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1.
Mol Psychiatry ; 21(2): 178-88, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26347316

RESUMO

Increased dosage of methyl-CpG-binding protein-2 (MeCP2) results in a dramatic neurodevelopmental phenotype with onset at birth. We generated induced pluripotent stem cells (iPSCs) from patients with the MECP2 duplication syndrome (MECP2dup), carrying different duplication sizes, to study the impact of increased MeCP2 dosage in human neurons. We show that cortical neurons derived from these different MECP2dup iPSC lines have increased synaptogenesis and dendritic complexity. In addition, using multi-electrodes arrays, we show that neuronal network synchronization was altered in MECP2dup-derived neurons. Given MeCP2 functions at the epigenetic level, we tested whether these alterations were reversible using a library of compounds with defined activity on epigenetic pathways. One histone deacetylase inhibitor, NCH-51, was validated as a potential clinical candidate. Interestingly, this compound has never been considered before as a therapeutic alternative for neurological disorders. Our model recapitulates early stages of the human MECP2 duplication syndrome and represents a promising cellular tool to facilitate therapeutic drug screening for severe neurodevelopmental disorders.


Assuntos
Proteína 2 de Ligação a Metil-CpG/genética , Proteína 2 de Ligação a Metil-CpG/fisiologia , Rede Nervosa/metabolismo , Diferenciação Celular , Dendritos/metabolismo , Dosagem de Genes/fisiologia , Duplicação Gênica/genética , Estudos de Associação Genética , Humanos , Células-Tronco Pluripotentes Induzidas , Masculino , Neurogênese , Neurônios
2.
Neurobiol Learn Mem ; 124: 71-87, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26220900

RESUMO

Autism Spectrum Disorders are a heterogeneous group of neurodevelopmental disorders, with rising incidence but little effective therapeutic intervention available. Currently two main clinical features are described to diagnose ASDs: impaired social interaction and communication, and repetitive behaviors. Much work has focused on understanding underlying causes of ASD by generating animal models of the disease, in the hope of discovering signaling pathways and cellular targets for drug intervention. Here we review how ASD behavioral phenotypes can be modeled in the mouse, the most common animal model currently in use in this field, and discuss examples of genetic mouse models of ASD with behavioral features that recapitulate various symptoms of ASD.


Assuntos
Transtorno do Espectro Autista/genética , Modelos Animais de Doenças , Pesquisa Translacional Biomédica , Agressão/fisiologia , Animais , Transtorno do Espectro Autista/psicologia , Comportamento Compulsivo/genética , Humanos , Relações Interpessoais , Transtornos da Memória/genética , Camundongos , Atividade Motora/genética , Comportamento Obsessivo/genética , Fenótipo , Transdução de Sinais , Vocalização Animal/fisiologia
3.
PLoS One ; 18(6): e0287062, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37379311

RESUMO

Breast cancer is the most common cancer among women worldwide. Molecular and clinical evidence indicated that Fragile X Messenger Ribonucleoprotein 1 (FMRP) plays a role in different types of cancer, including breast cancer. FMRP is an RNA binding protein that regulates the metabolism of a large group of mRNAs coding for proteins involved in both neural processes and in epithelial-mesenchymal transition, a pivotal mechanism that in cancer is associated to tumor progression, aggressiveness and chemoresistance. Here, we carried out a retrospective case-control study of 127 patients, to study the expression of FMRP and its correlation with metastasis formation in breast cancer. Consistent with previous findings, we found that FMRP levels are high in tumor tissue. Two categories have been analyzed, tumor with no metastases (referred as control tumors, 84 patients) and tumor with distant metastatic repetition, (referred as cases, 43 patients), with a follow-up of 7 years (mean). We found that FMRP levels were lower in both the nuclei and the cytoplasm in the cases compared to control tumors. Next, within the category cases (tumor with metastases) we evaluated FMRP expression in the specific sites of metastasis revealing a nuclear staining of FMRP. In addition, FMRP expression in both the nuclear and cytoplasmic compartment was significantly lower in patients who developed brain and bone metastases and higher in hepatic and pulmonary sites. While further studies are required to explore the underlying molecular mechanisms of FMRP expression and direct or inverse correlation with the secondary metastatic site, our findings suggest that FMRP levels might be considered a prognostic factor for site-specific metastasis.


