Conserved Nuclear Export Sequences in Schizosaccharomyces pombe Mex67 and Human TAP Function in mRNA Export by Direct Nuclear Pore Interactions

Thakurta, Anjan; Gopal, Ganesh; Yoon, Jin Ho; Saha, Tapas Kumar; Dhar, Ravi

doi:10.1074/jbc.m309731200

Cited by 13 publications

(15 citation statements)

References 32 publications

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“…Two novel nucleoporin-binding sites, NES I and NES II, located within residues 473-546 of Tap would satisfy the requirements for such alternate NPC-interacting regions. Each of these NESs can drive the nuclear export of fusion proteins (Thakurta et al, 2004). Mutations in the NES I (m9 mutation:V495A and G496A) and NES II (m6 mutation: I529A and V530A), previously shown to disrupt the interaction of these domains with nucleoporins (Thakurta et al, 2004), significantly reduced shuttling (Figure 9).…”

Section: Distribution Of the Tap Mutants In Vivomentioning

confidence: 94%

“…Each of these NESs can drive the nuclear export of fusion proteins (Thakurta et al, 2004). Mutations in the NES I (m9 mutation:V495A and G496A) and NES II (m6 mutation: I529A and V530A), previously shown to disrupt the interaction of these domains with nucleoporins (Thakurta et al, 2004), significantly reduced shuttling (Figure 9). These same mutations also disrupted Nxt1 binding ( Figure 2B).…”

Section: Distribution Of the Tap Mutants In Vivomentioning

confidence: 94%

“…In addition, we generated mutations within the newly identified NES I (m9; V495A and G497A) and NES II (m6; I529A and V530A) located within the Nxt1 heterodimerization region of Tap. Similar mutations were shown to disrupt the ability of these NESs to bind nucleoporins and disrupted their ability to exit the nucleus but the effect of these mutations on Nxt1 binding was not examined (Thakurta et al, 2004). Tap constructs (L383,386R, S585P, and Tap 1-569 ) with mutations in one of the two nucleoporin-binding domains were also tested.…”

Section: Tap Residues Involved In Nxt1 Bindingmentioning

confidence: 99%

“…This double-point mutant was included in the present study to elucidate the role of the NTF2-like hydrophobic fold in the interaction of Tap with nucleoporins. We also tested the effects of two additional mutations within the NTF2-like region, NES I m9 and NES II m6 , which have also been shown to disrupt nucleoporin binding (Thakurta et al, 2004). Maximal binding of Tap to the nucleoporins only occurs when its Nxt1-binding domain is intact and Nxt1 is included in the binding reaction ( Figure 3, present article; .…”

Section: Tap Mutations That Affect In Vitro Binding To Nucleoporinsmentioning

confidence: 99%

“…Mutation of two residues (L383,386R) within this second nucleoporin-binding region (Tap-NBR2) decreases nuclear rim association in permeabilized cells and reduces RNA export activity in a transfected cell assay (Fribourg et al, 2001). In addition to the Tap-NBR2, functional analyses have identified two other regions, referred to as nuclear export sequences (NES) I (residues 473-505) and NES II (residues 505-546) within the NTF2-like domain of Tap that can interact directly with nucleoporins to mediate the export of tethered glucocorticoid receptor-GFP fusion protein in HeLa cells (Thakurta et al, 2004). Mutations of residues 495-497 (VNG3 AAA; m9 mutant) within NES I and residues 529 -530 (IV3 AA; m6 mutant) within NES II abolished both the ability of these NESs to interact with nucleoporins and mediate Tap export function.…”

Section: Introductionmentioning

confidence: 99%

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Mutations in Tap Uncouple RNA Export Activity from Translocation through the Nuclear Pore Complex

