Nucleolin
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Nucleolin is the major nucleolar protein of animal, plant and yeast proliferating cells. It is enriched in the most soluble nuclear or nucleolar protein extract, containing ribonucleoproteins, from which it has been purified. It has a tripartite structure in which each domain accounts for different functions. Despite its multifunctionality, the best characterized role of nucleolin is in the primary cleavage of pre-rRNA, an early step of ribosome biogenesis. In the nucleolus of proliferating cells, nucleolin is mostly located in the dense fibrillar component, following a vectorial pattern, from the periphery of fibrillar centers outwards. This pattern is lost in quiescent cells in which nucleolin is present in low levels. Nucleolin is the most phosphorylated protein of the soluble nuclear extract. It is phosphorylated by casein kinase II and CDKA, and phosphorylation is closely associated with the role of nucleolin in proliferating cells. During mitosis, nucleolin is transported from the mother to the daughter cell nucleolus in the form of processing particles, together with pre-rRNA precursors and other nucleolar proteins. It forms part of prenucleolar bodies and plays a role in nucleologenesis. Recent studies on the nucleolin function, carried out on samples with inactivated nucleolin genes (siRNA downregulated or mutants) have evidenced that nucleolin is absolutely essential for cell proliferation, for the organization of the nucleolus and for transcription and processing of pre-rRNA. In plants, nucleolin controls the auxin responsiveness, thus being involved in the regulation of plant development.
- MeSH
- biogeneze organel MeSH
- buněčný cyklus fyziologie genetika MeSH
- fosfoproteiny biosyntéza genetika chemie MeSH
- fosforylace fyziologie genetika MeSH
- jaderné proteiny biosyntéza genetika chemie MeSH
- klinické laboratorní techniky využití MeSH
- kvasinky MeSH
- kyseliny indoloctové MeSH
- prekurzory RNA biosyntéza genetika chemie MeSH
- proliferace buněk MeSH
- proteiny vázající RNA biosyntéza genetika chemie MeSH
- ribonukleoproteiny malé jadérkové biosyntéza MeSH
- ribozomy genetika chemie MeSH
- rostliny MeSH
- statistika jako téma MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- přehledy MeSH
57 s. : il. ; 30 cm
A high level of nucleolin (NCL) expression is often associated with a poor prognosis of patients with lung cancer (LC), suggesting that NCL can be used as a possible biomarker. NCL has been shown to display a marked preference for the binding to G-quadruplexes (G4). Here, we investigate the formation of an RNA quadruplex structure in a sequence found in the human precursor pre-MIR150 with the potential to recognize NCL. Circular dichroism (CD) spectra of pre-MIR150 G4-forming sequence (designated by rG4) indicate the formation of a parallel quadruplex structure in KCl or when complexed with the well-known G4 ligand PhenDC3. The thermal stability of rG4 is very high, and further increases in the presence of PhenDC3. The binding affinities of rG4 to PhenDC3 and NCL RBD1,2 are similar with KD values in the nanomolar range. PAGE results suggest the formation of a ternary quadruplex-ligand-protein complex (rG4-PhenDC3-NCL RBD1,2), indicative that PhenDC3 does not prevent the binding of rG4 to NCL RBD1,2. Finally, rG4 can recognize NCL-positive cells and, when fluorescently labeled, can be used as a probe for this protein. ELISA experiments indicate altered NCL expression patterns in liquid biopsies of LC patients in a non-invasive manner, potentially helping the diagnosis, prognosis, and patient response to treatment. Hence, labeled rG4 could be used as a detection probe of LC in liquid biopsies.
