eukaryotic supergroup Excavata, shares biochemical characteristics with that of animals, which, together with fungi, belong to the supergroup Opisthokonta. However, the composition of the mitochondrial calcium uniporter (MCU) complex in trypanosomatids is quite peculiar, suggesting lineage-specific adaptations. In this work, we used Trypanosoma cruzi to study the role of orthologs for mitochondrial calcium uptake 1 (MICU1) and MICU2 in mitochondrial Ca2+ uptake. T. cruzi MICU1 (TcMICU1) and TcMICU2 have mitochondrial targeting signals, two canonical EF-hand calcium-binding domains, and localize to the mitochondria. Using the CRISPR/Cas9 system (i.e., clustered regularly interspaced short palindromic repeats with Cas9), we generated TcMICU1 and TcMICU2 knockout (-KO) cell lines. Ablation of either TcMICU1 or TcMICU2 showed a significantly reduced mitochondrial Ca2+ uptake in permeabilized epimastigotes without dissipation of the mitochondrial membrane potential or effects on the AMP/ATP ratio or citrate synthase activity. However, none of these proteins had a gatekeeper function at low cytosolic Ca2+ concentrations ([Ca2+]cyt), as occurs with their mammalian orthologs. TcMICU1-KO and TcMICU2-KO epimastigotes had a lower growth rate and impaired oxidative metabolism, while infective trypomastigotes have a reduced capacity to invade host cells and to replicate within them as amastigotes. The findings of this work, which is the first to study the role of MICU1 and MICU2 in organisms evolutionarily distant from animals, suggest that, although these components were probably present in the last eukaryotic common ancestor (LECA), they developed different roles during evolution of different eukaryotic supergroups. The work also provides new insights into the adaptations of trypanosomatids to their particular life styles.IMPORTANCE Trypanosoma cruzi is the etiologic agent of Chagas disease and belongs to the early-branching eukaryotic supergroup Excavata. Its mitochondrial calcium uniporter (MCU) subunit shares similarity with the animal ortholog that was important to discover its encoding gene. In animal cells, the MICU1 and MICU2 proteins act as Ca2+ sensors and gatekeepers of the MCU, preventing Ca2+ uptake under resting conditions and favoring it at high cytosolic Ca2+ concentrations ([Ca2+]cyt). Using the CRISPR/Cas9 technique, we generated TcMICU1 and TcMICU2 knockout cell lines and showed that MICU1 and -2 do not act as gatekeepers at low [Ca2+]cyt but are essential for normal growth, host cell invasion, and intracellular replication, revealing lineage-specific adaptations.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Cálcio/metabolismo , Mitocôndrias/metabolismo , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Proteínas de Protozoários/metabolismo , Trypanosoma cruzi/genética , Adaptação Fisiológica , Transporte Biológico , Sistemas CRISPR-Cas , Proteínas de Ligação ao Cálcio/genética , Proteínas de Transporte de Cátions , Citosol/química , Citosol/metabolismo , Técnicas de Inativação de Genes , Humanos , Proteínas de Transporte da Membrana Mitocondrial/genética , Proteínas de Protozoários/genética , Trypanosoma cruzi/patogenicidade
18.
Int J Mol Sci ; 20(9)2019 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-31075903

RESUMO

Boron (B) is a micronutrient for plant development, and its deficiency alters many physiological processes. However, the current knowledge on how plants are able to sense the B-starvation signal is still very limited. Recently, it has been reported that B deprivation induces an increase in cytosolic calcium concentration ([Ca2+]cyt) in Arabidopsis thaliana roots. The aim of this work was to research in Arabidopsis whether [Ca2+]cyt is restored to initial levels when B is resupplied and elucidate whether apoplastic Ca2+ is the major source for B-deficiency-induced rise in [Ca2+]cyt. The use of chemical compounds affecting Ca2+ homeostasis showed that the rise in root [Ca2+]cyt induced by B deficiency was predominantly owed to Ca2+ influx from the apoplast through plasma membrane Ca2+ channels in an IP3-independent manner. Furthermore, B resupply restored the root [Ca2+]cyt. Interestingly, expression levels of genes encoding Ca2+ transporters (ACA10, plasma membrane PIIB-type Ca2+-ATPase; and CAX3, vacuolar cation/proton exchanger) were upregulated by ethylene glycol tetraacetic acid (EGTA) and abscisic acid (ABA). The results pointed out that ACA10, and especially CAX3, would play a major role in the restoration of Ca2+ homeostasis after 24 h of B deficiency.


Assuntos
Arabidopsis/metabolismo , Boro/deficiência , Sinalização do Cálcio , Cálcio/metabolismo , Citosol/metabolismo , Arabidopsis/genética , Boro/farmacologia , Sinalização do Cálcio/efeitos dos fármacos , Citosol/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Plântula/efeitos dos fármacos , Plântula/metabolismo
19.
Nat Commun ; 10(1): 1994, 2019 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-31040279

RESUMO

Lignin biosynthesis is evolutionarily conserved among higher plants and features a critical 3-hydroxylation reaction involving phenolic esters. However, increasing evidence questions the involvement of a single pathway to lignin formation in vascular plants. Here we describe an enzyme catalyzing the direct 3-hydroxylation of 4-coumarate to caffeate in lignin biosynthesis as a bifunctional peroxidase that oxidizes both ascorbate and 4-coumarate at comparable rates. A combination of biochemical and genetic evidence in the model plants Brachypodium distachyon and Arabidopsis thaliana supports a role for this coumarate 3-hydroxylase (C3H) in the early steps of lignin biosynthesis. The subsequent efficient O-methylation of caffeate to ferulate in grasses is substantiated by in vivo biochemical assays. Our results identify C3H as the only non-membrane bound hydroxylase in the lignin pathway and revise the currently accepted models of lignin biosynthesis, suggesting new gene targets to improve forage and bioenergy crops.


Assuntos
Citosol/enzimologia , Lignina/biossíntese , Arabidopsis/metabolismo , Ascorbato Peroxidases , Brachypodium/metabolismo , Ácidos Cafeicos/metabolismo , Ácidos Cumáricos/metabolismo , Oxigenases de Função Mista/genética , Oxigenases de Função Mista/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
20.
Chimia (Aarau) ; 73(6): 395-405, 2019 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-31118122

RESUMO

Besides their medical importance, the parasitic protozoan Trypanosoma brucei and its relatives are experimentally highly accessible model systems for many cell biological processes. Trypanosomes are phylogenetically essentially unrelated to the popular model eukaryotes, such as yeast and animals, and thus show several unique features, many of which are connected to RNA. Here we review the tRNA biology of trypanosomes. Even though tRNAs were already discovered 60 years ago, owing to current technological advances in the field, research on tRNA biology has seen a Renaissance in recent years. First we discuss the extensive mitochondrial tRNA import process and the consequences it has for the parasite. Next we focus on trypanosomal aminoacyl-tRNA synthetases, some of which may be exploited as drug targets. Furthermore, we summarize what is known about trypanosomal tRNA modifications in both the cytosol and the mitochondrion. Finally, we provide an overview on the emerging field of tRNA-derived fragments and their possible function as translation regulators.


Assuntos
Trypanosoma , Animais , Citosol , Mitocôndrias , RNA de Transferência
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