Identification of different roles for RanGDP and RanGTP in nuclear protein import.

Görlich, Dirk; Panté, Nelly; Kutay, Ulrike; Aebi, Ueli; Bischoff, F. Ralf

doi:10.1002/j.1460-2075.1996.tb00943.x

Cited by 604 publications

(664 citation statements)

References 75 publications

Supporting

Mentioning

624

Contrasting

Unclassified

Order By: Relevance

“…Furthermore, we observed that importin ␤ potentiates the binding of importin ␣ to GR ( Figure 3F). Similar results have been observed for the SV40 NLS (Rexach and Blobel, 1995;Gorlich et al, 1996). It will be interesting to determine the role that importin ␤ plays in this interaction, whether it simply stabilizes the importin ␣-GR NL1 interaction or can bind to NL1 directly.…”

Section: Discussionsupporting

confidence: 74%

“…Because importin ␤ binds importin ␣ and the importin ␣/importin ␤/SV40 NLS complex can be dissociated by RanQ69L (Rexach and Blobel, 1995;Gorlich et al, 1996), we investigated the role of importin ␤ in the binding of importin ␣ to the NL1 of GR. The presence of recombinant importin ␤ potentiated the binding of importin ␣ but not importin 7 to a purified GR fragment (407-556) containing NL1 ( Figure 3F, compare lanes 3-4 and 5-6), suggesting that importin ␤ plays an important role in importin ␣ but not importin 7 mediated GR nuclear import.…”

Section: ϩH) (B) Padh-n795mentioning

confidence: 99%

See 1 more Smart Citation

Importin 7 and Importin α/Importin β Are Nuclear Import Receptors for the Glucocorticoid Receptor

Freedman

2004

MBoC

192

151

View full text Add to dashboard Cite

The vertebrate glucocorticoid receptor (GR) is cytoplasmic without hormone and localizes to the nucleus after hormone binding. GR has two nuclear localization signals (NLS): NL1 is similar in sequence to the SV40 NLS; NL2 is poorly defined, residing in the ligand-binding domain. We found that GR displayed similar hormone-regulated compartmentalization in Saccharomyces cerevisiae and required the Sxm1 nuclear import receptor for NL2-mediated import. Two metazoan homologues of Sxm1, importin 7 and importin 8, bound both NL1 and NL2, whereas importin ␣ selectively bound NL1. In an in vitro nuclear import assay, both importin 7 and the importin ␣-importin ␤ heterodimer could import a GR NL1 fragment. Under these conditions, full-length GR localized to nuclei in the presence but not absence of an unidentified component in cell extracts. Interestingly, importin 7, importin 8, and importin ␣ bound GR even in the absence of hormone; thus, hormonal control of localization is exerted at a step downstream of import receptor binding. INTRODUCTIONTo survive in a complex environment, cells sense external cues and commonly respond by altering gene expression, in part by regulating the intracellular localization and activity of transcriptional regulatory factors. For example, the intracellular localization of the yeast Pho4 and mammalian SREBP factors is altered in response to changes in external phosphate and circulating sterol concentrations, respectively (Kaffman et al., 1998b;Nagoshi et al., 1999). Similarly, changes in blood glucose levels and a wide range of physiological stress signals modulate the circulating level of glucocorticoids, changing the activity and localization of the glucocorticoid receptor (GR). Within minutes of glucocorticoid binding, GR enters the nucleus and activates or represses target gene transcription (Picard and Yamamoto, 1987). However, hormone binding is reversible; after hormone withdrawal, GR locates back to the cytoplasm (t 1/2 ϭ 10 h; Hache et al., 1999).Proteins and RNAs enter and exit the nucleus through the nuclear pore, a 125-MDa complex in the nuclear envelope (Stoffler et al., 1999). Small molecules readily diffuse through the nuclear pore, whereas molecules greater than ϳ40 kDa require import and export machinery for transport (Davis, 1995;Pante and Aebi, 1995). The first nuclear localization sequence (NLS) was identified in the SV40 large T-antigen (Kalderon et al., 1984). Development of an in vitro nuclear import assay facilitated the identification of two proteins that import substrates containing SV40 NLS-like domains. The adapter molecule importin ␣ binds to the NLS of the substrate and the importin ␤ transport receptor, creating a trimeric complex that imports the substrate into the nucleus through the nuclear pore (Strom and Weis, 2001).Based on similarity to importin ␤, a family of nuclear import and export receptors was identified and shown to transport a wide variety of proteins and RNAs into and out of the nucleus. After reaching the nucleoplasm, import receptors bind the...

