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1.
Life Sci ; 234: 116768, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31445027

RESUMO

In prostate cancer development, the androgen receptor (AR) signaling plays a crucial role during both formation of early prostate lesions and progression to the lethal, incurable castration resistant stage. Accordingly, numerous approaches have been developed to inhibit AR activity including androgen deprivation therapy, application of the AR antagonists as well as the use of taxanes. However, these treatments, although effective initially, resistance inevitably occur for most of the patients within several years and limiting the therapeutic efficacy. Of note, alterations and reactivation of the AR signaling pathway have been demonstrated as the major reasons for the observed resistance. Accumulating evidences have suggested that synthesis of AR splicing variants, in particular, the constitutively active AR-V7, is one of the most important mechanisms that contribute to the abnormal AR signaling. In addition, clinical data also highlight the potential of using AR-V7 as a predictive biomarker and a therapeutic target in metastatic castration resistant prostate cancer (mCRPC). In this review, we summarize the recent findings concerning the specific role of AR-V7 in CRPC progression, drug resistance and its potential value in clinical assessment.


Assuntos
Antagonistas de Receptores de Andrógenos/uso terapêutico , Antineoplásicos/uso terapêutico , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Receptores Androgênicos/genética , Taxoides/uso terapêutico , Antagonistas de Receptores de Andrógenos/farmacologia , Animais , Antineoplásicos/farmacologia , Progressão da Doença , Resistencia a Medicamentos Antineoplásicos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologia , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Receptores Androgênicos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Taxoides/farmacologia
3.
Dokl Biochem Biophys ; 486(1): 224-228, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31367827

RESUMO

TRF2 protein (TBP-related factor 2) can substitute for TBP forming alternative transcription initiation complexes on TATA-less promoters, including the promoters of histone H1 and piRNA clusters required for transposon repression. The Drosophilatrf2 gene codes for two isoforms: a "short" and a "long" one, in which the same short TRF2 sequence is preceded by a long N-terminal domain. Here, we demonstrated that the long TFR2 isoform has a greater functional activity than the short isoform by expressing each of them at a reduced rate under the endogenous promoters. Expression of the long isoform alone affects neither the flies' viability nor the sex ratio. Expression of the short isoform alone leads to the phenotype described for the trf2 gene insufficiency and derepression of transposable elements, that is, decreased viability, disturbance of homologous chromosome pairing and segregation, and apparent female-biased sex ratio.


Assuntos
Drosophila melanogaster/metabolismo , Proteína 2 de Ligação a Repetições Teloméricas/química , Proteína 2 de Ligação a Repetições Teloméricas/metabolismo , Animais , Drosophila melanogaster/genética , Mutação , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismo , Proteína 2 de Ligação a Repetições Teloméricas/genética
4.
Nat Commun ; 10(1): 3004, 2019 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-31285436

RESUMO

Identity determining transcription factors (TFs), or core regulatory (CR) TFs, are governed by cell-type specific super enhancers (SEs). Drugs to selectively inhibit CR circuitry are of high interest for cancer treatment. In alveolar rhabdomyosarcoma, PAX3-FOXO1 activates SEs to induce the expression of other CR TFs, providing a model system for studying cancer cell addiction to CR transcription. Using chemical genetics, the systematic screening of chemical matter for a biological outcome, here we report on a screen for epigenetic chemical probes able to distinguish between SE-driven transcription and constitutive transcription. We find that chemical probes along the acetylation-axis, and not the methylation-axis, selectively disrupt CR transcription. Additionally, we find that histone deacetylases (HDACs) are essential for CR TF transcription. We further dissect the contribution of HDAC isoforms using selective inhibitors, including the newly developed selective HDAC3 inhibitor LW3. We show HDAC1/2/3 are the co-essential isoforms that when co-inhibited halt CR transcription, making CR TF sites hyper-accessible and disrupting chromatin looping.


