Search results for "Nuclear transport"
showing 10 items of 26 documents
Probing Differential Binding Mechanisms of Phenylalanine-Glycine-Rich Nucleoporins by Single-Molecule FRET
2018
Abstract Phenylalanine-glycine-rich nucleoporins (FG-Nups) are intrinsically disordered proteins, constituting the selective barrier of the nuclear pore complex. They are highly dynamic under physiological conditions and studying their interaction with nuclear transport receptors (NTRs) is key to understanding the molecular mechanism of nucleocytoplasmic transport. Distinct conformational features of FG-Nups interacting with diverse NTRs can be detected by multiparameter single-molecule fluorescence energy transfer (smFRET), which is a powerful technique for studying the dynamics and interactions of biomolecules in solution. Here we provide a detailed protocol utilizing smFRET to reveal dif…
Two differential binding mechanisms of FG-nucleoporins and nuclear transport receptors
2018
Summary Phenylalanine-glycine-rich nucleoporins (FG-Nups) are intrinsically disordered proteins, constituting the selective barrier of the nuclear pore complex (NPC). Previous studies showed that nuclear transport receptors (NTRs) were found to interact with FG-Nups by forming an “archetypal-fuzzy” complex through the rapid formation and breakage of interactions with many individual FG motifs. Here, we use single-molecule studies combined with atomistic simulations to show that, in sharp contrast, FG-Nup214 undergoes a coupled reconfiguration-binding mechanism when interacting with the export receptor CRM1. Association and dissociation rate constants are more than an order of magnitude lowe…
Beyond the Transport Function of Import Receptors: What’s All the FUS about?
2018
Nuclear import receptors are central players in transporting protein cargoes into the nucleus. Moving beyond this role, four newly published articles describe a function in regulating supramolecular assemblies by fine-tuning the phase separating properties of RNA-binding proteins, which has implications for a variety of devastating neurodegenerative disorders.
A comparative study of the degradation of yeast cyclins Cln1 and Cln2.
2016
The yeast cyclins Cln1 and Cln2 are very similar in both sequence and function, but some differences in their functionality and localization have been recently described. The control of Cln1 and Cln2 cellular levels is crucial for proper cell cycle initiation. In this work, we analyzed the degradation patterns of Cln1 and Cln2 in order to further investigate the possible differences between them. Both cyclins show the same half‐life but, while Cln2 degradation depends on ubiquitin ligases SCFG rr1 and SCFC dc4, Cln1 is affected only by SCFG rr1. Degradation analysis of chimeric cyclins, constructed by combining fragments from Cln1 and Cln2, identifies the N‐terminal sequence of the proteins…
Protoparvovirus Knocking at the Nuclear Door
2017
Protoparvoviruses target the nucleus due to their dependence on the cellular reproduction machinery during the replication and expression of their single-stranded DNA genome. In recent years, our understanding of the multistep process of the capsid nuclear import has improved, and led to the discovery of unique viral nuclear entry strategies. Preceded by endosomal transport, endosomal escape and microtubule-mediated movement to the vicinity of the nuclear envelope, the protoparvoviruses interact with the nuclear pore complexes. The capsids are transported actively across the nuclear pore complexes using nuclear import receptors. The nuclear import is sometimes accompanied by structural chan…
Different pathways for the nuclear import of yeast RNA polymerase II
2015
Recent studies suggest that RNA polymerase II (Pol II) has to be fully assembled before being imported into the nucleus, while other reports indicate a distinct mechanism to import large and small subunits. In yeast, Iwr1 binds to the holoenzyme assembled in the cytoplasm and directs its nuclear entry. However, as IWR1 is not an essential gene, Iwr1-independent pathway(s) for the nuclear import of Pol II must exist. In this paper, we investigate the transport into the nucleus of several large and small Pol II subunits in the mutants of genes involved in Pol II biogenesis. We also analyse subcellular localization in the presence of drugs that can potentially affect Pol II nuclear import. Our…
Mid-region parathyroid hormone-related protein (PTHrP) binds chromatin of MDA-MB231 breast cancer cells and isolated oligonucleotides “in vitro”
2006
We have previously shown that PTHrP(38-94)-amide restrains growth and invasion "in vitro", causes striking toxicity and accelerates death of some breast cancer cell lines, the most responsive being MDA-MB231 whose tumorigenesis was also attenuated "in vivo". PTHrP(38-94)-amide contains the domain implicated in the nuclear import of PTHrP. Although the nucleus was identified as a destination for mid-region PTHrP, evidence for direct DNA-binding capability is lacking to date. Here, we examined the localization of PTHrP(38-94)-amide within MDA-MB231 cells and within metaphase spread preparations and characterized its DNA-binding properties, employing a combination of immunocytochemical, cytoge…
XPO1E571K Mutation Modifies Exportin 1 Localisation and Interactome in B-cell Lymphoma
2020
The XPO1 gene encodes exportin 1 (XPO1) that controls the nuclear export of cargo proteins and RNAs. Almost 25% of primary mediastinal B-cell lymphoma (PMBL) and classical Hodgkin lymphoma (cHL) cases harboured a recurrent XPO1 point mutation (NM_003400, chr2:g61718472C>
Spatial regulation of the Start repressor Whi5
2009
The Saccharomyces cerevisiae Start repressor Whi5, the functional analogue of mammalian pRB, shuttles between the nucleus and the cytoplasm throughout the cell cycle: enters into the nucleus at the end of mitosis and remains nuclear until Start. We studied the mechanisms involved in this spatial regulation. The nuclear import depends on the beta-karyopherins of the classical import pathway Kap95 and Cse1. Whi5 contains a monopartite and a bipartite classical NLS localized in its N-terminal region which are functionally redundant. A fragment of Whi5 containing these NLSs is able to constitutively accumulate a GFP(4) protein inside the nucleus throughout the cell cycle, which suggests that th…
Cargo transport through the nuclear pore complex at a glance.
2021
ABSTRACT Bidirectional transport of macromolecules across the nuclear envelope is a hallmark of eukaryotic cells, in which the genetic material is compartmentalized inside the nucleus. The nuclear pore complex (NPC) is the major gateway to the nucleus and it regulates nucleocytoplasmic transport, which is key to processes including transcriptional regulation and cell cycle control. Accordingly, components of the nuclear transport machinery are often found to be dysregulated or hijacked in diseases. In this Cell Science at a Glance article and accompanying poster, we provide an overview of our current understanding of cargo transport through the NPC, from the basic transport signals and mach…