Karsten Weis | Swiss Federal Institute of Technology (ETH) (original) (raw)

Papers by Karsten Weis

Methods in Enzymology, 2000

Current Molecular Medicine, 2001

Molecular biological investigations of HIV have made fundamental contributions to our understandi... more Molecular biological investigations of HIV have made fundamental contributions to our understanding of eukaryotic biology. These studies elucidated new paradigms in transcription, RNA and protein export from the nucleus to the cytoplasm, cellular activation, morphology and vesicular trafficking.

Nature Chemical Biology, 2006

The goal of high-throughput screening (HTS) from the perspective of the biologist is to identify ... more The goal of high-throughput screening (HTS) from the perspective of the biologist is to identify a highly specific small molecule that can be used to inhibit a protein in its normal biological context. Although several useful small molecules have been identified with HTS, there are many challenges to be considered when contemplating a screen, especially by those unfamiliar with chemical biology.

Biophysical Journal, 2014

Science, 1995

Import of proteins into the nucleus is a two-step process, involving nuclear localization sequenc... more Import of proteins into the nucleus is a two-step process, involving nuclear localization sequence (NLS)-dependent docking of the substrate at the nuclear envelope followed by translocation through the nuclear pore. A recombinant human protein, hSRP1alpha, bound in vitro specifically and directly to substrates containing either a simple or bipartite NLS motif. hSRP1alpha promoted docking of import substrates to the nuclear

Proceedings of the National Academy of Sciences, 1999

Mitosis is triggered in vertebrate cells by the cyclin B1-Cdc2 complex. The activation of this co... more Mitosis is triggered in vertebrate cells by the cyclin B1-Cdc2 complex. The activation of this complex at the end of G2 phase is accompanied by its translocation from the cytoplasm to the nucleus. We used digitonin-permeabilized human cells to analyze the mechanism by which cyclin B1-Cdc2 is imported into the nucleus. Cyclin B1-Cdc2 import was not blocked by inhibitors of the importin alpha-dependent import pathway or by dominant negative versions of the GTPase Ran or importin beta. However, the rate of cyclin B1 import was decreased by immunodepletion of importin beta from cytosol. Purified importin beta promoted cyclin B1 import in the absence of cytosol or Ran and in the presence of the dominant negative Ran mutant. We conclude that cyclin B1 import is mediated by an unusual importin beta-dependent mechanism that does not require Ran.

Cytosolic proteins bearing a classical nuclear localization signal enter the nucleus bound to a h... more Cytosolic proteins bearing a classical nuclear localization signal enter the nucleus bound to a heterodimer of importin-␣ and importin-␤ (also called karyopherin-␣ and -␤). The formation of this heterodimer involves the importin-␤-binding (IBB) domain of importin-␣, a highly basic amino-terminal region of roughly 40 amino-acid residues. Here we report the crystal structure of human importin-␤ bound to the IBB domain of importin-␣, determined at 2.5 Å and 2.3 Å resolution in two crystal forms. Importin-␤ consists of 19 tandemly repeated HEAT motifs and wraps intimately around the IBB domain. The association involves two separate regions of importin-␤, recognizing structurally distinct parts of the IBB domain: an amino-terminal extended moiety and a carboxy-terminal helix. The structure indicates that significant conformational changes occur when importin-␤ binds or releases the IBB domain domain and suggests how dissociation of the importin-␣/␤ heterodimer may be achieved upon nuclear entry.

Genome biology, 2001

In recent years, our understanding of macromolecular transport processes across the nuclear envel... more In recent years, our understanding of macromolecular transport processes across the nuclear envelope has grown dramatically, and a large number of soluble transport receptors mediating either nuclear import or nuclear export have been identified. Most of these receptors belong to one large family of proteins, all of which share homology with the protein import receptor importin beta (also named karyopherin beta). Members of this family have been classified as importins or exportins on the basis of the direction they carry their cargo. To date, the family includes 14 members in the yeast Saccharomyces cerevisiae and at least 22 members in humans. Importins and exportins are regulated by the small GTPase Ran, which is thought to be highly enriched in the nucleus in its GTP-bound form. Importins recognize their substrates in the cytoplasm and transport them through nuclear pores into the nucleus. In the nucleoplasm, RanGTP binds to importins, inducing the release of import cargoes. In ...