Assuntos
Elementos Facilitadores Genéticos/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Inibidores de Histona Desacetilases/farmacologia , Histona Desacetilases/metabolismo , Rabdomiossarcoma/genética , Acetilação/efeitos dos fármacos , Linhagem Celular Tumoral , Cromatina/efeitos dos fármacos , Cromatina/metabolismo , Genômica/métodos , Sequenciamento de Nucleotídeos em Larga Escala , Ensaios de Triagem em Larga Escala , Inibidores de Histona Desacetilases/química , Histona Desacetilases/química , Humanos , Simulação de Dinâmica Molecular , Sondas Moleculares/química , Sondas Moleculares/farmacologia , Proteínas de Fusão Oncogênica/genética , Fatores de Transcrição Box Pareados/genética , Cultura Primária de Células , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismo , Rabdomiossarcoma/patologia , Análise de Sequência de RNA , Transcrição Genética/efeitos dos fármacos
5.
Exp Parasitol ; 204: 107721, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31288023

RESUMO

BACKGROUND: Toxoplasma gondii (T. gondii) is an obligate intracellular protozoan able to infect humans and it is common in pregnant women. During pregnancy and lactation, there are changes in the concentration of 17ß-estradiol (E2), progesterone (Prg), and prolactin (PRL). It is known that a proinflamatory response reduces the susceptibility to be infected, and this response may change according to hormonal impairment. Monocytes and macrophages are the main barrier against many intracellular microorganisms, due to their ability to produce cytokines. The aim of this work was to determine the effect of E2, progesterone, and PRL on the infective capacity of T. gondii, proinflamatory immune response modulation and the expression of hormonal receptors on THP-1 cell stimulated with T. gondii. METHODS: The THP-1 cells were infected with 1500 T. gondii tachyzoites, of RH strain. Stimuli were conducted with recombinant PRL (200 ng/mL), E2 (40 nM) y Prg (40 nM). MTT assays were performed to evaluate cellular viability. Western blot assays were carried out to evaluate the expression of the hormonal receptors (PRLR, ERα, and ERß). Cytokines produced were measured with a magnetic bead kit directed to 17 cytokines. RESULTS: Stimuli with E2 and Prg increased T. gondii infection in monocytes after 48 h; however, no differences in infection were observed in PRL stimulus. The E2 decreased the secretion of IL-12 and IL-1ß and PRL did not modify the production of these cytokines in THP-1 cells stimulated with T. gondii; however, both hormones increased the production of IL-10. Besides, PRL augmented the production of IL-4 and IL-13. In contrast, Prg reduced these cytokines. Our results show that T. gondii induces the expression of ERα and ERß and lowers PRLR. The hormones modify the expression of the receptors of other hormones: Prg decreases PRLR, ERß and increases ERα; E2 diminishes PRLR; and PRL decreases ERα and ERß expression. CONCLUSION: The hormones can increase T. gondii infection and could be mediating an anti-inflammatory response in THP-1 cells. T. gondii induces changes in the expression of hormonal receptors.


Assuntos
Citocinas/metabolismo , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/metabolismo , Receptores da Prolactina/metabolismo , Células THP-1/metabolismo , Toxoplasma/fisiologia , Animais , Corantes , Estradiol/metabolismo , Feminino , Humanos , Camundongos , Progesterona/metabolismo , Prolactina/metabolismo , Isoformas de Proteínas/metabolismo , Células THP-1/imunologia , Células THP-1/parasitologia , Sais de Tetrazólio , Tiazóis , Toxoplasma/crescimento & desenvolvimento
6.
Nat Commun ; 10(1): 2767, 2019 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-31235694

RESUMO

The coactivator PGC-1α1 is activated by exercise training in skeletal muscle and promotes fatigue-resistance. In exercised muscle, PGC-1α1 enhances the expression of kynurenine aminotransferases (Kats), which convert kynurenine into kynurenic acid. This reduces kynurenine-associated neurotoxicity and generates glutamate as a byproduct. Here, we show that PGC-1α1 elevates aspartate and glutamate levels and increases the expression of glycolysis and malate-aspartate shuttle (MAS) genes. These interconnected processes improve energy utilization and transfer fuel-derived electrons to mitochondrial respiration. This PGC-1α1-dependent mechanism allows trained muscle to use kynurenine metabolism to increase the bioenergetic efficiency of glucose oxidation. Kat inhibition with carbidopa impairs aspartate biosynthesis, mitochondrial respiration, and reduces exercise performance and muscle force in mice. Our findings show that PGC-1α1 activates the MAS in skeletal muscle, supported by kynurenine catabolism, as part of the adaptations to endurance exercise. This crosstalk between kynurenine metabolism and the MAS may have important physiological and clinical implications.