- MeSH
- aminokyselinové motivy fyziologie MeSH
- dospělí MeSH
- fosfoproteiny biosyntéza genetika MeSH
- G-kvadruplexy * MeSH
- genový targeting metody MeSH
- kultivované buňky MeSH
- leukocyty mononukleární metabolismus MeSH
- lidé MeSH
- nádory plic genetika metabolismus terapie MeSH
- proteiny vázající RNA biosyntéza genetika MeSH
- regulace genové exprese u nádorů MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Nucleolin is a multifunctional RNA Binding Protein (RBP) with diverse subcellular localizations, including the nucleolus in all eukaryotic cells, the plasma membrane in tumor cells, and the axon in neurons. Here we show that the glycine arginine rich (GAR) domain of nucleolin drives subcellular localization via protein-protein interactions with a kinesin light chain. In addition, GAR sequences mediate plasma membrane interactions of nucleolin. Both these modalities are in addition to the already reported involvement of the GAR domain in liquid-liquid phase separation in the nucleolus. Nucleolin transport to axons requires the GAR domain, and heterozygous GAR deletion mice reveal reduced axonal localization of nucleolin cargo mRNAs and enhanced sensory neuron growth. Thus, the GAR domain governs axonal transport of a growth controlling RNA-RBP complex in neurons, and is a versatile localization determinant for different subcellular compartments. Localization determination by GAR domains may explain why GAR mutants in diverse RBPs are associated with neurodegenerative disease.
- MeSH
- axonální transport genetika MeSH
- buněčné jadérko metabolismus ultrastruktura MeSH
- exprese genu MeSH
- fosfoproteiny chemie genetika metabolismus MeSH
- HEK293 buňky MeSH
- HeLa buňky MeSH
- kineziny genetika metabolismus MeSH
- lidé MeSH
- messenger RNA genetika metabolismus MeSH
- mutace MeSH
- myši inbrední BALB C MeSH
- myši inbrední C57BL MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- nervus ischiadicus cytologie metabolismus MeSH
- neurony cytologie metabolismus MeSH
- primární buněčná kultura MeSH
- proteinové domény MeSH
- proteiny vázající RNA chemie genetika metabolismus MeSH
- sekvence aminokyselin MeSH
- spinální ganglia cytologie metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
C-terminal mutations of the nucleolar protein nucleophosmin (NPM) are the most frequent genetic aberration detected in acute myeloid leukemia (AML) with normal karyotype. The mutations cause aberrant cytoplasmic localization of NPM and lead to loss of functions associated with NPM nucleolar localization, e.g. in ribosome biogenesis or DNA-damage repair. NPM has many interaction partners and some of them were proved to interact also with the mutated form (NPMmut) and due to this interaction thereby to be withdrawn from their site of action. We analyzed the impact of the mutation on NPM interaction with nucleolin (NCL) which is also prevalently localized into the nucleolus and cooperates with wild-type NPM (NPMwt) in many cellular processes. We revealed that the NCL-NPM complex formation is completely abolished by the mutation and that the presence/absence of the interaction is not affected by drugs causing genotoxic stress or differentiation. Deregulation resulting from changes of NCL/NPMwt ratio may contribute to leukemogenesis.
- MeSH
- akutní myeloidní leukemie genetika metabolismus patologie MeSH
- buněčné jadérko genetika metabolismus patologie MeSH
- fosfoproteiny genetika metabolismus MeSH
- HEK293 buňky MeSH
- jaderné proteiny genetika metabolismus MeSH
- lidé MeSH
- multiproteinové komplexy genetika metabolismus MeSH
- mutace * MeSH
- nádorové buněčné linie MeSH
- nádorové proteiny genetika metabolismus MeSH
- proteiny vázající RNA genetika metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Protein aggregates and abnormal proteins are toxic and associated with neurodegenerative diseases. There are several mechanisms to help cells get rid of aggregates but little is known on how cells prevent aggregate-prone proteins from being synthesised. The EBNA1 of the Epstein-Barr virus (EBV) evades the immune system by suppressing its own mRNA translation initiation in order to minimize the production of antigenic peptides for the major histocompatibility (MHC) class I pathway. Here we show that the emerging peptide of the disordered glycine-alanine repeat (GAr) within EBNA1 dislodges the nascent polypeptide-associated complex (NAC) from the ribosome. This results in the recruitment of nucleolin to the GAr-encoding mRNA and suppression of mRNA translation initiation in cis. Suppressing NAC alpha (NACA) expression prevents nucleolin from binding to the GAr mRNA and overcomes GAr-mediated translation inhibition. Taken together, these observations suggest that EBNA1 exploits a nascent protein quality control pathway to regulate its own rate of synthesis that is based on sensing the nascent GAr peptide by NAC followed by the recruitment of nucleolin to the GAr-encoding RNA sequence.