Proceedings of the National Academy of Sciences, 1999

Transport of macromolecules across the nuclear envelope is an active process that depends on solu... more Transport of macromolecules across the nuclear envelope is an active process that depends on soluble factors including the GTPase Ran. Ran-GTP is predominantly located in the nucleus and has been shown to regulate cargo binding and release of import and export receptors in their respective target compartments. Recently, it was shown that transport of receptor-cargo complexes across the nuclear pore complex (NPC) does not depend on GTP-hydrolysis by Ran; however, the mechanism of translocation is still poorly understood. Here, we show that the direction of transport through the NPC can be inverted in the presence of high concentrations of cytoplasmic Ran-GTP. Under these conditions, two different classes of export cargoes are transported into the nucleus in the absence of GTP hydrolysis. The inverted transport is very rapid and can be blocked by known inhibitors of nuclear protein export. These results suggest that the NPC functions as a facilitated transport channel, allowing the selective translocation of receptor-cargo complexes. We conclude that the directionality of nucleocytoplasmic transport is determined mainly by the compartmentalized distribution of Ran-GTP.

Cell, 1994

Acute promyelocytic leukemia (APL) is characterized by a specific t(15;17) translocation that fus... more Acute promyelocytic leukemia (APL) is characterized by a specific t(15;17) translocation that fuses the retinoic acid receptor alpha (RAR alpha) to a novel gene product, PML. The involvement of RAR alpha is particularly intriguing in view of the efficient therapeutic effect of retinoic acid (RA) in this disease. In this report, we show that PML is specifically localized within a discrete subnuclear compartment corresponding to nuclear bodies recognized by patient autoimmune sera. In APL cells, the PML-RAR alpha hybrid displays an abnormal localization and directs RXR and other nuclear antigens into aberrant structures that are tightly bound to chromatin. This suggests that the hybrid could exert a dominant negative effect by diverting a subset of proteins from their natural sites of action. Interestingly, treatment of APL cells with RA induces a complete relocalization of each of these proteins. We propose that the beneficial role of RA in promoting myeloid differentiation in APL might be related to its ability to restore a normal subnuclear organization.

The Journal of Cell Biology, 1996

Kinetic competition experiments have demonstrated that at least some factors required for the nuc... more Kinetic competition experiments have demonstrated that at least some factors required for the nuclear import of proteins and U snRNPs are distinct. Both import processes require energy, and in the case of protein import, the energy requirement is known to be at least partly met by GTP hydrolysis by the Ran GTPase. We have compared the effects of nonhydrolyzable GTP analogues and two mutant Ran proteins on the nuclear import of proteins and U snRNPs in vitro. The mutant Ran proteins have different defects; Q69L (glutamine 69 changed to leucine) is defective in GTP hydrolysis while T24N (threonine 24 changed to asparagine) is defective in binding GTP. Both protein and snRNP import are sensitive either to the presence of the two mutant Ran proteins, which act as dominant negative inhibitors of nuclear import, or to incubation with nonhydrolyzable GTP analogues. This demonstrates that there is a requirement for a GTPase activity for the import of U snRNPs, as well as proteins, into the nucleus. The dominant negative effects of the two mutant Ran proteins indicate that the pathways of protein and snRNP import share at least one common component.

Journal of Cell Biology, 1998

While the Vpr protein of HIV-1 has been implicated in import of the viral preintegration complex ... more While the Vpr protein of HIV-1 has been implicated in import of the viral preintegration complex across the nuclear pore complex (NPC) of nondividing cellular hosts, the mechanism by which Vpr enters the nucleus remains unknown. We now demonstrate that Vpr contains two discrete nuclear targeting signals that use two different import pathways, both of which are distinct from the classical nuclear localization signal (NLS)-and the M9-dependent pathways. Vpr import does not appear to require Ran-mediated GTP hydrolysis and persists under conditions of low energy. Competition experiments further suggest that Vpr directly engages the NPC at two discrete sites. These sites appear to form distal components of a common import pathway used by NLS-and M9-containing proteins. Together, our data suggest that Vpr bypasses many of the soluble receptors involved in import of cellular cargoes. Rather, this viral protein appears to directly access the NPC, a property that may help to ensure the capacity of HIV to replicate in nondividing cellular hosts.