Assuntos
Metabolismo Energético/fisiologia , Fadiga/fisiopatologia , Cinurenina/metabolismo , Músculo Esquelético/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Adaptação Fisiológica , Animais , Aspartato Aminotransferases/metabolismo , Ácido Aspártico/metabolismo , Carbidopa/farmacologia , Respiração Celular/efeitos dos fármacos , Respiração Celular/fisiologia , Metabolismo Energético/efeitos dos fármacos , Glicólise/fisiologia , Malatos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Modelos Animais , Músculo Esquelético/fisiopatologia , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Condicionamento Físico Animal/fisiologia , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Transaminases/antagonistas & inibidores , Transaminases/metabolismo
7.
Folia Histochem Cytobiol ; 57(2): 84-93, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31198984

RESUMO

INTRODUCTION: WNT5A (Wnt family member 5A) belongs to the WNT family of secreted signaling glycoproteins that play essential role in developmental, physiological and pathological processes. WNT5A was shown to take part in carcinogenesis process playing both oncogenic and suppressor functions in various types of human malignancies. This study aimed to assess the expression of the WNT5A gene at the mRNA and protein levels in the specimens derived from endometrial cancer (EC) or unchanged control endometrium. The associations between the WNT5A expression levels and clinicopathological characteristics and survival of EC patients were evaluated. MATERIALS AND METHODS: Total RNA was isolated in order to assess the relative amounts of WNT5A mRNA by quantitative polymerase chain reaction (QPCR) in samples of unchanged endometrial control (n = 8) and tumor samples of EC patients (n = 28). Immunohistochemistry (IHC) was used to determine the presence of WNT5A protein in the sections of formalin-fixed, paraffin-embedded tissue specimens derived from unchanged endome-trial controls (n = 6) and EC tumors (n = 19). Significance of differences in WNT5A expression levels between the studied groups of EC patients and correlations between the WNT5A and demographic data, pathological features, hematological parameters and overall survival of the patients were evaluated by statistical analysis. RESULTS: The level of WNT5A mRNA was decreased in EC in comparison to unchanged endometrium. WNT5A expression was associated with primary tumor invasion status exhibiting reduced level of transcripts in EC that involved organs beyond the uterus when compared to the uterus-confined cancers. WNT5A immunoreactivity was visualized in the cytoplasm and nuclei of EC cells as well as in the luminal and glandular epithelial cells of unchanged endometrium. WNT5A mRNA expression levels correlated negatively with cytoplasmic, and positively with nuclear immunoreactivity of the WNT5A protein in the EC cells. In addition, the relationships between blood leucocyte count (in particular granulocytes and lymphocytes) of patients with EC and their WNT5A mRNA and protein expression levels were established. A positive correlation between the nuclear immunoexpression of WNT5A protein in the cancer cells in cell nuclei and mean platelet volume in blood was also found. CONCLUSIONS: The results of the first study of WNT5A expression at the transcript and protein levels indicate that it could be considered as a potential marker of molecular changes that take place during EC development.


Assuntos
Neoplasias do Endométrio/genética , Neoplasias do Endométrio/metabolismo , RNA Mensageiro/metabolismo , Proteína Wnt-5a/genética , Proteína Wnt-5a/metabolismo , Adulto , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/imunologia , Biomarcadores Tumorais/metabolismo , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Regulação para Baixo , Neoplasias do Endométrio/diagnóstico , Neoplasias do Endométrio/mortalidade , Endométrio/patologia , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Imuno-Histoquímica , Pessoa de Meia-Idade , Prognóstico , Isoformas de Proteínas/genética , Isoformas de Proteínas/imunologia , Isoformas de Proteínas/metabolismo , Proteína Wnt-5a/imunologia
8.
Nat Commun ; 10(1): 2692, 2019 06 19.
Artigo em Inglês | MEDLINE | ID: mdl-31217420

RESUMO

Sphingomyelin phosphodiesterase acid-like 3b (SMPDL3b) is a lipid raft enzyme that regulates plasma membrane (PM) fluidity. Here we report that SMPDL3b excess, as observed in podocytes in diabetic kidney disease (DKD), impairs insulin receptor isoform B-dependent pro-survival insulin signaling by interfering with insulin receptor isoforms binding to caveolin-1 in the PM. SMPDL3b excess affects the production of active sphingolipids resulting in decreased ceramide-1-phosphate (C1P) content as observed in human podocytes in vitro and in kidney cortexes of diabetic db/db mice in vivo. Podocyte-specific Smpdl3b deficiency in db/db mice is sufficient to restore kidney cortex C1P content and to protect from DKD. Exogenous administration of C1P restores IR signaling in vitro and prevents established DKD progression in vivo. Taken together, we identify SMPDL3b as a modulator of insulin signaling and demonstrate that supplementation with exogenous C1P may represent a lipid therapeutic strategy to treat diabetic complications such as DKD.