Assuntos
Neoplasias da Mama , Síndrome do Cromossomo X Frágil , Neoplasias Mamárias Animais , Animais , Humanos , Feminino , Proteína do X Frágil da Deficiência Intelectual/genética , Proteína do X Frágil da Deficiência Intelectual/metabolismo , Estudos de Casos e Controles , Estudos Retrospectivos , Proteínas/metabolismo , Neoplasias da Mama/genética , Síndrome do Cromossomo X Frágil/genética
4.
Eur Neuropsychopharmacol ; 48: 49-88, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33781629

RESUMO

In the last decade there has been a revolution in terms of genetic findings in neurodevelopmental disorders (NDDs), with many discoveries critical for understanding their aetiology and pathophysiology. Clinical trials in single-gene disorders such as fragile X syndrome highlight the challenges of investigating new drug targets in NDDs. Incorporating a developmental perspective into the process of drug development for NDDs could help to overcome some of the current difficulties in identifying and testing new treatments. This paper provides a summary of the proceedings of the 'New Frontiers Meeting' on neurodevelopmental disorders organised by the European College of Neuropsychopharmacology in conjunction with the Innovative Medicines Initiative-sponsored AIMS-2-TRIALS consortium. It brought together experts in developmental genetics, autism, NDDs, and clinical trials from academia and industry, regulators, patient and family associations, and other stakeholders. The meeting sought to provide a platform for focused communication on scientific insights, challenges, and methodologies that might be applicable to the development of CNS treatments from a neurodevelopmental perspective. Multidisciplinary translational consortia to develop basic and clinical research in parallel could be pivotal to advance knowledge in the field. Although implementation of clinical trials for NDDs in paediatric populations is widely acknowledged as essential, safety concerns should guide each aspect of their design. Industry and academia should join forces to improve knowledge of the biology of brain development, identify the optimal timing of interventions, and translate these findings into new drugs, allowing for the needs of users and families, with support from regulatory agencies.


Assuntos
Transtorno Autístico , Transtornos do Neurodesenvolvimento , Criança , Descoberta de Drogas/métodos , Humanos , Transtornos do Neurodesenvolvimento/tratamento farmacológico , Transtornos do Neurodesenvolvimento/genética
5.
Neuroscience ; 151(3): 791-801, 2008 Feb 06.
Artigo em Inglês | MEDLINE | ID: mdl-18178317

RESUMO

Ionotropic purinergic receptors (P2XR) are ATP-gated cationic channels composed of seven known subunits (P2X(1-7)R) and involved in different functions in neural tissue. Although their presence has been demonstrated in the brain, few studies have investigated their expression pattern. In particular, ionotropic purinergic receptor subunit type 1 (P2X(1)R) has been observed in the cerebellum and in brainstem nuclei. The present study investigates the P2X(1)R expression pattern in the rat forebrain using immunohistochemistry. The specificity of the immunolabeling has been verified by Western blotting and in situ hybridization methods. P2X(1)R immunoreactivity was specifically localized in neurons, dendrites and axons throughout the forebrain. Characteristic differences in the distribution of P2X(1)R were observed in different cortical areas. In prefrontal, cingulate and perirhinal cortices, very intense labeling was present in neuronal bodies. In frontal, parietal, temporal and occipital cortices, immunostaining was lighter and mainly found in dendrites and axons. The hippocampal formation was intensely labeled. Labeling was present almost exclusively in dendrites and axons and never in neuronal bodies. The diencephalon was devoid of P2X(1)R positive neurons or fibers except for the medial habenular nucleus, which showed very intense P2X(1)R immunostaining. Furthermore, two subcortical regions, namely, the nucleus centralis of the amygdala and the bed nucleus of the stria terminalis, showed intense P2X(1)R neuronal labeling. Present data indicate that P2X(1)R are prevalent in forebrain areas involved in the integration of cognitive, limbic and autonomic functions.


Assuntos
Neurônios/metabolismo , Prosencéfalo/citologia , Receptores Purinérgicos P2/metabolismo , Animais , Proteína Glial Fibrilar Ácida/metabolismo , Masculino , Neurônios/ultraestrutura , Fosfopiruvato Hidratase/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Receptores Purinérgicos P2/genética , Receptores Purinérgicos P2X
6.
Oncogene ; 20(33): 4596-600, 2001 Jul 27.
Artigo em Inglês | MEDLINE | ID: mdl-11494157