Lévesque

Bor

Matzat

et al. 2006

MBoC

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Interactions between transport receptors and phenylalanine-glycine (FG) repeats on nucleoporins drive the translocation of receptor-cargo complexes through nuclear pores. Tap, a transport receptor that mediates nuclear export of cellular mRNAs, contains a UBA-like and NTF2-like folds that can associate directly with FG repeats. In addition, two nuclear export sequences (NESs) within the NTF2-like region can also interact with nucleoporins. The Tap-RNA complex was shown to bind to three nucleoporins, Nup98, p62, and RanBP2, and these interactions were enhanced by Nxt1. Mutations in the Tap-UBA region abolished interactions with all three nucleoporins, whereas the effect of point mutations within the NTF2-like domain of Tap known to disrupt Nxt1 binding or nucleoporin binding were nucleoporin dependent. A mutation in any of these Tap domains was sufficient to reduce RNA export but was not sufficient to disrupt Tap interaction with the NPC in vivo or its nucleocytoplasmic shuttling. However, shuttling activity was reduced or abolished by combined mutations within the UBA and either the Nxt1-binding domain or NESs. These data suggest that Tap requires both the UBA-and NTF2-like domains to mediate the export of RNA cargo, but can move through the pores independently of these domains when free of RNA cargo. INTRODUCTIONNuclear transport plays an important role in cell function by selectively segregating macromolecules to the nuclear or cytoplasmic compartments. This biased distribution contributes to the regulation of many proteins including transcription factors, cell cycle regulators, and cell signaling components (Komeili and O'Shea, 2000;Macara, 2001;Carmo-Fonseca, 2002). Regulated nuclear export of mRNA serves as a quality control step to ensure that only properly spliced mRNA is exported to the cytoplasm for translation (Lei and Silver, 2002;Stutz and Izaurralde, 2003). All nucleocytoplasmic traffic must go through large, proteinaceous channels known as nuclear pore complexes (NPCs; Suntharalingam and Wente, 2003). Although small molecules (Ͻ40 kDa) can diffuse freely through the NPC, movement of most proteins and RNAs across nuclear pores requires binding to soluble import or export receptors (Macara, 2001;Weis, 2002;Bednenko et al., 2003;Pemberton and Paschal, 2005). Import and export receptors, in turn, mediate protein and RNA translocation through highly transient interactions with nucleoporins in the NPC.The majority of nuclear transport receptors belong to the karyopherin/importin ␤ family of proteins (Macara, 2001;Bednenko et al., 2003). Crm1, the best-characterized export receptor and a member of the importin ␤ family, mediates the export of proteins, U snRNAs, and 5S RNAs (Fornerod et al., 1997;Mattaj and Englmeier, 1998;Cullen, 2003). The transport receptor believed to be responsible for the bulk of mRNA export in eukaryotes is Tap/NXF1, a factor that is unrelated to the importin ␤ family (Katahira et al., 1999;Cullen, 2003). Tap also mediates nuclear export of certain retroviral mRNAs, such as the Mas...

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Section: Distribution Of the Tap Mutants In Vivomentioning

confidence: 94%

Section: Distribution Of the Tap Mutants In Vivomentioning

confidence: 94%

Section: Tap Residues Involved In Nxt1 Bindingmentioning

confidence: 99%

Section: Tap Mutations That Affect In Vitro Binding To Nucleoporinsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Mutations in Tap Uncouple RNA Export Activity from Translocation through the Nuclear Pore Complex

Lévesque

Bor

Matzat

et al. 2006

MBoC

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Current awareness on yeast

2004

Yeast

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In order to keep subscribers up‐to‐date with the latest developments in their field, this current awareness service is provided by John Wiley & Sons and contains newly‐published material on yeasts. Each bibliography is divided into 10 sections. 1 Books, Reviews & Symposia; 2 General; 3 Biochemistry; 4 Biotechnology; 5 Cell Biology; 6 Gene Expression; 7 Genetics; 8 Physiology; 9 Medical Mycology; 10 Recombinant DNA Technology. Within each section, articles are listed in alphabetical order with respect to author. If, in the preceding period, no publications are located relevant to any one of these headings, that section will be omitted. (5 weeks journals ‐ search completed 21st. July 2004)

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Genome‐wide screen of fission yeast mutants for sensitivity to 6‐azauracil, an inhibitor of transcriptional elongation

Zhou

Liu

Shi

et al. 2015

Yeast

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Abstract6-Azauracil (6 AU) inhibits enzymes in nucleoside synthesis and depletes the intracellular GTP/UTP pool. Mutations in transcriptional elongation machinery, as well as mutations in a variety of other pathways, exaggerate the growth defect of cells in the presence of 6 AU. Thus, identification of mutations that render cells sensitive to 6 AU will benefit study on the basis of 6 AU-sensitive phenotype. Here we performed a genome-wide screen of a fission yeast deletion library. Of 3235 single-gene deletions, 66 mutants displayed at least 50% drop of fitness in the presence of 6 AU and 60 mutants were reported for the first time; five deletions showed synthetic decrease of fitness when combined with deletion of set3 + , which encodes a transcriptional regulator. Genes conferring tolerance to 6 AU were enriched in various processes, especially in chromosome segregation. Accordingly, genes encoding subunits of CLRC complex and spindle pole body were over-represented. Mutants were subjected to an in vivo transcript length-dependent reporter assay to assess the potential roles of deleted genes in transcriptional elongation. As with the deletions known to affect elongation, nab2Δ, nxt1Δ, rhp18Δ, SPAC24C9.08Δ, clr3Δ and ncs1Δset3Δ mutants exhibited defects in expressing long transcripts. New 6 AU-sensitive mutants identified here will help to elucidate the mechanism of action of 6 AU in the cells. Meanwhile, our study revealed novel genes potentially involved in transcriptional elongation and provided valuable targets for transcription study.

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Conserved Nuclear Export Sequences in Schizosaccharomyces pombe Mex67 and Human TAP Function in mRNA Export by Direct Nuclear Pore Interactions

Cited by 13 publications

References 32 publications

Mutations in Tap Uncouple RNA Export Activity from Translocation through the Nuclear Pore Complex

Mutations in Tap Uncouple RNA Export Activity from Translocation through the Nuclear Pore Complex

Current awareness on yeast

Genome‐wide screen of fission yeast mutants for sensitivity to 6‐azauracil, an inhibitor of transcriptional elongation

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