Assuntos
Antígenos CD/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 3/metabolismo , Nefropatias Diabéticas/patologia , Insulina/metabolismo , Receptor de Insulina/metabolismo , Esfingomielina Fosfodiesterase/metabolismo , Animais , Caveolina 1/metabolismo , Linhagem Celular , Membrana Celular/metabolismo , Ceramidas/metabolismo , Ceramidas/uso terapêutico , Nucleotídeo Cíclico Fosfodiesterase do Tipo 3/genética , Nefropatias Diabéticas/tratamento farmacológico , Modelos Animais de Doenças , Feminino , Humanos , Masculino , Camundongos , Camundongos Knockout , Podócitos/citologia , Podócitos/metabolismo , Isoformas de Proteínas/metabolismo , Transdução de Sinais , Resultado do Tratamento
9.
Biochemistry (Mosc) ; 84(4): 329-345, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31228925

RESUMO

The review describes functional and structural features of different isoforms of prolactin receptor, mechanisms of signaling pathway activation, and molecular messengers involved in the transmission and termination of signal from the prolactin receptor isoforms. Changes in the ratio between prolactin receptor isoforms, key mediators of prolactin signal transduction and termination in various organs and tissues, are analyzed. Special attention is given to the role of molecular mediators and the ratio between the isoforms in normal physiological functions and pathologies. Approaches for therapeutic correction of prolactin signaling impairments are discussed.


Assuntos
Prolactina/metabolismo , Receptores da Prolactina/metabolismo , Animais , Humanos , Neoplasias/metabolismo , Neoplasias/patologia , Proteínas Inibidoras de STAT Ativados/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Receptores da Prolactina/genética , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais , Proteínas Supressoras da Sinalização de Citocina/metabolismo
10.
Proc Jpn Acad Ser B Phys Biol Sci ; 95(5): 198-210, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31080188

RESUMO

Iron plays many important roles in the brain, including involvement in myelination, neurotransmission and electron transfer in the respiratory chain. Transferrin (Tf), an iron transporter, is mainly biosynthesized in the liver, but can also be biosynthesized in the brain; i.e., by oligodendrocytes and the choroid plexus, a cerebrospinal fluid (CSF) producing tissue. The CSF contains two Tf isoforms, brain-type Tf and serum-type Tf, which differ in their glycan structures. Brain-type Tf is uniquely glycolsylated with biantennary asialo- and agalacto-complex type N-glycans that carry bisecting ß1,4-GlcNAc and core α1,6-Fuc. The glycans of serum-type Tf in the CSF are similar to those of Tf in serum. Biochemical analyses reveal that the apparent molecular size of brain-type Tf is smaller than that of serum-type Tf, and that hydrophobic patches are exposed on brain-type Tf as demonstrated by hydrophobic probe binding studies. We found that brain-type Tf levels were decreased in idiopathic normal pressure hydrocephalus, in which CSF production is suspected to decrease, while brain-type Tf increased in spontaneous intracranial hypotension, in which CSF production is suspected to increase. These results suggest that brain-type Tf could be a biomarker of altered CSF production.