RESUMO

Mutations in the 5' UTR which cause increment/decrement of translation efficiency have been recently described as a novel molecular mechanism of disease. Alterations in the consensus sequence for the translation initiation may promote context-dependent leaky scanning of ribosomes and/or initiation from a downstream AUG codon. Initiation of translation from a downstream in-frame AUG codon in BRCA1 gene was recently identified in normal cells and possibly in breast cancer. Here we present further insight into BRCA1 translational pathophysiology investigating the role of the canonical structure of the initiation consensus sequence of BRCA1. We have analysed the effect of a somatic point mutation (117 G>C) in position -3 with respect to the AUG of the BRCA1 gene, identified in a highly aggressive sporadic breast cancer. We constructed chimeric genes encoding the luciferase reporter sequence downstream of the wild type or the mutated BRCA1 5'UTR. These transcripts were tested for their activity in in vitro and in vivo systems. In in vitro transcription/translation assays the estimated translation efficiency of the construct with the mutated BRCA1 5'UTR was 30-50% lower than that with the wild type BRCA1 5'UTR. The same chimeric genes were analysed for their expression in vivo by transient transfection in human cells. While the two constructs were equally transcribed, the plasmid carrying the mutated sequence produced 70% less luciferase activity compared to the wild type sequence. Finally, to obtain a direct evaluation on translational efficiency in vivo, we analysed mRNA translation on translationally active and non-active ribosomes separated from transfected cells. Mutant mRNA was partially localized in subpolysomal particles analytically confirming a polysome recruitment defect. Thus, characterization of BRCA1 5'UTR and translation efficiency seems to provide new insight into BRCA1 role in breast and ovarian cancer pathogenesis.


Assuntos
Regiões 5' não Traduzidas/genética , Neoplasias da Mama/genética , Carcinoma/genética , Genes BRCA1 , Mutação , Biossíntese de Proteínas , Bacteriófago T7/genética , Linhagem Celular , Sistema Livre de Células , Sequência Consenso , Feminino , Genes Reporter , Genes Sintéticos , Humanos , Rim , Luciferases/biossíntese , Luciferases/genética , Iniciação Traducional da Cadeia Peptídica/genética , Regiões Promotoras Genéticas , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/genética
7.
J Neurosci ; 20(10): RC76, 2000 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-10783400

RESUMO

The presence of specific mRNAs in dendrites and at synapses is well established, but a direct and reliable demonstration that they are associated with polysomes is still missing. To address this point we analyzed the polysomal association of the mRNAs for the alpha-subunit of Ca(2+)/calmodulin-dependent protein kinase II (alpha-CaMKII), for type 1 inositol 1,4,5-trisphosphate receptor (InsP3R1) and for the activity-regulated cytoskeleton-associated protein (Arc) in a synaptosomal preparation devoid of contaminating material from neuronal and glial perikarya. We show that a fraction of alpha-CaMKII, InsP3R1, and Arc mRNAs present in synaptosomes is indeed associated with polysomes. Moreover, we show that polysomal association of alpha-CaMKII mRNA, but not InsP3R1 and Arc mRNAs, increases with depolarization of the synaptosomal membrane. Finally, we show that the synthesis of alpha-CaMKII protein increases with stimulation. Dendritic mRNA recruitment onto polysomes in response to synaptic stimulation might represent one of the mechanisms underlying the processes of learning and memory.


Assuntos
Proteínas Quinases Dependentes de Cálcio-Calmodulina/metabolismo , Polirribossomos/metabolismo , RNA Mensageiro/metabolismo , Sinaptossomos/metabolismo , Animais , Encéfalo/ultraestrutura , Canais de Cálcio/metabolismo , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina , Proteínas do Citoesqueleto/metabolismo , Técnicas In Vitro , Receptores de Inositol 1,4,5-Trifosfato , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Eletrônica , Proteínas do Tecido Nervoso/metabolismo , Polirribossomos/enzimologia , Cloreto de Potássio/farmacologia , Receptores Citoplasmáticos e Nucleares/metabolismo , Estimulação Química , Sinaptossomos/enzimologia
8.
Biochim Biophys Acta ; 1216(3): 475-8, 1993 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-8268230

RESUMO

The cloning and sequencing of a cDNA for human ribosomal protein L4 is reported. The corresponding mRNA has a very short 5' untranslated region initiating with a sequence of 12 pyrimidines, characteristic of all vertebrate ribosomal protein mRNAs. The deduced amino acid sequence shows that human ribosomal protein L4 has 425 amino acid residues and a calculated molecular mass of 47,821 Da. Comparison with the homologous counterparts of Xenopus, Drosophila and yeast shows that this protein has a very conserved amino-terminus region and an extremely divergent carboxyl-terminus portion.