Trends in Biochemical Sciences, 1998

The EMBO Journal, 2002

An important control step in the regulation of cytoplasmic mRNA turnover is the removal of the m ... more An important control step in the regulation of cytoplasmic mRNA turnover is the removal of the m 7 G cap structure at the 5¢ end of the message. Here, we describe the functional characterization of Dhh1, a highly conserved member of the family of DEAD box-containing proteins, as a regulator of mRNA decapping in Saccharomyces cerevisiae. Dhh1 is a cytoplasmic protein and is shown to be in a complex with the mRNA degradation factor Pat1/Mtr1 and with the 5¢±3¢ exoribonuclease Xrn1. Dhh1 speci®cally affects mRNA turnover in the deadenylation-dependent decay pathway, but does not act on the degradation of nonsense-containing mRNAs. Cells that lack dhh1 accumulate degradation intermediates that have lost their poly(A) tail but contain an intact 5¢ cap structure, suggesting that Dhh1 is required for ef®cient decapping in vivo. Furthermore, recombinant Dhh1 is able to stimulate the activity of the puri®ed decapping enzyme Dcp1 in an in vitro decapping assay. We propose that the DEAD box protein Dhh1 regulates the access of the decapping enzyme to the m 7 G cap by modulating the structure at the 5¢ end of mRNAs. Keywords: Dcp1/DEAD box helicase/Dhh1/mRNA decapping/mRNA turnover

The Journal of Cell Biology, 2004

ucleocytoplasmic transport occurs through gigantic proteinaceous channels called nuclear pore com... more ucleocytoplasmic transport occurs through gigantic proteinaceous channels called nuclear pore complexes (NPCs). Translocation through the NPC is exquisitely selective and is mediated by interactions between soluble transport carriers and insoluble NPC proteins that contain phenylalanine-glycine (FG) repeats. Although most FG nucleoporins (Nups) are organized symmetrically about the planar axis of the nuclear envelope, very few localize exclusively to one side of the NPC. We constructed Saccharomyces cerevisiae mutants with N asymmetric FG repeats either deleted or swapped to generate NPCs with inverted FG asymmetry. The mutant Nups localize properly within the NPC and exhibit exchanged binding specificity for the export factor Xpo1. Surprisingly, we were unable to detect any defects in the Kap95, Kap121, Xpo1, or mRNA transport pathways in cells expressing the mutant FG Nups. These findings suggest that the biased distribution of FG repeats is not required for major nucleocytoplasmic trafficking events across the NPC.

Journal of Biological Chemistry, 2005

Coilin is a marker protein for the Cajal body, a subnuclear domain acting as a site for assembly ... more Coilin is a marker protein for the Cajal body, a subnuclear domain acting as a site for assembly and maturation of nuclear RNA-protein complexes. Using a yeast two-hybrid screen to identify coilin-interacting proteins, we have identified hCINAP (human coilin interacting nuclear ATPase protein), a nuclear factor of 172 amino acids with a P-loop nucleotide binding motif and ATPase activity. The hCINAP protein sequence is highly conserved across its full-length from human to plants and yeast and is ubiquitously expressed in all human tissues and cell lines tested. The yeast orthologue of CINAP is a single copy, essential gene. Tagged hCINAP is present in complexes containing coilin in mammalian cells and recombinant, Escherichia coli expressed hCINAP binds directly to coilin in vitro. The 214 carboxyl-terminal residues of coilin appear essential for the interaction with hCINAP. Both immunofluorescence and fluorescent protein tagging show that hCINAP is specifically nuclear and distributed in a widespread, diffuse nucleoplasmic pattern, excluding nucleoli, with some concentration also in Cajal bodies. Overexpression of hCINAP in HeLa cells results in a decrease in the average number of Cajal bodies per nucleus, consistent with it affecting either the stability of Cajal bodies and/or their rate of assembly. The hCINAP mRNA is an alternatively spliced transcript from the TAF9 locus, which encodes the basal transcription factor subunit TAFIID 32 . However, hCINAP and TAFIID 32 mRNAs are translated from different ATG codons and use distinct reading frames, resulting in them having no identity in their respective protein sequences.