Assuntos
Doenças do Sistema Nervoso/líquido cefalorraquidiano , Transferrina/líquido cefalorraquidiano , Animais , Encéfalo/metabolismo , Humanos , Polissacarídeos/metabolismo , Isoformas de Proteínas/líquido cefalorraquidiano , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismo , Transferrina/química , Transferrina/metabolismo
11.
Int J Mol Sci ; 20(10)2019 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-31091671

RESUMO

The human Na+/H+ exchanger isoform 1 (NHE1) is a plasma membrane transport protein that plays an important role in pH regulation in mammalian cells. Because of the generation of protons by intermediary metabolism as well as the negative membrane potential, protons accumulate within the cytosol. Extracellular signal-regulated kinase (ERK)-mediated regulation of NHE1 is important in several human pathologies including in the myocardium in heart disease, as well as in breast cancer as a trigger for growth and metastasis. NHE1 has a N-terminal, a 500 amino acid membrane domain, and a C-terminal 315 amino acid cytosolic domain. The C-terminal domain regulates the membrane domain and its effects on transport are modified by protein binding and phosphorylation. Here, we discuss the physiological regulation of NHE1 by ERK, with an emphasis on the critical effects on structure and function. ERK binds directly to the cytosolic domain at specific binding domains. ERK also phosphorylates NHE1 directly at multiple sites, which enhance NHE1 activity with subsequent downstream physiological effects. The NHE1 cytosolic regulatory tail possesses both ordered and disordered regions, and the disordered regions are stabilized by ERK-mediated phosphorylation at a phosphorylation motif. Overall, ERK pathway mediated phosphorylation modulates the NHE1 tail, and affects the activity, structure, and function of this membrane protein.


Assuntos
Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Trocador 1 de Sódio-Hidrogênio/metabolismo , Animais , Humanos , Fosforilação , Domínios Proteicos , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Trocador 1 de Sódio-Hidrogênio/química , Trocador 1 de Sódio-Hidrogênio/genética
12.
Cell Mol Life Sci ; 76(15): 2917-2932, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31123777

RESUMO

Protein arginine methyltransferases (PRMTs) catalyze the methyl transfer to the arginine residues of protein substrates and are classified into three major types based on the final form of the methylated arginine. Recent studies have shown a strong correlation between PRMT expression level and the prognosis of cancer patients. Currently, crystal structures of eight PRMT members have been determined. Kinetic and structural studies have shown that all PRMTs share similar, but unique catalytic and substrate recognition mechanism. In this review, we discuss the structural similarities and differences of different PRMT members, focusing on their overall structure, S-adenosyl-L-methionine-binding pocket, substrate arginine recognition and catalytic mechanisms. Since PRMTs are valuable targets for drug discovery, we also rationally classify the known PRMT inhibitors into five classes and discuss their mechanisms of action at the atomic level.


Assuntos
Proteína-Arginina N-Metiltransferases/metabolismo , Arginina/metabolismo , Sítios de Ligação , Domínio Catalítico , Inibidores Enzimáticos/química , Inibidores Enzimáticos/metabolismo , Humanos , Metilação , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/metabolismo , Proteína-Arginina N-Metiltransferases/antagonistas & inibidores , Especificidade por Substrato
13.
Cell Mol Life Sci ; 76(15): 2885-2898, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31098676

RESUMO

The mixed lineage leukemia (MLL) family of proteins became known initially for the leukemia link of its founding member. Over the decades, the MLL family has been recognized as an important class of histone H3 lysine 4 (H3K4) methyltransferases that control key aspects of normal cell physiology and development. Here, we provide a brief history of the discovery and study of this family of proteins. We address two main questions: why are there so many H3K4 methyltransferases in mammals; and is H3K4 methylation their key function?


Assuntos
Histona-Lisina N-Metiltransferase/metabolismo , Proteína de Leucina Linfoide-Mieloide/metabolismo , Animais , Histona-Lisina N-Metiltransferase/química , Histonas/metabolismo , Humanos , Metilação , Proteína de Leucina Linfoide-Mieloide/química , Proteína de Leucina Linfoide-Mieloide/classificação , Domínios Proteicos , Isoformas de Proteínas/química , Isoformas de Proteínas/classificação , Isoformas de Proteínas/metabolismo
14.
Nat Commun ; 10(1): 2178, 2019 05 16.
Artigo em Inglês | MEDLINE | ID: mdl-31097717

RESUMO

T-cells play a crucial role in progression of autoimmunity, including vitiligo, yet the initial steps triggering their activation and tissue damage remain unknown. Here we demonstrate increased presence of type-1 innate lymphoid cells (NK and ILC1)-producing interferon gamma (IFNγ) in the blood and in non-lesional skin of vitiligo patients. Melanocytes of vitiligo patients have strong basal expression of chemokine-receptor-3 (CXCR3) isoform B which is directly regulated by IFNγ. CXCR3B activation by CXCL10 at the surface of cultured human melanocytes induces their apoptosis. The remaining melanocytes, activated by the IFNγ production, express co-stimulatory markers which trigger T-cell proliferation and subsequent anti-melanocytic immunity. Inhibiting the CXCR3B activation prevents this apoptosis and the further activation of T cells. Our results emphasize the key role of CXCR3B in apoptosis of melanocytes and identify CXCR3B as a potential target to prevent and to treat vitiligo by acting at the early stages of melanocyte destruction.