show abstract

Section: Discussionsupporting

confidence: 74%

Section: ϩH) (B) Padh-n795mentioning

confidence: 99%

Importin 7 and Importin α/Importin β Are Nuclear Import Receptors for the Glucocorticoid Receptor

Freedman

2004

MBoC

192

151

View full text Add to dashboard Cite

show abstract

“…The predicted reduction in SAM biosynthesis and hypomethylation of pre-tRNAs might impede their proper folding and processing, as occurs in gcd10 and gcd14 mutants defective in methylation of m1A 58 (Anderson et al 1998;Calvo et al 1999). It is also interesting to note that Ran-dependent nucleocytoplasmic transport is dependent on both GTP-and GDPbound forms of the Ran GTPase, and that defects in tRNA maturation have been observed in nucleoporin mutants (Gorlich et al 1996;Sharma et al 1996;Izaurralde et al 1997). Hence, the abnormal GTP/ GDP ratio in the gua1-G388D mutant could contribute to the observed defects in tRNA maturation by interfering with Ran function.…”

Section: Resultsmentioning

confidence: 99%

Guanine Nucleotide Pool Imbalance Impairs Multiple Steps of Protein Synthesis and Disrupts GCN4 Translational Control in Saccharomyces cerevisiae

et al. 2011

View full text Add to dashboard Cite

Purine nucleotides are structural components of the genetic material, function as phosphate donors, participate in cellular signaling, are cofactors in enzymatic reactions, and constitute the main carriers of cellular energy. Thus, imbalances in A/G nucleotide biosynthesis affect nearly the whole cellular metabolism and must be tightly regulated. We have identified a substitution mutation (G388D) that reduces the activity of the GMP synthase Gua1 in budding yeast and the total G-nucleotide pool, leading to precipitous reductions in the GDP/GTP ratio and ATP level in vivo. gua1-G388D strongly reduces the rate of growth, impairs general protein synthesis, and derepresses translation of GCN4 mRNA, encoding a transcriptional activator of diverse amino acid biosynthetic enzymes. Although processing of pre-tRNA i Met and other tRNA precursors, and the aminoacylation of tRNA iMet are also strongly impaired in gua1-G388D cells, tRNA i GTP complex (TC) and the multifactor complex (MFC) required for translation initiation accumulate $10-fold in gua1-G388D cells and, to a lesser extent, in wild-type (WT) cells treated with 6-azauracil (6AU). Consistently, addition of an external supply of guanine reverts all the phenotypes of gua1-G388D cells, but not those of gua1-G388D Dhpt1 mutants unable to refill the internal GMP pool through the salvage pathway. These and other findings suggest that a defect in guanine nucleotide biosynthesis evokes a reduction in the rate of general protein synthesis by impairing multiple steps of the process, disrupts the gene-specific reinitiation mechanism for translation of GCN4 mRNA and has far-reaching effects in cell biology and metabolism.

show abstract

“…Accordingly it has been suggested that Ran⅐GDP is the active form of the protein at the cytoplasmic side of the nuclear pore and that GTP-bound Ran is required to terminate translocation at the nucleoplasmic side with the Ran⅐GDP/GTP cycle providing directionality of transport. Evidence for this model includes the findings that Ran⅐GDP can bind to the NPC via its interaction with p10/NTF2, that a Ran mutant locked in the GTP-bound state fails to support nuclear protein import and prevents docking of an import substrate at the NPC, and that Ran⅐GTP can bind to importin ␤ thereby inducing the dissociation of importin ␣ from importin ␤ and most likely the termination of translocation (15,23,24).Genes encoding the principle regulators of the Ran GTPase cycle have also been shown to be essential for viability in budding and fission yeasts. In Saccharomyces cerevisiae and Schizosaccharomyces pombe, these genes encode the Ran homologues Gsp1p and spi1p (25,26), the RCC1 homologues Prp20p and pim1p (26,27), and the RanGAP homologues Rna1p and rna1p, respectively (28, 29).…”

mentioning

confidence: 99%

mentioning

confidence: 99%

The Acidic C-terminal Domain of rna1p Is Required for the Binding of Ran·GTP and for RanGAP Activity

