The Experts below are selected from a list of 17304 Experts worldwide ranked by ideXlab platform
Daniel Levy - One of the best experts on this subject based on the ideXlab platform.
-
the nucleoporin elys regulates Nuclear size by controlling npc number and Nuclear Import capacity
EMBO Reports, 2019Co-Authors: Predrag Jevtic, Andria C Schibler, Chase C Wesley, Gianluca Pegoraro, Tom Misteli, Daniel LevyAbstract:Abstract How intracellular organelles acquire their characteristic sizes is a fundamental question in cell biology. Given stereotypical changes in Nuclear size in cancer, it is Important to understand the mechanisms that control Nuclear size in human cells. Using a high‐throughput imaging RNAi screen, we identify and mechanistically characterize ELYS, a nucleoporin required for post‐mitotic Nuclear pore complex (NPC) assembly, as a determinant of Nuclear size in mammalian cells. ELYS knockdown results in small nuclei, reduced Nuclear lamin B2 localization, lower NPC density, and decreased Nuclear Import. Increasing Nuclear Import by Importin α overexpression rescues Nuclear size and lamin B2 Import, while inhibiting Importin α/β‐mediated Nuclear Import decreases Nuclear size. Conversely, ELYS overexpression increases Nuclear size, enriches Nuclear lamin B2 at the Nuclear periphery, and elevates NPC density and Nuclear Import. Consistent with these observations, knockdown or inhibition of exportin 1 increases Nuclear size. Thus, we identify ELYS as a novel positive effector of mammalian Nuclear size and propose that Nuclear size is sensitive to NPC density and Nuclear Import capacity.
-
Altering the levels of Nuclear Import factors in early Xenopus laevis embryos affects later development.
PLOS ONE, 2019Co-Authors: Predrag Jevtić, Richik Nilay Mukherjee, Pan Chen, Daniel LevyAbstract:More than just a container for DNA, the Nuclear envelope carries out a wide variety of critical and highly regulated cellular functions. One of these functions is Nuclear Import, and in this study we investigate how altering the levels of Nuclear transport factors impacts developmental progression and organismal size. During early Xenopus laevis embryogenesis, the timing of a key developmental event, the midblastula transition (MBT), is sensitive to Nuclear Import factor levels. How might altering Nuclear Import factors and MBT timing in the early embryo affect downstream development of the organism? We microinjected X. laevis two-cell embryos with mRNA to increase levels of Importin α or NTF2, resulting in differential amounts of Nuclear Import factors in the two halves of the embryo. Compared to controls, these embryos exhibited delayed gastrulation, curved neural plates, and bent tadpoles with different sized eyes. Furthermore, embryos microinjected with NTF2 developed into smaller froglets compared to control microinjected embryos. We propose that altering Nuclear Import factors and Nuclear size affects MBT timing, cell size, and cell number, subsequently disrupting later development. Thus, altering Nuclear Import factors early in development can affect function and size at the organismal level.
-
Altering Nuclear Import in early Xenopus laevis embryos affects later development
bioRxiv, 2019Co-Authors: Predrag Jevtić, Daniel LevyAbstract:More than just a container for DNA, the nucleus carries out a wide variety of critical and highly regulated cellular functions. One of these functions is Nuclear Import, and in this study we investigate how altering Nuclear Import impacts developmental progression and organismal size. During early Xenopus laevis embryogenesis, the timing of a key developmental event, the midblastula transition (MBT), is sensitive to Nuclear Import factor levels. How might altering Nuclear Import and MBT timing in the early embryo affect downstream development of the organism? We microinjected X. laevis two-cell embryos to increase levels of Importin α or NTF2, resulting in differential amounts of Nuclear Import factors in the two halves of the embryo. Compared to controls, these embryos exhibited delayed gastrulation, curved neural plates, and bent tadpoles with different sized eyes. Furthermore, embryos microinjected with NTF2 developed into smaller froglets compared to control microinjected embryos. We propose that altering Nuclear Import and size affects MBT timing, cell size, and cell number, subsequently disrupting later development. Thus, altering Nuclear Import early in development can affect function and size at the organismal level.