Chromosoma, 2006

... 2000). About a third of the nucleoporins belong to the so-called FG Nup subclass, defined by ... more ... 2000). About a third of the nucleoporins belong to the so-called FG Nup subclass, defined by the presence of domains rich in ... 2004). Nup159 is an FG-containing Nup that localizes to the cytoplasmic fibrils of the NPC and is required for mRNA export (Gorsch et al. ...

Current Opinion in Cell Biology, 2011

Nuclear pore complexes (NPCs) are highly selective transport gates that enable the bi-directional... more Nuclear pore complexes (NPCs) are highly selective transport gates that enable the bi-directional traffic of macromolecules across the nuclear envelope (NE). NPCs are located at the fusion pores between the inner and outer membranes of the NE and are built from a common set of 30 different proteins, nucleoporins. Remarkably, recent proteomic, bioinformatic, and structural studies have provided firm evidence that key structural nucleoporins share common ancestry with elements of coated vesicles, indicating an evolutionary link between these structures. This has provided novel insight into the origin of NPCs and may help us to better functionally characterize these fundamental components of eukaryotic cells.

Cell, 2007

In this issue, provide new insight into the selective barrier that controls protein traffic throu... more In this issue, provide new insight into the selective barrier that controls protein traffic through the nuclear pore complex. They show that a single protein domain of the nuclear pore protein Nsp1 can form a hydrogel that allows highly selective access of nuclear transport receptors and their cargos, but rejects other proteins of similar size.

Methods in Enzymology, 2000

Current Molecular Medicine, 2001

Molecular biological investigations of HIV have made fundamental contributions to our understandi... more Molecular biological investigations of HIV have made fundamental contributions to our understanding of eukaryotic biology. These studies elucidated new paradigms in transcription, RNA and protein export from the nucleus to the cytoplasm, cellular activation, morphology and vesicular trafficking.

Nature Chemical Biology, 2006

The goal of high-throughput screening (HTS) from the perspective of the biologist is to identify ... more The goal of high-throughput screening (HTS) from the perspective of the biologist is to identify a highly specific small molecule that can be used to inhibit a protein in its normal biological context. Although several useful small molecules have been identified with HTS, there are many challenges to be considered when contemplating a screen, especially by those unfamiliar with chemical biology.

Biophysical Journal, 2014

Science, 1995

Import of proteins into the nucleus is a two-step process, involving nuclear localization sequenc... more Import of proteins into the nucleus is a two-step process, involving nuclear localization sequence (NLS)-dependent docking of the substrate at the nuclear envelope followed by translocation through the nuclear pore. A recombinant human protein, hSRP1alpha, bound in vitro specifically and directly to substrates containing either a simple or bipartite NLS motif. hSRP1alpha promoted docking of import substrates to the nuclear

Proceedings of the National Academy of Sciences, 1999

Mitosis is triggered in vertebrate cells by the cyclin B1-Cdc2 complex. The activation of this co... more Mitosis is triggered in vertebrate cells by the cyclin B1-Cdc2 complex. The activation of this complex at the end of G2 phase is accompanied by its translocation from the cytoplasm to the nucleus. We used digitonin-permeabilized human cells to analyze the mechanism by which cyclin B1-Cdc2 is imported into the nucleus. Cyclin B1-Cdc2 import was not blocked by inhibitors of the importin alpha-dependent import pathway or by dominant negative versions of the GTPase Ran or importin beta. However, the rate of cyclin B1 import was decreased by immunodepletion of importin beta from cytosol. Purified importin beta promoted cyclin B1 import in the absence of cytosol or Ran and in the presence of the dominant negative Ran mutant. We conclude that cyclin B1 import is mediated by an unusual importin beta-dependent mechanism that does not require Ran.