Haberland¹,

Becker²,

Gerke³

1997

Journal of Biological Chemistry

View full text Add to dashboard Cite

The small GTP binding protein Ran is an essential component of the nuclear protein import machinery whose GTPase cycle is regulated by the nuclear guanosine nucleotide exchange factor RCC1 and by the cytosolic GTPase activating protein RanGAP. In the yeasts Schizosaccharomyces pombe and Saccharomyces cerevisiae the RanGAP activity is encoded by the RNA1 genes which are essential for cell viability and nucleocytoplasmic transport in vivo. Although of limited sequence identity the two yeast proteins show a conserved structural organization characterized by an N-terminal domain of eight leucine-rich repeats, motifs implicated in protein-protein interactions, and a C-terminal domain rich in acidic amino acid residues. By analyzing the RanGAP activity of a series of recombinantly expressed rna1p mutant derivatives, we show that the highly acidic sequence in the C-terminal domain of both yeast proteins is indispensable for activating Ran-mediated GTP hydrolysis. Chemical cross-linking reveals that the same sequence in rna1p is required for rna1p⅐Ran complex formation indicating that the loss of GAP activity in the C-terminally truncated rna1p mutants results from an impaired interaction with Ran. The predominant species stabilized through the covalent cross-link is a rna1p⅐Ran heterodimer whose formation requires the GTP-bound conformation of Ran. As the acidic C-terminal domain of rna1p is required for establishing the interaction with Ran, the leucine-rich repeats domain in rna1p is potentially available for additional protein interactions perhaps required for directing a fraction of rna1p to the nuclear pore.The import of karyophilic proteins into the nucleus which occurs through the nuclear pore complex (NPC) 1 is an energydependent process specified by nuclear localization signals (NLSs) on the import substrate (for reviews see Refs. 1-4). Using digitonin-permeabilized HeLa cells as an in vitro transport system, it was shown that the import of substrate proteins is a multistep process depending on four cytosolic factors (5, 6) (for reviews see Refs. 7-10). Two of these factors form a heterodimeric complex serving as the NLS receptor. The smaller 60-kDa subunit of this complex, also known as importin ␣, is responsible for NLS binding, whereas the larger 97-kDa subunit, importin ␤, most likely mediates a targeting of the substrate-NLS receptor complex to the NPC. After docking the entire substrate-receptor complex is thought to travel through the NPC and then to dissociate close to or at the nucleoplasmic side of the pore (for review see Ref. 11). The two other factors identified in the in vitro transport system as essential components of the nuclear import machinery are the GTP-binding protein Ran/TC4 (herein referred to as Ran (12, 13)) and a small protein of 10 kDa, p10/NTF2. The latter can interact with NPC proteins, Ran, and importin ␤ and appears to be involved in the formation of a pentameric complex including p10, Ran in its GDP-bound form, the two NLS receptor subunits and a nuclear pore protein (14, 15).R...

show abstract

Identification of different roles for RanGDP and RanGTP in nuclear protein import.

Cited by 604 publications

References 75 publications

Importin 7 and Importin α/Importin β Are Nuclear Import Receptors for the Glucocorticoid Receptor

Importin 7 and Importin α/Importin β Are Nuclear Import Receptors for the Glucocorticoid Receptor

Guanine Nucleotide Pool Imbalance Impairs Multiple Steps of Protein Synthesis and Disrupts GCN4 Translational Control in Saccharomyces cerevisiae

The Acidic C-terminal Domain of rna1p Is Required for the Binding of Ran·GTP and for RanGAP Activity

Contact Info

Product

Resources

About