-
The nucleoporin ELYS regulates Nuclear size by controlling NPC number and Nuclear Import capacity
bioRxiv, 2019Co-Authors: Predrag Jevtić, Andria C Schibler, Gianluca Pegoraro, Tom Misteli, Daniel LevyAbstract:How intracellular organelles acquire their characteristic sizes is a fundamental cell biological question. Given the stereotypical changes in Nuclear size in cancer, it is particularly Important to understand the mechanisms that control Nuclear size in human cells. Here we use a high-throughput imaging RNAi screen to identify and mechanistically characterize ELYS, a nucleoporin required for postmitotic Nuclear pore complex (NPC) assembly, as a determinant of Nuclear size in mammalian cells. We show that ELYS knockdown results in small nuclei, the accumulation of cytoplasmic lamin aggregates, reduced Nuclear lamin B2 localization, lower NPC density, and decreased Nuclear Import. Increasing Nuclear Import by Importin α overexpression rescues Nuclear size and lamin B2 Import, while inhibiting Importin α/β Nuclear Import decreases Nuclear size. Conversely, ELYS overexpression leads to increased Nuclear size, enrichment of Nuclear lamin B2 staining at the Nuclear periphery, and elevated NPC density and Nuclear Import. Consistent with these observations, knockdown or inhibition of exportin 1 increases Nuclear size. In summary, we identify ELYS and NPC density as novel positive effectors of mammalian Nuclear size and propose that Nuclear size is controlled by Nuclear Import capacity.
A Fassati - One of the best experts on this subject based on the ideXlab platform.
-
structural basis for Nuclear Import of splicing factors by human transportin 3
Proceedings of the National Academy of Sciences of the United States of America, 2014Co-Authors: Goedele N Maertens, N Cook, Weifeng Wang, Stephen Hare, S S Gupta, Ilker Oztop, Ophelie Cosnefroy, Ambrosius P Snijders, Vineet N Kewalramani, A FassatiAbstract:Transportin 3 (Tnpo3, Transportin-SR2) is implicated in Nuclear Import of splicing factors and HIV-1 replication. Herein, we show that the majority of cellular Tnpo3 binding partners contain arginine-serine (RS) repeat domains and present crystal structures of human Tnpo3 in its free as well as GTPase Ran- and alternative splicing factor/splicing factor 2 (ASF/SF2)-bound forms. The flexible β-karyopherin fold of Tnpo3 embraces the RNA recognition motif and RS domains of the cargo. A constellation of charged residues on and around the arginine-rich helix of Tnpo3 HEAT repeat 15 engage the phosphorylated RS domain and are critical for the recognition and Nuclear Import of ASF/SF2. Mutations in the same region of Tnpo3 impair its interaction with the cleavage and polyadenylation specificity factor 6 (CPSF6) and its ability to support HIV-1 replication. Steric incompatibility of the RS domain and RanGTP engagement by Tnpo3 provides the mechanism for cargo release in the nucleus. Our results elucidate the structural bases for Nuclear Import of splicing factors and the Tnpo3–CPSF6 nexus in HIV-1 biology.