Cytosolic proteins bearing a classical nuclear localization signal enter the nucleus bound to a h... more Cytosolic proteins bearing a classical nuclear localization signal enter the nucleus bound to a heterodimer of importin-␣ and importin-␤ (also called karyopherin-␣ and -␤). The formation of this heterodimer involves the importin-␤-binding (IBB) domain of importin-␣, a highly basic amino-terminal region of roughly 40 amino-acid residues. Here we report the crystal structure of human importin-␤ bound to the IBB domain of importin-␣, determined at 2.5 Å and 2.3 Å resolution in two crystal forms. Importin-␤ consists of 19 tandemly repeated HEAT motifs and wraps intimately around the IBB domain. The association involves two separate regions of importin-␤, recognizing structurally distinct parts of the IBB domain: an amino-terminal extended moiety and a carboxy-terminal helix. The structure indicates that significant conformational changes occur when importin-␤ binds or releases the IBB domain domain and suggests how dissociation of the importin-␣/␤ heterodimer may be achieved upon nuclear entry.

Genome biology, 2001

In recent years, our understanding of macromolecular transport processes across the nuclear envel... more In recent years, our understanding of macromolecular transport processes across the nuclear envelope has grown dramatically, and a large number of soluble transport receptors mediating either nuclear import or nuclear export have been identified. Most of these receptors belong to one large family of proteins, all of which share homology with the protein import receptor importin beta (also named karyopherin beta). Members of this family have been classified as importins or exportins on the basis of the direction they carry their cargo. To date, the family includes 14 members in the yeast Saccharomyces cerevisiae and at least 22 members in humans. Importins and exportins are regulated by the small GTPase Ran, which is thought to be highly enriched in the nucleus in its GTP-bound form. Importins recognize their substrates in the cytoplasm and transport them through nuclear pores into the nucleus. In the nucleoplasm, RanGTP binds to importins, inducing the release of import cargoes. In ...

Proceedings of the National Academy of Sciences, 1999

Transport of macromolecules across the nuclear envelope is an active process that depends on solu... more Transport of macromolecules across the nuclear envelope is an active process that depends on soluble factors including the GTPase Ran. Ran-GTP is predominantly located in the nucleus and has been shown to regulate cargo binding and release of import and export receptors in their respective target compartments. Recently, it was shown that transport of receptor-cargo complexes across the nuclear pore complex (NPC) does not depend on GTP-hydrolysis by Ran; however, the mechanism of translocation is still poorly understood. Here, we show that the direction of transport through the NPC can be inverted in the presence of high concentrations of cytoplasmic Ran-GTP. Under these conditions, two different classes of export cargoes are transported into the nucleus in the absence of GTP hydrolysis. The inverted transport is very rapid and can be blocked by known inhibitors of nuclear protein export. These results suggest that the NPC functions as a facilitated transport channel, allowing the selective translocation of receptor-cargo complexes. We conclude that the directionality of nucleocytoplasmic transport is determined mainly by the compartmentalized distribution of Ran-GTP.

Cell, 1994

Acute promyelocytic leukemia (APL) is characterized by a specific t(15;17) translocation that fus... more Acute promyelocytic leukemia (APL) is characterized by a specific t(15;17) translocation that fuses the retinoic acid receptor alpha (RAR alpha) to a novel gene product, PML. The involvement of RAR alpha is particularly intriguing in view of the efficient therapeutic effect of retinoic acid (RA) in this disease. In this report, we show that PML is specifically localized within a discrete subnuclear compartment corresponding to nuclear bodies recognized by patient autoimmune sera. In APL cells, the PML-RAR alpha hybrid displays an abnormal localization and directs RXR and other nuclear antigens into aberrant structures that are tightly bound to chromatin. This suggests that the hybrid could exert a dominant negative effect by diverting a subset of proteins from their natural sites of action. Interestingly, treatment of APL cells with RA induces a complete relocalization of each of these proteins. We propose that the beneficial role of RA in promoting myeloid differentiation in APL might be related to its ability to restore a normal subnuclear organization.

