1,721,443 research outputs found
Importin alpha binds to an unusual bipartite nuclear localization signal in the heterogeneous ribonucleoprotein type I
No full textThe heterogeneous nuclear ribonucleoprotein (hnRNP) type I, a modulator of alternative splicing, localizes in the nucleoplasm of mammalian cells and in a discrete perinucleolar structure. HnRNP I contains a novel type of bipartite nuclear localization signal (NLS) at the N-terminus of the protein that we have previously named nuclear determinant localization type I (NLD-I). Recently, a neural counterpart of hnRNP I has been identified that contains a putative NLS with two strings of basic amino acids separated by a spacer of 30 residues. In the present study we show that the neural hnRNP I NLS is necessary and sufficient for nuclear localization and represents a variant of the novel bipartite NLS present in the NLD-I domain. Furthermore, we demonstrate that the NLD-I is transported into the nucleus by cytoplasmic factor(s) with active transport modality. Binding assays using recombinant importin α show an interaction with NLD-I similar to that of SV40 large T antigen NLS. Deletion analysis indicates that both stretches of basic residues are necessary for binding to importin α. The above experimental results lead to the conclusion that importin α acts as cytoplasmic receptor for proteins characterized by a bipartite NLS signal that extends up to 37 residues.The heterogeneous nuclear ribonucleoprotein (hnRNP) type I, a modulator of alternative splicing, localizes in the nucleoplasm of mammalian cells and in a discrete perinucleolar structure. HnRNP I contains a novel type of bipartite nuclear localization signal (NLS) at the N-terminus of the protein that we have previously named nuclear determinant localization type I (NLD-I). Recently, a neural counterpart of hnRNP I has been identified that contains a putative NLS with two strings of basic amino acids separated by a spacer of 30 residues. In the present study we show that the neural hnRNP I NLS is necessary and sufficient for nuclear localization and represents a variant of the novel bipartite NLS present in the NLD-I domain. Furthermore, we demonstrate that the NLD-I is transported into the nucleus by cytoplasmic factor(s) with active transport modality. Binding assays using recombinant importin alpha show an interaction with NLD-I similar to that of SV40 large T antigen NLS. Deletion analysis indicates that both stretches of basic residues are necessary for binding to importin alpha. The above experimental results lead to the conclusion that importin alpha acts as cytoplasmic receptor for proteins characterized by a bipartite NLS signal that extends up to 37 residues
Post-transcriptional Gene Regulation: Role of the Polypyrimidine Tract-binding Protein (PTB) and Associated Factors in "From the hallowed halls of Herpesvirology".
No full textThe overall processes of gene expression, from mRNA biogenesis to translation and degradation, require RNA messenger ribonucleoprotein (mRNP) complexes assembly, differentially combined with individual transcripts in order to mediate the post-transcriptional events. One of those events, the alternative splicing of pre-mRNA, is responsible for the wide variety and complexity levels of transcriptome and proteome. Large-scale expressed sequence tag and genome-wide analyses estimate that more than 60% of human genes undergo alternative splicing with a differential tissue distribution. More than 15% of human genetic diseases are associated to mutations in the consensus splice sites and disruption of splicing regulatory networks contributes to various diseases, including cancer. Among the best characterized splicing regulators is the polypyrimidine tract binding protein (PTB) and its paralog nPTB. Highly conserved in vertebrates, PTB has been implicated in several cellular process related to post-transcriptional regulation. In addition to the alternative splicing regulation, PTB is involved into polyadenylation, mRNA stability and initiation of translation. For the latter, several studies have described the PTB recruitment in IRES-mediated translation initiation or propagation of several viruses including hepatitis C virus and picornaviruses. To perform these functions PTB is regulated in its intracellular localization, shuttling between nucleus and cytoplasm. Tissue specific PTB isoforms and co-regulators, the ribonucleoproteins Raver 1 and 2, have been recently identified. Mechanisms of post-transcriptional regulation mediated by PTB, nPTB and Raver proteins are described in the present review
Biopolymer production from agro-industrial wastes: bacterial strains selection and genetic improvement.