-
tRNAs promote Nuclear Import of HIV-1 intracellular reverse transcription complexes
PLOS BIOL, 2006Co-Authors: A FassatiAbstract:Infection of non-dividing cells is a biological property of HIV-1 crucial for virus transmission and AIDS pathogenesis. This property depends on Nuclear Import of the intracellular reverse transcription and pre-integration complexes (RTCs/PICs). To identify cellular factors involved in Nuclear Import of HIV-1 RTCs, cytosolic extracts were fractionated by chromatography and Import activity examined by the Nuclear Import assay. A near-homogeneous fraction was obtained, which was active in inducing Nuclear Import of purified and labeled RTCs. The active fraction contained tRNAs, mostly with defective 3' CCA ends. Such tRNAs promoted HIV-1 RTC Nuclear Import when synthesized in vitro. Active tRNAs were incorporated into and recovered from virus particles. Mutational analyses indicated that the anticodon loop mediated binding to the viral complex whereas the T-arm may interact with cellular factors involved in Nuclear Import. These tRNA species efficiently accumulated into the nucleus on their own in a energy- and temperature-dependent way. An HIV-1 mutant containing MLV gag did not incorporate tRNA species capable of inducing HIV-1 RTC Nuclear Import and failed to infect cell cycle-arrested cells. Here we provide evidence that at least some tRNA species can be Imported into the nucleus of human cells and promote HIV-1 Nuclear Import.
-
Nuclear Import of HIV‐1 intracellular reverse transcription complexes is mediated by Importin 7.
The EMBO Journal, 2003Co-Authors: A Fassati, Dirk Görlich, Ian F. Harrison, Lyubov Zaytseva, José-manuel MingotAbstract:Human immunodeficiency virus type 1 (HIV-1), like other lentiviruses, can infect non-dividing cells. This property depends on the active Nuclear Import of its intracellular reverse transcription complex (RTC). We have studied Nuclear Import of purified HIV-1 RTCs in primary macrophages and found that Importin 7, an Import receptor for ribosomal proteins and histone H1, is involved in the process. Nuclear Import of RTCs requires, in addition, energy and the components of the Ran system. Depletion of Importin 7 from cultured cells by small interfering RNA inhibits HIV-1 infection. These results provide a new insight into the molecular mechanism for HIV-1 Nuclear Import and reveal potential targets for therapeutic intervention.
Anil K Jaiswal - One of the best experts on this subject based on the ideXlab platform.
-
prothymosin alpha mediates Nuclear Import of the inrf2 cul3 rbx1 complex to degrade Nuclear nrf2
Journal of Biological Chemistry, 2009Co-Authors: Suryakant K Niture, Anil K JaiswalAbstract:Abstract Nrf2-mediated coordinated induction of a battery of defensive genes is a critical mechanism in cellular protection and survival. INrf2 (Keap1), an inhibitor of Nrf2, functions as an adaptor for Cul3·Rbx1-mediated degradation of Nrf2. A majority of the INrf2/Cul3·Rbx1 complex is localized in the cytosol that degrades cytosolic Nrf2. However, 10-15% of INrf2 is also localized inside the nucleus. INrf2 does not contain a defined Nuclear Import signal, and the mechanism of Nuclear Import and its function inside the nucleus remain obscure. Present studies demonstrate that the DGR region of INrf2 is required for Nuclear Import of INrf2. Studies also demonstrate that Cul3 and Rbx1 are also Imported inside the nucleus in complex with INrf2. Interestingly, Nrf2 and prothymosin-α both bind to the DGR region of INrf2. However, it is prothymosin-α and not Nrf2 that mediates Nuclear Import of INrf2/Cul3·Rbx1 complex. Antioxidant treatment increases Nuclear Import of INrf2/Cul3·Rbx1 complex. The INrf2/Cul3·Rbx1 complex inside the nucleus exchanges prothymosin-α with Nrf2, resulting in degradation of Nrf2. These results led to the conclusion that prothymosin-α-mediated Nuclear Import of INrf2/Cul3·Rbx1 complex leads to ubiquitination and degradation of Nrf2 inside the nucleus presumably to regulate Nuclear level of Nrf2 and rapidly switch off the activation of Nrf2 downstream gene expression.
-
Prothymosin-alpha mediates Nuclear Import of the INrf2/Cul3 Rbx1 complex to degrade Nuclear Nrf2.