- MeSH
- alanin MeSH
- fosfoproteiny MeSH
- glycin MeSH
- infekce virem Epsteina-Barrové * MeSH
- lidé MeSH
- messenger RNA genetika metabolismus MeSH
- peptidy genetika MeSH
- proteinové agregáty MeSH
- proteiny vázající RNA metabolismus MeSH
- virus Epsteinův-Barrové - jaderné antigeny metabolismus MeSH
- virus Epsteinův-Barrové * genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- MeSH
- blastocysta metabolismus MeSH
- buněčné jadérko metabolismus MeSH
- chromozomální proteiny, nehistonové metabolismus MeSH
- DNA metabolismus MeSH
- fluorescenční mikroskopie MeSH
- fosfoproteiny metabolismus MeSH
- jaderné proteiny metabolismus MeSH
- myši MeSH
- těhotenství MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- těhotenství MeSH
- ženské pohlaví MeSH
- zvířata MeSH
Nucleolin is a major highly phosphorylated nucleolar protein involved in the regulation of r-chromatin condensation/expansion and rDNA transcription as well as in rRNA processing. The nucleolar protein homologous to the mammalian nucleolin and to the onion nucleolin-like protein NopA100, was detected in the nuclear soluble protein fraction, and in the nuclear matrix fractionfrom Lepidium sativum root meristematic cells, using the selective silver staining method and the cross-reaction with the anti-NopA100 antibody. In 2-DE Western blots of both nuclear fractions, the nucleolin-like protein was revealed as a smear on the level of 90 kDa extending through a certain range of pI. In both extracts obtained from seedlings germinated and grown under slow clinorotation, the extension of the pI range was shorter and the molecular weight diapason was thinner than in the 1 g control; moreover, in the nuclear matrix fraction, the spread of the pI range was separated into two clusters. The results obtained could indicate a lower phosphorylation of the protein, suggesting a decrease in the activity of L. sativum nucleolin-like protein under clinorotation.
Nucleic acid aptamers can specifically bind to target molecules on the cell membrane that mediate their entrance into the cells. Their small size, high binding affinity, specificity, good biocompatibility, stability and low immunogenicity make them ideal drug delivery systems for cancer therapy. These biopharmaceuticals have potential for the delivery of anticancer compounds to diseased tissues, increasing their effectiveness while mitigating the off-target toxicity towards healthy cells. Herein, we have studied two quadruplex-forming DNA sequences derived from the nucleolin-targeted aptamer AS1411 as supramolecular carriers for the cancer-selective delivery of acridine orange derivatives (C3, C5 and C8) in cervical cancer cells. The devised delivery strategy relied on the non-covalent association of the acridine derivatives and the G-quadruplex (G4) structures. This association is done with a high binding strength, as suggested by the obtained KD values in the 10-6-10-7 M range, leading to the thermal stabilization of the G4 structures, particularly for C8. The stability of the resulting supramolecular conjugates was evaluated in fetal bovine serum, which proved their resistance against serum nucleases up to 48 h. Previous studies showed that the tested acridine orange derivatives were cytotoxic towards cervical cancer cells (HeLa) and non-malignant cells. However, when conjugated to AS1411 derivatives, the cytotoxicity of the free ligands towards non-malignant cells was restrained. Furthermore, conjugated C3 showed an enhanced cytotoxicity against HeLa cancer cells. Confocal microscopy indicated that both G4 sequences appear to colocalize with nucleolin, suggesting their ability to recognize and bind nucleolin on the cell surface. Additionally, the results confirmed the internalization of these delivery systems into HeLa cancer cells and their sustained cell trafficking, although being able to dissociate intracellularly to deliver C8 to the nucleoli. Overall, we showed that AS1411-derived G4s can be used as a potential cancer drug delivery system for cervical cancer.
- MeSH
- akridinová oranž aplikace a dávkování analogy a deriváty chemie MeSH
- aptamery nukleotidové aplikace a dávkování chemie MeSH
- buněčné linie MeSH
- G-kvadruplexy * MeSH
- lékové transportní systémy * MeSH
- lidé MeSH
- ligandy MeSH
- nádory děložního čípku metabolismus MeSH
- oligodeoxyribonukleotidy aplikace a dávkování chemie MeSH
- viabilita buněk účinky léků MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