Assuntos
Proteínas Ribossômicas/biossíntese , Proteínas Ribossômicas/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , DNA Complementar/química , DNA Complementar/metabolismo , Drosophila melanogaster/genética , Humanos , Dados de Sequência Molecular , Saccharomyces cerevisiae/genética , Homologia de Sequência de Aminoácidos , Xenopus laevis
9.
Gene ; 67(1): 69-74, 1988 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-2843441

RESUMO

Recombinant clones specific for ribosomal protein (r-protein) S8 have been isolated from a Xenopus laevis cDNA bank. Sequence analysis shows that they are of two types, derived from two different gene copies originating from gene duplication. The two cDNAs differ in several silent sites and code for the same S8 protein whose complete amino acid sequence has been derived. Sequence comparison of S8 mRNAs with those for other X. laevis r-proteins, has revealed interesting similarities in the 5' and 3' untranslated regions. These could be involved in r-protein synthesis regulation which we have previously shown to occur mainly at post-transcriptional and translational levels.


Assuntos
DNA/isolamento & purificação , RNA Mensageiro/genética , Proteínas Ribossômicas/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , DNA/genética , Enzimas de Restrição do DNA , Genes , Dados de Sequência Molecular , Xenopus laevis
10.
Gene ; 119(2): 283-6, 1992 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-1398110

RESUMO

The genes coding for ribosomal proteins (r-proteins) in Xenopus laevis have a stretch of about 20 pyrimidines (Y) at their 5' end, in the middle of which is localized the main transcription start point (tsp). To obtain information about its possible functional significance, we have introduced by site-directed mutagenesis one or more purines at various positions within the oligo(Y) tract present at the 5' end of the gene coding for r-protein S19 of X. laevis. The effect of these mutations on transcription and translation of a reporter-coding sequence has been evaluated by DNA microinjection in X. laevis oocytes. We show that an uninterrupted stretch of pyrimidines is not necessary for efficient transcription and translation of the reporter gene in the X. laevis oocyte. This finding does not exclude the possibility that this sequence is involved in transcription and/or translation regulation in some developmental situations different from oogenesis.


Assuntos
Oócitos/metabolismo , Pirimidinas , Proteínas Ribossômicas/genética , Transcrição Gênica , Animais , Sequência de Bases , Clonagem Molecular , DNA , Regulação da Expressão Gênica/genética , Microinjeções , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Xenopus laevis
11.
Gene ; 132(2): 255-60, 1993 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-8224872

RESUMO

We present here the cloning and the entire sequence of one of the two gene copies coding for ribosomal protein (r-protein) S8 in Xenopus laevis (corresponding to r-protein S7 in rat) and its flanking regions. The S8a gene contains seven exons and six introns for a total length of about 12,700 bp coding for a mRNA of 663 nucleotides (nt) plus a poly(A) tail. Mapping of the 5' end of the gene has shown that the transcription start point is located in a pyrimidine-rich tract, as has been observed for all r-protein-encoding genes of X. laevis and other vertebrates so far characterized. A computer analysis of the S8a sequence has revealed the presence of a 220-nt sequence repeated, with some variations, once in each of the six introns. RNA analysis by hybridization with oligo probes specific for the two gene copies coding for r-protein S8 has demonstrated that the two of them are expressed at similar levels both in oocytes and in embryos.


Assuntos
Proteínas Ribossômicas/genética , Xenopus laevis/genética , Animais , Sequência de Bases , Southern Blotting , Clonagem Molecular , DNA , Éxons , Íntrons , Dados de Sequência Molecular , Mapeamento por Restrição
12.
Hernia ; 5(3): 124-8, 2001 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-11759796

RESUMO

Giant incisional hernias with total loss of substance are an ominous pathological condition characterized by massive depletion of muscular and fascial tissue, by complete loss of the anatomical and physiological function of the abdominal wall and by severe respiratory and visceral involvement. Over a 10-year period we operated 270 patients with voluminous incisional hernias, 12 of which had a total loss of substance. There was no intraoperative mortality. One patient died of myocardial infarction on the fifth and one died of intestinal occlusion and peritonitis the 11th postoperative day. Early postoperative complications occurred in only one patient who had skin necrosis with an infection at the polypropylene mesh. This was successfully treated with systemic antibiotic therapy and topical medication of the wound. There was also one minor recurrence over the pubis 1 year after the operation that required a new operation to replace the mesh. No respiratory complications occurred and all patients were normally active. The good results reported in our series encourage us to continue in this direction even though these patients are at high risk.