Assuntos
Autoimunidade , Melanócitos/imunologia , Receptores CXCR3/metabolismo , Linfócitos T/imunologia , Vitiligo/imunologia , Adulto , Idoso , Apoptose/imunologia , Biópsia , Células Cultivadas , Quimiocina CXCL10/metabolismo , Feminino , Humanos , Imunidade Inata , Interferon gama/imunologia , Interferon gama/metabolismo , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Ativação Linfocitária , Masculino , Melanócitos/metabolismo , Pessoa de Meia-Idade , Cultura Primária de Células , Isoformas de Proteínas/imunologia , Isoformas de Proteínas/metabolismo , Receptores CXCR3/imunologia , Pele/citologia , Pele/patologia , Linfócitos T/metabolismo , Vitiligo/sangue , Vitiligo/patologia
15.
Plant Physiol Biochem ; 140: 68-77, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31085448

RESUMO

Mo-CBP3 is a chitin-binding 2S albumin from Moringa oleifera. This seed storage protein is resistant to thermal denaturation and shows biological activities that might be of practical use, such as antifungal properties against Candida sp., a pathogen that causes candidiasis, and against Fusarium solani, a soil fungus that can cause diseases in plants and humans. Previous work has demonstrated that Mo-CBP3 is a mixture of isoforms encoded by members of a small multigene family. Mature Mo-CBP3 is a small protein (∼14 kDa), constituted by a small chain of approximately 4 kDa and a large chain of 8 kDa, which are held together by disulfide bridges. However, a more comprehensive picture on the spectrum of Mo-CBP3 isoforms which are found in mature seeds, is still lacking. In this work, genomic DNA fragments were obtained from M. oleifera leaves, cloned and completely sequenced, thus revealing new genes encoding Mo-CBP3. Moreover, mass spectrometry analysis showed that the mature protein is a complex mixture of isoforms with a remarkable number of molecular mass variants. Using computational predictions and calculations, most (∼86%) of the experimentally determined masses were assigned to amino acid sequences deduced from DNA fragments. The results suggested that the complex mixture of Mo-CBP3 isoforms originates from proteins encoded by closely related genes, whose products undergo different combinations of distinct post-translational modifications, including cleavage at the N- and C-terminal ends of both subunits, cyclization of N-terminal Gln, as well as Pro hydroxylation, Ser/Thr phosphorylation, and Met oxidation.


Assuntos
Moringa oleifera/química , Proteínas de Plantas/metabolismo , Isoformas de Proteínas/metabolismo , Humanos , Proteínas de Plantas/química , Isoformas de Proteínas/química , Processamento de Proteína Pós-Traducional
16.
Food Chem ; 290: 216-228, 2019 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-31000040

RESUMO

Blueberry is one of the richest phenolic sources, providing health benefits. To study blueberry phenolic biosynthesis, we investigated phenolics and proteomics at three typical fruit maturation phases. Multiple isoforms of enzymes and multiple members of transcription factors involved in phenolic biosynthesis were divergent and differently regulated. Regulation of some proteins resulted in change of phenolic content. During fruit maturation, down-regulation of VcOMT (CUFF.177.1) and VcLAR2 (CUFF.16780.1) was associated with decreases of ferulic acid and catechin, respectively; Up-regulation of VcFLS (CUFF.41155.1), and VcF3'5'H (CUFF.51711.1) and VcF3'5'H (gene.g10884.t1.1) likely drove increases of their products (quercetin and myricetin); Up-regulation of VcUFGALT (CUFF.20951.1) and VcUFGT73 (4333_g.1) and down-regulation of VcU5GT (CUFF.51258.1) were correlated to accumulation of anthocyanins with 3-glucoside/galactoside. Additionally, four TFs, VcAPRR2 (CUFF.24826.1), VcbHLH3 (CUFF.37765.1), VcWD (CUFF.28282.2) and VcWD (CUFF.28273.1) were probably related to regulation of anthocyanin biosynthesis. These proteins were potential targets for genetic improvement in a breeding program.