Journal of Biological Chemistry, 2009Co-Authors: Suryakant K Niture, Anil K JaiswalAbstract:Nrf2-mediated coordinated induction of a battery of defensive genes is a critical mechanism in cellular protection and survival. INrf2 (Keap1), an inhibitor of Nrf2, functions as an adaptor for Cul3 Rbx1-mediated degradation of Nrf2. A majority of the INrf2/Cul3 Rbx1 complex is localized in the cytosol that degrades cytosolic Nrf2. However, 10-15% of INrf2 is also localized inside the nucleus. INrf2 does not contain a defined Nuclear Import signal, and the mechanism of Nuclear Import and its function inside the nucleus remain obscure. Present studies demonstrate that the DGR region of INrf2 is required for Nuclear Import of INrf2. Studies also demonstrate that Cul3 and Rbx1 are also Imported inside the nucleus in complex with INrf2. Interestingly, Nrf2 and prothymosin-alpha both bind to the DGR region of INrf2. However, it is prothymosin-alpha and not Nrf2 that mediates Nuclear Import of INrf2/Cul3 Rbx1 complex. Antioxidant treatment increases Nuclear Import of INrf2/Cul3 Rbx1 complex. The INrf2/Cul3 Rbx1 complex inside the nucleus exchanges prothymosin-alpha with Nrf2, resulting in degradation of Nrf2. These results led to the conclusion that prothymosin-alpha-mediated Nuclear Import of INrf2/Cul3 Rbx1 complex leads to ubiquitination and degradation of Nrf2 inside the nucleus presumably to regulate Nuclear level of Nrf2 and rapidly switch off the activation of Nrf2 downstream gene expression.
Gideon Dreyfuss - One of the best experts on this subject based on the ideXlab platform.
-
Transportin-SR, a Nuclear Import Receptor for SR Proteins
Journal of Cell Biology, 1999Co-Authors: Naoyuki Kataoka, Jennifer L. Bachorik, Gideon DreyfussAbstract:The SR proteins, a group of abundant arginine/serine (RS)-rich proteins, are essential pre-mRNA splicing factors that are localized in the nucleus. The RS domain of these proteins serves as a Nuclear localization signal. We found that RS domain–bearing proteins do not utilize any of the known Nuclear Import receptors and identified a novel Nuclear Import receptor specific for SR proteins. The SR protein Import receptor, termed transportin-SR (TRN-SR), binds specifically and directly to the RS domains of ASF/SF2 and SC35 as well as several other SR proteins. The Nuclear transport regulator RanGTP abolishes this interaction. Recombinant TRN-SR mediates Nuclear Import of RS domain– bearing proteins in vitro. TRN-SR has amino acid sequence similarity to several members of the Importin β/transportin family. These findings strongly suggest that TRN-SR is a Nuclear Import receptor for the SR protein family.
-
Functional Conservation of the Transportin Nuclear Import Pathway in Divergent Organisms
Molecular and Cellular Biology, 1998Co-Authors: Mikiko C. Siomi, Micheline Fromont, Jean-christophe Rain, Fan Wang, Pierre Legrain, Gideon DreyfussAbstract:Human transportin1 (hTRN1) is the Nuclear Import receptor for a group of pre-mRNA/mRNA-binding proteins (heterogeneous Nuclear ribonucleoproteins [hnRNP]) represented by hnRNP A1, which shuttle continuously between the nucleus and the cytoplasm. hTRN1 interacts with the M9 region of hnRNP A1, a 38-amino-acid domain rich in Gly, Ser, and Asn, and mediates the Nuclear Import of M9-bearing proteins in vitro. Saccharomyces cerevisiae transportin (yTRN; also known as YBR017c or Kap104p) has been identified and cloned. To understanding the Nuclear Import mediated by yTRN, we searched with a yeast two-hybrid system for proteins that interact with it. In an exhaustive screen of the S. cerevisiae genome, the most frequently selected open reading frame was the Nuclear mRNA-binding protein, Nab2p. We delineated a ca.-50-amino-acid region in Nab2p, termed NAB35, which specifically binds yTRN and is similar to the M9 motif. NAB35 also interacts with hTRN1 and functions as a Nuclear localization signal in mammalian cells. Interestingly, yTRN can also mediate the Import of NAB35-bearing proteins into mammalian nuclei in vitro. We also report on additional substrates for TRN as well as sequences of Drosophila melanogaster, Xenopus laevis, and Schizosaccharomyces pombe TRNs. Together, these findings demonstrate that both the M9 signal and the Nuclear Import machinery utilized by the transportin pathway are conserved in evolution.