The Journal of Cell Biology, 1996

Kinetic competition experiments have demonstrated that at least some factors required for the nuc... more Kinetic competition experiments have demonstrated that at least some factors required for the nuclear import of proteins and U snRNPs are distinct. Both import processes require energy, and in the case of protein import, the energy requirement is known to be at least partly met by GTP hydrolysis by the Ran GTPase. We have compared the effects of nonhydrolyzable GTP analogues and two mutant Ran proteins on the nuclear import of proteins and U snRNPs in vitro. The mutant Ran proteins have different defects; Q69L (glutamine 69 changed to leucine) is defective in GTP hydrolysis while T24N (threonine 24 changed to asparagine) is defective in binding GTP. Both protein and snRNP import are sensitive either to the presence of the two mutant Ran proteins, which act as dominant negative inhibitors of nuclear import, or to incubation with nonhydrolyzable GTP analogues. This demonstrates that there is a requirement for a GTPase activity for the import of U snRNPs, as well as proteins, into the nucleus. The dominant negative effects of the two mutant Ran proteins indicate that the pathways of protein and snRNP import share at least one common component.

Journal of Cell Biology, 1998

While the Vpr protein of HIV-1 has been implicated in import of the viral preintegration complex ... more While the Vpr protein of HIV-1 has been implicated in import of the viral preintegration complex across the nuclear pore complex (NPC) of nondividing cellular hosts, the mechanism by which Vpr enters the nucleus remains unknown. We now demonstrate that Vpr contains two discrete nuclear targeting signals that use two different import pathways, both of which are distinct from the classical nuclear localization signal (NLS)-and the M9-dependent pathways. Vpr import does not appear to require Ran-mediated GTP hydrolysis and persists under conditions of low energy. Competition experiments further suggest that Vpr directly engages the NPC at two discrete sites. These sites appear to form distal components of a common import pathway used by NLS-and M9-containing proteins. Together, our data suggest that Vpr bypasses many of the soluble receptors involved in import of cellular cargoes. Rather, this viral protein appears to directly access the NPC, a property that may help to ensure the capacity of HIV to replicate in nondividing cellular hosts.

Trends in Biochemical Sciences, 1998

The EMBO Journal, 2002

An important control step in the regulation of cytoplasmic mRNA turnover is the removal of the m ... more An important control step in the regulation of cytoplasmic mRNA turnover is the removal of the m 7 G cap structure at the 5¢ end of the message. Here, we describe the functional characterization of Dhh1, a highly conserved member of the family of DEAD box-containing proteins, as a regulator of mRNA decapping in Saccharomyces cerevisiae. Dhh1 is a cytoplasmic protein and is shown to be in a complex with the mRNA degradation factor Pat1/Mtr1 and with the 5¢±3¢ exoribonuclease Xrn1. Dhh1 speci®cally affects mRNA turnover in the deadenylation-dependent decay pathway, but does not act on the degradation of nonsense-containing mRNAs. Cells that lack dhh1 accumulate degradation intermediates that have lost their poly(A) tail but contain an intact 5¢ cap structure, suggesting that Dhh1 is required for ef®cient decapping in vivo. Furthermore, recombinant Dhh1 is able to stimulate the activity of the puri®ed decapping enzyme Dcp1 in an in vitro decapping assay. We propose that the DEAD box protein Dhh1 regulates the access of the decapping enzyme to the m 7 G cap by modulating the structure at the 5¢ end of mRNAs. Keywords: Dcp1/DEAD box helicase/Dhh1/mRNA decapping/mRNA turnover