Assuntos
Mirtilos Azuis (Planta)/metabolismo , Fenóis/metabolismo , Proteínas de Plantas/metabolismo , Sequência de Aminoácidos , Antocianinas/análise , Antocianinas/metabolismo , Mirtilos Azuis (Planta)/química , Mirtilos Azuis (Planta)/genética , Cromatografia Líquida de Alta Pressão , Frutas/química , Frutas/genética , Frutas/metabolismo , Ontologia Genética , Peptídeos/análise , Proteínas de Plantas/química , Proteínas de Plantas/genética , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Alinhamento de Sequência , Espectrometria de Massas em Tandem , Regulação para Cima
17.
Plant Mol Biol ; 100(4-5): 379-390, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30968308

RESUMO

KEY MESSAGE: Functions of most splice isoforms that are generated by alternative splicing are unknown. We show that two splice variants that encode proteins differing in only eight amino acids have distinct functions in a stress response. Serine/arginine-rich (SR) and SR-like proteins, a conserved family of RNA binding proteins across eukaryotes, play important roles in pre-mRNA splicing and other post-transcriptional processes. Pre-mRNAs of SR and SR-like proteins undergo extensive alternative splicing in response to diverse stresses and produce multiple splice isoforms. However, the functions of most splice isoforms remain elusive. Alternative splicing of pre-mRNA of Arabidopsis SR45, which encodes an SR-like splicing regulator, generates two isoforms (long-SR45.1 and short-SR45.2). The proteins encoded by these two isoforms differ in eight amino acids. Here, we investigated the role of SR45 and its splice variants in salt stress tolerance. The loss of SR45 resulted in enhanced sensitivity to salt stress and changes in expression and splicing of genes involved in regulating salt stress response. Interestingly, only the long isoform (SR45.1) rescued the salt-sensitive phenotype as well as the altered gene expression and splicing patterns in the mutant. These results suggest that SR45 positively regulates salt tolerance. Furthermore, only the long isoform is required for SR45-mediated salt tolerance.


Assuntos
Proteínas de Arabidopsis/fisiologia , Arabidopsis/genética , Proteínas de Ligação a RNA/fisiologia , Tolerância ao Sal/genética , Processamento Alternativo , Arabidopsis/metabolismo , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Homeostase , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Isoformas de Proteínas/fisiologia , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Transdução de Sinais , Estresse Fisiológico
18.
Fish Shellfish Immunol ; 89: 701-709, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31004801

RESUMO

Based on the transcriptome database, we screened out four ferritin subunit genes (MnFer2-5) from the oriental river prawn Macrobrachium nipponense, which encode two non-secretory and two secretory peptides. MnFer2 and 4 possess a strictly conserved ferroxidase site, and MnFer3 has a non-typical ferroxidase site. MnFer5 seems to be a number of ferritin families, which has a distinct dinuclear metal binding motif, but lacks an iron ion channel, a ferroxidase site and a nucleation site. Diverse tissue-specific transcriptions of the four genes indicate their functional diversity in the prawn. Among them, MnFer2 is mainly expressed in hepatopancreas and intestines, MnFer3 and 4 are predominantly expressed in gills, and MnFer5 is widely expressed in various tissues with high presence in intestines, hepatopancreas and haemocytes. The transcription of all the four MnFer genes can be strongly induced by doxorubicin, indicating the involvement of these ferritin subunits in protection from oxidative stress. Upon Aeromonas hydrophila infection, only MnFer5 is persistently up-regulated, while other subunits including MnFer2-4 are down-regulated during the early stage, followed by recovery and even a slight increase at 48 h post bacterial challenge. Moreover, the iron binding capacity of recombinant MnFer2 is also demonstrated in vitro. The E. coli expressing MnFer2 displays increased resistance to hydrogen peroxidase cytotoxicity. These results suggest a protective role of ferritins from M. nipponense in iron homeostasis, redox biology and antibacterial immunity and shed light on the molecule evolution of crustacean ferritin subunits.