Assuntos
Hérnia Ventral/cirurgia , Complicações Pós-Operatórias/cirurgia , Próteses e Implantes , Telas Cirúrgicas , Idoso , Idoso de 80 Anos ou mais , Feminino , Seguimentos , Hérnia Ventral/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Infarto do Miocárdio/etiologia , Peritonite/etiologia , Complicações Pós-Operatórias/patologia , Infecções Cutâneas Estafilocócicas/etiologia , Staphylococcus aureus/isolamento & purificação , Telas Cirúrgicas/microbiologia
13.
Minerva Med ; 75(37): 2105-9, 1984 Sep 29.
Artigo em Italiano | MEDLINE | ID: mdl-6483265

RESUMO

Intra-operative esophageal manometric evaluations are examined. The data refer to: extramucosal myotomy by thoracotomy; hiatal hernia repair by laparatomy; pharingo-crico-myotomy by cervicotomy. The utility of this intra-operative measurement clearly appears, especially in order to an immediate evaluation of surgical correction.


Assuntos
Acalasia Esofágica/cirurgia , Hérnia Diafragmática/cirurgia , Hérnia Hiatal/cirurgia , Manometria/métodos , Esôfago/fisiopatologia , Humanos , Período Intraoperatório , Pressão
14.
Minerva Gastroenterol Dietol ; 46(1): 19-22, 2000 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-16498346

RESUMO

BACKGROUND: In this retrospective investigation the symptoms, signs, and laboratory findings collected in 2 groups of patients with simple and complicated acute appendicitis, respectively, have been observed in order to give some indication for a correct diagnosis and surgical treatment. METHODS: A total of 103 consecutive patients affected by simple and complicated acute appendicitis submitted to surgical operation have been studied. RESULTS: Data collected show statistically significant differences between clinical presentation of simple and complicated acute appendicitis. CONCLUSIONS: The conclusion is draws in that anamnesis and clinical examination of the patients affected by acute appendicitis are the best indications for an exact diagnosis and to select patients who need an immediate operation.

17.
Oncogene ; 28(43): 3814-24, 2009 Oct 29.
Artigo em Inglês | MEDLINE | ID: mdl-19668229

RESUMO

Reticulons are a family of highly conserved proteins, localized in the endoplasmic reticulum (ER) and involved in different cellular functions, such as intracellular membrane trafficking, apoptosis and nuclear envelope formation. The reticulon protein family consists of four members, but their specific functions are presently poorly understood. RTN-1C overexpression triggers apoptosis, regulating ER stress versus DNA damage-induced cell death in a mutually exclusive way. The different RTN isoforms share a C-terminal reticulon homology domain containing two hydrophobic segments and a 66-amino acid hydrophilic loop. In the C-terminal region of RTN-1C, a unique consensus sequence (GAKRH) has recently been identified, showing 100% identity with the DNA-binding domain of histone H4. In this study, we show that this sequence is essential for RTN-1C-mediated apoptosis. It is noteworthy that the lysine 204 present in this region is post-translationally modified by acetylation and that this event is associated with a significant decrease in histone deacetylase activity and contributes to RTN-1C binding to DNA. These data demonstrate a molecular mechanism by which RTN-1C controls apoptosis and indicate this protein to be a novel potential target for cancer therapy.


Assuntos
Retículo Endoplasmático/metabolismo , Inibidores de Histona Desacetilases , Proteínas do Tecido Nervoso/fisiologia , Tumores Neuroectodérmicos/metabolismo , Acetilação , Apoptose , Linhagem Celular Tumoral , DNA/metabolismo , Humanos , Proteínas do Tecido Nervoso/química
18.
Mol Cell Biochem ; 272(1-2): 19-28, 2005 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-16010968

RESUMO

Lysophosphatidic acid (LPA) is a lipid mediator with multiple biological functions. In the present study we investigated the possible role of atrial natriuretic peptide (ANP), a hormone affecting cardiovascular homeostasis and inducing antimitogenic effects in different cell types, on LPA-induced cell growth and reactive oxygen species (ROS) production in rat aortic smooth muscle (RASM) cells. Both LPA effects on cell growth and levels of ROS were totally abrogated by physiological concentrations of ANP, without modifying the overexpression of LPA-receptors. These effects were also affected by cell pretreatment with wortmannin, an inhibitor of phosphatidylinositol 3-kinase (PI3K). Moreover, the LPA-induced activation of Akt, a downstream target of PI3K, was completely inhibited by physiological concentrations of ANP, which were also able to inhibit p42/p44 phosphorylation. Taken together, our data suggest that PI3K may represent an important step in the LPA signal transduction pathway responsible for ROS generation and DNA synthesis in RASM cells. At same time, the enzyme could also represent an essential target for the antiproliferative effects of ANP.