Full text linkThis work is included in a wider research program carried out at the Dipartimento di Agronomia Animali Alimenti Risorse Naturali e Ambiente (DAFNAE) within the EU project ANIMPOL (Biotechnological conversion of carbon containing wastes for eco‐efficient production of high added value products). The aim of the entire project was to produce high-added-value products using inexpensive agricultural by-products as carbon source. The fatty by-products from slaughterhouses have been adopted for its conversion into biopolymers such as polyhydroxyalkanoates (PHA) by properly selected and/or developed microbes.
In more detail the objective of this research was to look for, select and characterise bacterial strains able to utilise, as carbon source, low cost industrial lipid wastes such as triacylglycerols (TAGs) from animal fats, with the final goal to produce PHAs. For this reason lipolytic bacteria were isolated from a variety of different environments, such as soil or waste water of a slaughterhouse. Several bacterial strains were found to possess remarkable lipolytic activities but not efficient PHAs production capabilities.
As a consequence, a molecular biology program started in order to obtain a microbial strain capable of both hydrolysing lipids and producing high levels of PHAs.
Delftia acidovorans DSM39, a well-known producer of PHAs with high molar fractions of 4-hydroxybutyrate (4HB), although unable to metabolize lipids, was selected as host strain. On the other hand, Pseudomonas stutzeri BT3, the most efficient lipase producer among the obtained isolates, was designated as potential donor of lipolytic genes.
The lipC and lipH sequences of P. stutzeri BT3 were successfully co-expressed into D. acidovorans DSM39 and the resulting recombinant strain displayed high extracellular enzymatic activity on corn oil. The PHAs production from corn oil achieved high levels (26% of cell dry weight, with about 7% of 4HB). Surprisingly, the recombinant strain produced greater values directly from slaughterhouse residues such as udder and lard (43 and 39%, respectively, with almost 7% of 4HB).
Moreover, this work proved the ability of the recombinant D. acidovorans DSM39 strain to produce PHAs with significant percentage of 4HB, without the supplementation of any precursor in the liquid broth. This research paves the way to the efficient one-step conversion of fatty residues into PHAs having valuable properties exploitable in several medical and industrial applications
Functional characterization of the human ribonucleoprotein, PTB-binding 2/Raver2 promoter region.
No full textStudio delle proteine nucleari hnRNP. Applicabilita’ del nuovo gene marcatore in grado di esprimere la “Green Fluorescent Protein” (GFP) ad aumentata fluorescenza
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Identification and analysis of the human neural polypyrimidine tract binding protein (nPTB) gene promoter region.
No full textNeural polypyrimidine tract-binding protein nPTB, originally identified as the neuronal counterpart of the hnRNPI/PTB protein, is an RNA binding protein involved into tissue-specific alternative splicing regulation. Here we describe the functional characterization of the promoter sequence of nPTB in HeLa and neuroblastoma cells. By means of genomic sequence analysis we have isolated and cloned a 1587-base pair region upstream the human nPTB coding region. The nPTB proximal promoter, although rich in G+C content and presenting putative binding sites for the transcription factors Sp1, NF-1, NF-kB and Oct-1, lacks a typical TATA box. Luciferase transient expression assays using deletion mutants have identified the proximal promoter region at -125 relative to the transcription start site. Alignment of human, murine and chimpanzee genomic sequences upstream the nPTB exon 1 has provided evidences for the evolutionary conservation of specific transcription factor binding sites
Raver1 expression and alternative splicing events during skeletal muscle differentiation
No full textCoinvolgimento della DNA polimerasi cellulare nella replicazione dei virus herpes simplex di tipo 1.
No full textExpression of the late (g2) genes in herpes simplex virus 1 (HSV-1). Analysis of the binding sites for the major viral regulatory portein, ICP 4, in the 5' transcribed non coding domain of a late gene
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