Assuntos
Ferritinas/genética , Ferritinas/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Estresse Oxidativo/imunologia , Penaeidae/genética , Penaeidae/imunologia , Sequência de Aminoácidos , Animais , Proteínas de Artrópodes/química , Proteínas de Artrópodes/genética , Proteínas de Artrópodes/imunologia , Sequência de Bases , Relação Dose-Resposta a Droga , Doxorrubicina/farmacologia , Ferritinas/química , Perfilação da Expressão Gênica , Homeostase/imunologia , Oxirredução , Palaemonidae , Filogenia , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Distribuição Aleatória , Alinhamento de Sequência
19.
Mol Biotechnol ; 61(6): 442-450, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30980224

RESUMO

Soil salinity imposes a serious threat to the productivity of agricultural crops. Among several other transporters, high-affinity K+ transporter (HKT)'s play an important role in reducing the phytotoxicity of Na+. Expression of Eutrema salsugineum (a halophyte) HKT1;2 is induced upon salt exposure. To elucidate the role of its promoter, we compared the sequences of HKT1;2 promoters from E. salsugineum (1822 bp) and E. botschantzevii (1811 bp) with Arabidopsis thaliana HKT1;1 (846 bp) promoter. In silico analysis predicted several cis-acting regulatory elements (GT-1 elements, core motifs of DRE/CRT, MYC/MYB-recognition sites and ACGT elements). Activities of the three promoters were analyzed by measuring HKT1;1 and/or HKT1;2 transcript level in the Athkt1;1 mutant plants. NaCl tolerance of the transgenics was also assessed. Our results depicted that expressing either AtHKT1;1 or EsHKT1;2 coding regions under the control of AtHKT1;1 promoter, almost reversed the hypersensitivity of the mutant for salt, on contrarily, when AtHKT1;1 coding sequence expressed under either Es or EbHKT1;2 promoters did not. Changes in shoot Na+/K+ concentrations under salt exposure is significantly consistent with the complementation ability of the mutant. The transcript concentration for genes under the control of either of Eutrema promoters, at control level was very less. This may suggest that either an important upstream response motif is missed or that A. thaliana misses a transcriptional regulator that is essential for salt-inducible HKT1 expression in Eutrema.


Assuntos
Arabidopsis/genética , Brassicaceae/genética , Proteínas de Transporte de Cátions/genética , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/genética , Tolerância ao Sal/genética , Simportadores/genética , Arabidopsis/efeitos dos fármacos , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Sequência de Bases , Brassicaceae/efeitos dos fármacos , Brassicaceae/crescimento & desenvolvimento , Brassicaceae/metabolismo , Proteínas de Transporte de Cátions/metabolismo , Teste de Complementação Genética , Transporte de Íons/efeitos dos fármacos , Mutação , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/genética , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/metabolismo , Proteínas de Plantas/metabolismo , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Brotos de Planta/efeitos dos fármacos , Brotos de Planta/genética , Brotos de Planta/crescimento & desenvolvimento , Brotos de Planta/metabolismo , Plantas Geneticamente Modificadas , Potássio/metabolismo , Regiões Promotoras Genéticas , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Alinhamento de Sequência , Sódio/metabolismo , Cloreto de Sódio/farmacologia , Especificidade da Espécie , Estresse Fisiológico/genética , Simportadores/metabolismo
20.
Int J Mol Sci ; 20(8)2019 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-31022851

RESUMO

Laccases are blue multi-copper oxidases with an extensive number of actual and potential industrial applications. It is known that laccases from different fungal strains may vary in properties; however, the reason of this remains unclear. In the current study we have isolated and characterized seven laccases from different strains of Steccherinum ochraceum obtained from regions of central Russia. Although all seven laccases had the same primary sequences, there was a little variation in their molecular weights and thermostabilities. Moreover, statistically significant differences in laccases' catalytic parameters of oxidation of phenolic substrates and ABTS were observed. After the deglycosylation of four selected laccases by Endo H and PNGase F, their affinities to pyrocatechol and ABTS became the same, suggesting a substantial role of N-linked glycosylation in moderation of enzymatic properties of laccases.


Assuntos
Proteínas Fúngicas/metabolismo , Lacase/metabolismo , Polyporales/enzimologia , Sequência de Aminoácidos , Proteínas Fúngicas/química , Glicosilação , Lacase/química , Modelos Moleculares , Polyporales/química , Polyporales/metabolismo , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismo , Especificidade por Substrato
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