The Journal of Cell Biology, 2004

ucleocytoplasmic transport occurs through gigantic proteinaceous channels called nuclear pore com... more ucleocytoplasmic transport occurs through gigantic proteinaceous channels called nuclear pore complexes (NPCs). Translocation through the NPC is exquisitely selective and is mediated by interactions between soluble transport carriers and insoluble NPC proteins that contain phenylalanine-glycine (FG) repeats. Although most FG nucleoporins (Nups) are organized symmetrically about the planar axis of the nuclear envelope, very few localize exclusively to one side of the NPC. We constructed Saccharomyces cerevisiae mutants with N asymmetric FG repeats either deleted or swapped to generate NPCs with inverted FG asymmetry. The mutant Nups localize properly within the NPC and exhibit exchanged binding specificity for the export factor Xpo1. Surprisingly, we were unable to detect any defects in the Kap95, Kap121, Xpo1, or mRNA transport pathways in cells expressing the mutant FG Nups. These findings suggest that the biased distribution of FG repeats is not required for major nucleocytoplasmic trafficking events across the NPC.

Journal of Biological Chemistry, 2005

Coilin is a marker protein for the Cajal body, a subnuclear domain acting as a site for assembly ... more Coilin is a marker protein for the Cajal body, a subnuclear domain acting as a site for assembly and maturation of nuclear RNA-protein complexes. Using a yeast two-hybrid screen to identify coilin-interacting proteins, we have identified hCINAP (human coilin interacting nuclear ATPase protein), a nuclear factor of 172 amino acids with a P-loop nucleotide binding motif and ATPase activity. The hCINAP protein sequence is highly conserved across its full-length from human to plants and yeast and is ubiquitously expressed in all human tissues and cell lines tested. The yeast orthologue of CINAP is a single copy, essential gene. Tagged hCINAP is present in complexes containing coilin in mammalian cells and recombinant, Escherichia coli expressed hCINAP binds directly to coilin in vitro. The 214 carboxyl-terminal residues of coilin appear essential for the interaction with hCINAP. Both immunofluorescence and fluorescent protein tagging show that hCINAP is specifically nuclear and distributed in a widespread, diffuse nucleoplasmic pattern, excluding nucleoli, with some concentration also in Cajal bodies. Overexpression of hCINAP in HeLa cells results in a decrease in the average number of Cajal bodies per nucleus, consistent with it affecting either the stability of Cajal bodies and/or their rate of assembly. The hCINAP mRNA is an alternatively spliced transcript from the TAF9 locus, which encodes the basal transcription factor subunit TAFIID 32 . However, hCINAP and TAFIID 32 mRNAs are translated from different ATG codons and use distinct reading frames, resulting in them having no identity in their respective protein sequences.

Chromosoma, 2006

... 2000). About a third of the nucleoporins belong to the so-called FG Nup subclass, defined by ... more ... 2000). About a third of the nucleoporins belong to the so-called FG Nup subclass, defined by the presence of domains rich in ... 2004). Nup159 is an FG-containing Nup that localizes to the cytoplasmic fibrils of the NPC and is required for mRNA export (Gorsch et al. ...

Current Opinion in Cell Biology, 2011

Nuclear pore complexes (NPCs) are highly selective transport gates that enable the bi-directional... more Nuclear pore complexes (NPCs) are highly selective transport gates that enable the bi-directional traffic of macromolecules across the nuclear envelope (NE). NPCs are located at the fusion pores between the inner and outer membranes of the NE and are built from a common set of 30 different proteins, nucleoporins. Remarkably, recent proteomic, bioinformatic, and structural studies have provided firm evidence that key structural nucleoporins share common ancestry with elements of coated vesicles, indicating an evolutionary link between these structures. This has provided novel insight into the origin of NPCs and may help us to better functionally characterize these fundamental components of eukaryotic cells.

Cell, 2007

In this issue, provide new insight into the selective barrier that controls protein traffic throu... more In this issue, provide new insight into the selective barrier that controls protein traffic through the nuclear pore complex. They show that a single protein domain of the nuclear pore protein Nsp1 can form a hydrogel that allows highly selective access of nuclear transport receptors and their cargos, but rejects other proteins of similar size.