Assuntos
Fator Natriurético Atrial/fisiologia , Lisofosfolipídeos/antagonistas & inibidores , Músculo Liso Vascular/efeitos dos fármacos , Androstadienos/farmacologia , Animais , Aorta/citologia , Fator Natriurético Atrial/farmacologia , Células Cultivadas , Replicação do DNA/efeitos dos fármacos , Ativação Enzimática , Lisofosfolipídeos/farmacologia , Masculino , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Músculo Liso Vascular/citologia , Músculo Liso Vascular/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/agonistas , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-akt , Ratos , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo , Receptores de Ácidos Lisofosfatídicos/genética , Receptores de Ácidos Lisofosfatídicos/metabolismo , Transdução de Sinais , Wortmanina
19.
J Biol Chem ; 273(18): 10868-73, 1998 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-9556561

RESUMO

The nucleolar proteins Gar1p and fibrillarin possess a typical nucleolar glycine/arginine-rich domain and belong to ribonucleoprotein particles. Both proteins are essential for yeast cell growth and are required for pre-rRNA processing. In addition, Gar1p is involved in pre-rRNA pseudouridylation, whereas fibrillarin is required for pre-rRNA methylation. Gar1p and fibrillarin are each associated with a different subset of the small nucleolar RNAs (snoRNAs). Gar1p is co-immunoprecipitated with the H/ACA family of snoRNAs, whereas fibrillarin is co-immunoprecipitated with the C/D family. However, attempts to demonstrate direct interactions between fibrillarin and snoRNAs have failed, and such interactions between Gar1p and the H/ACA snoRNAs had not yet been reported. Among the H/ACA snoRNAs associated with Gar1p, one can distinguish a large group of snoRNAs that are not essential in yeast and serve as guides for pseudouridine synthesis onto the pre-rRNA molecule. In contrast, the two snoRNAs snR10 and snR30 are required for normal cell growth and for pre-rRNA cleavage. We show here that Gar1p interacts in vitro directly and specifically with these two snoRNAs. Deletion analysis of Gar1p indicates that a major RNA binding element, which is extremely well conserved throughout evolution, lies in the middle of the protein. However, this domain alone binds poorly to the target RNAs and an accessory domain is required to restore efficient binding. The accessory domain can be either one of the glycine/arginine-rich domains or a second element of the core of the protein that is highly conserved between different species.


Assuntos
Proteínas Fúngicas/metabolismo , Proteínas Nucleares/metabolismo , RNA Nuclear Pequeno/metabolismo , Proteínas de Ligação a RNA/metabolismo , Ribonucleoproteínas Nucleolares Pequenas , Proteínas de Saccharomyces cerevisiae , Sequência de Aminoácidos , Proteínas Fúngicas/química , Dados de Sequência Molecular , Proteínas Nucleares/química , Ligação Proteica , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Deleção de Sequência , Homologia de Sequência de Aminoácidos
20.
Mol Gen Genet ; 234(1): 60-4, 1992 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-1495485

RESUMO

Ribosomal protein synthesis is regulated by controlling the fraction of mRNA associated with polysomes. It is known that this value changes in different developmental stages during Xenopus embryogenesis or, more generally, with changing cell growth conditions. We present here an analysis of the proportion of mRNA loaded on polysomes, carried out with probes for five different ribosomal proteins on several batches of Xenopus embryos obtained from different individuals. The results obtained indicate the existence of probe-dependent and individual differences, which reflect genetic variations in the cis- and trans-acting regulatory elements responsible for translational regulation. The fraction of ribosomal protein mRNA loaded onto polysomes can be used as an index of an individual's capacity for ribosome production.


Assuntos
Regulação da Expressão Gênica , Variação Genética , Biossíntese de Proteínas , Proteínas Ribossômicas/biossíntese , Animais , Autorradiografia , Polirribossomos/metabolismo , RNA Mensageiro/genética , Proteínas Ribossômicas/genética , Xenopus laevis
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