-
transportin mediated Nuclear Import of heterogeneous Nuclear rnp proteins
Journal of Cell Biology, 1997Co-Authors: Mikiko C. Siomi, Naoyuki Kataoka, Paul S Eder, Gideon DreyfussAbstract:Heterogeneous Nuclear ribonucleoprotein (hnRNP) A1 is an abundant Nuclear protein that plays an Important role in pre-mRNA processing and mRNA export from the nucleus. A1 shuttles rapidly between the nucleus and the cytoplasm, and a 38-amino acid domain, M9, serves as the bidirectional transport signal of A1. Recently, a 90-kD protein, transportin, was identified as the mediator of A1 Nuclear Import. In this study, we show that transportin mediates the Nuclear Import of additional hnRNP proteins, including hnRNP F. We have also isolated and sequenced a novel transportin homolog, transportin2, which may differ from transportin1 in its substrate specificity. Immunostaining shows that transportin1 is localized both in the cytoplasm and the nucleoplasm, and Nuclear rim staining is also observed. The Nuclear localization of A1 is dependent on ongoing RNA polymerase II transcription. Interestingly, a pyruvate kinase–M9 fusion, which normally localizes in the nucleus, also accumulates in the cytoplasm when RNA polymerase II is inhibited. Thus, M9 itself is a specific sensor for transcription-dependent Nuclear transport. Transportin1–A1 complexes can be isolated from the cytoplasm and the nucleoplasm, but transportin1 is not detectable in hnRNP complexes. RanGTP causes dissociation of A1-transportin1 complexes in vitro. Thus, it is likely that after Nuclear Import, A1 dissociates from transportin1 by RanGTP and becomes incorporated into hnRNP complexes, where A1 functions in pre-mRNA processing.
Didier Trono - One of the best experts on this subject based on the ideXlab platform.
-
role of the karyopherin pathway in human immunodeficiency virus type 1 Nuclear Import
Journal of Virology, 1996Co-Authors: Philippe Gallay, V Stitt, Cynthia L Mundy, Marjorie A Oettinger, Didier TronoAbstract:The interaction of the human immunodeficiency virus type 1 (HIV-1) nucleoprotein complex with the cell Nuclear Import machinery is necessary for viral replication in macrophages and for the establishment of infection in quiescent T lymphocytes. The karyophilic properties of two viral proteins, matrix (MA) and Vpr, are keys to this process. Here, we show that an early step of HIV-1 Nuclear Import is the recognition of the MA Nuclear localization signal (NLS) by Rch1, a member of the karyopherin-alpha family. Furthermore, we demonstrate that an N-terminally truncated form of Rch1 which binds MA but fails to localize to the nucleus efficiently blocks MA- but not Vpr-mediated HIV-1 Nuclear Import. Correspondingly, NLS peptide inhibits the Nuclear migration of MA but not that of Vpr and prevents the infection of terminally differentiated macrophages by vpr-defective virus but not wild-type virus. These results are consistent with a model in which Rch1 or another member of the karyopherin-alpha family, through the recognition of the MA NLS, participates in docking the HIV-1 nucleoprotein complex at the Nuclear pore. In addition, our data suggest that Vpr governs HIV-1 Nuclear Import through a distinct pathway.
