Unconventional tethering of Ulp1 to the transport channel of the nuclear pore complex by karyopherins.

The ubiquitin-like protein SUMO-1 (small ubiquitin-related modifier 1) is covalently attached to substrate proteins by ligases and cleaved by isopeptidases. Yeast has two SUMO-1-deconjugating enzymes, Ulp1 and Ulp2, which are located at nuclear pores and in the nucleoplasm, respectively. Here we show that the catalytic C-domain of Ulp1 must be excluded from the nucleoplasm for cell viability.

A Noc complex specifically involved in the formation and nuclear export of ribosomal 40 S subunits.

Formation and nuclear export of 60 S pre-ribosomes requires many factors including the heterodimeric Noc1-Noc2 and Noc2-Noc3 complexes. Here, we report another Noc complex with a specific role in 40 S subunit biogenesis. This complex consists of Noc4p, which exhibits the conserved Noc domain and is homologous to Noc1p, and Nop14p, a nucleolar protein with a role in 40 S subunit formation.

Analysis of gamma c-family cytokine target genes. Identification of dual-specificity phosphatase 5 (DUSP5) as a regulator of mitogen-activated protein kinase activity in interleukin-2 signaling.

Interleukin (IL)-2, IL-4, IL-7, IL-9, IL-15, and IL-21 form a family of cytokines based on their sharing the common cytokine receptor gamma chain, gamma(c), which is mutated in X-linked severe combined immunodeficiency (SCID). As a step toward further elucidating the mechanism of action of these cytokines in T-cell biology, we compared the gene expression profiles of IL-2, IL-4, IL-7, and IL-15 in T cells using cDNA microarrays. IL-2, IL-7, and IL-15 each induced a highly similar set of genes, whereas IL-4 induced distinct genes correlating with differential STAT protein activation by this cytokine.

The mRNA export machinery requires the novel Sac3p-Thp1p complex to dock at the nucleoplasmic entrance of the nuclear pores.

Yra1p and Sub2p are components of the TREX complex, which couples transcription elongation with nuclear export of mRNAs. Here, we report a genetic interaction between Yra1p and a conserved protein Sac3p, which previously was found to interact with Sub2p. In vivo, Sac3p forms a stable complex with Thp1p, which was reported to function in transcription elongation.

60S pre-ribosome formation viewed from assembly in the nucleolus until export to the cytoplasm.

60S ribosomes undergo initial assembly in the nucleolus before export to the cytoplasm and recent analyses have identified several nucleolar pre-60S particles. To unravel the steps in the pathway of ribosome formation, we have purified the pre-60S ribosomes associated with proteins predicted to act at different stages as the pre-ribosomes transit from the nucleolus through the nucleoplasm and are then exported to the cytoplasm for final maturation. About 50 non-ribosomal proteins are associated with the early nucleolar pre-60S ribosomes.

90S pre-ribosomes include the 35S pre-rRNA, the U3 snoRNP, and 40S subunit processing factors but predominantly lack 60S synthesis factors.

We report the characterization of early pre-ribosomal particles. Twelve TAP-tagged components each showed nucleolar localization, sedimented at approximately 90S on sucrose gradients, and coprecipitated both the 35S pre-rRNA and the U3 snoRNA. Thirty-five non-ribosomal proteins were coprecipitated, including proteins associated with U3 (Nop56p, Nop58p, Sof1p, Rrp9, Dhr1p, Imp3p, Imp4p, and Mpp10p) and other factors required for 18S rRNA synthesis (Nop14p, Bms1p, and Krr1p).

Blimp-1 orchestrates plasma cell differentiation by extinguishing the mature B cell gene expression program.

Blimp-1, a transcriptional repressor, drives the terminal differentiation of B cells to plasma cells. Using DNA microarrays, we found that introduction of Blimp-1 into B cells blocked expression of a remarkably large set of genes, while a much smaller number was induced. Blimp-1 initiated this cascade of gene expression changes by directly repressing genes encoding several transcription factors, including Spi-B and Id3, that regulate signaling by the B cell receptor.

The use of molecular profiling to predict survival after chemotherapy for diffuse large-B-cell lymphoma.

Background: The survival of patients with diffuse large-B-cell lymphoma after chemotherapy is influenced by molecular features of the tumors. We used the gene-expression profiles of these lymphomas to develop a molecular predictor of survival.

Methods: Biopsy samples of diffuse large-B-cell lymphoma from 240 patients were examined for gene expression with the use of DNA microarrays and analyzed for genomic abnormalities.

International guidelines and experiences on health claims in Europe.

The relationship between nutrition and health is gaining public acceptance and consumers are increasingly health-conscious and want to obtain more information about the food they buy. There is a legal void with regard to health claims in the European Union. The framework labelling legislation prohibits 'attributing to any foodstuff the property of preventing, treating or curing a human disease or referring to such properties'.

Nutrition labelling: European Union and United Kingdom perspectives.

The growing public interest in the relationship between diet and health and increasing public health problems in Europe were among the determining factors which led the European Commission to propose harmonized legislation on nutrition labelling. The Directive which was adopted in 1990 primarily aimed at providing information which helps consumers to make an informed choice and assist action in the area of nutrition education for the public. The provisions of the Directive are voluntary but become obligatory if the manufacturer decides to make a 'nutritional claim'.

Rlp7p is associated with 60S preribosomes, restricted to the granular component of the nucleolus, and required for pre-rRNA processing.

Many analyses have examined subnucleolar structures in eukaryotic cells, but the relationship between morphological structures, pre-rRNA processing, and ribosomal particle assembly has remained unclear. Using a visual assay for export of the 60S ribosomal subunit, we isolated a ts-lethal mutation, rix9-1, which causes nucleolar accumulation of an Rpl25p-eGFP reporter construct. The mutation results in a single amino acid substitution (F176S) in Rlp7p, an essential nucleolar protein related to ribosomal protein Rpl7p.

TREX is a conserved complex coupling transcription with messenger RNA export.

The essential yeast proteins Yra1 and Sub2 are messenger RNA export factors that have conserved counterparts in metazoans, designated Aly and UAP56, respectively. These factors couple the machineries that function in splicing and export of mRNA. Here we show that both Yra1 and Sub2 are stoichiometrically associated with the heterotetrameric THO complex, which functions in transcription in yeast.

An intron in the YRA1 gene is required to control Yra1 protein expression and mRNA export in yeast.

Yra1p is an essential and conserved mRNA export factor in yeast. Strikingly, removal of the intron from YRA1 causes a dominant-negative growth phenotype and a concomitant inhibition of mRNA export. However, both defects are neutralized by replacement of the intron of YRA1 by a different intron.

A conserved mRNA export machinery coupled to pre-mRNA splicing.

Recent advances have led to a new understanding of how mRNAs are exported from the nucleus to the cytoplasm. This process requires a heterodimeric mRNA export receptor that is part of an elaborate machinery conserved from yeast to humans. Export of mRNAs is coupled to upstream steps in gene expression, such as pre-mRNA splicing, and to downstream events, including nonsense-mediated decay.

Structure of the C-terminal FG-nucleoporin binding domain of Tap/NXF1.

The vertebrate Tap protein is a member of the NXF family of shuttling transport receptors for nuclear export of mRNA. Tap has a modular structure, and its most C-terminal domain is important for binding to FG repeat-containing nuclear pore proteins (FG-nucleoporins) and is sufficient to mediate nuclear shuttling. We report the solution structure of this C-terminal domain, which is based on a distinctive arrangement of four alpha-helices and is joined to the next module by a flexible 12-residue Pro-rich linker.

Modular self-assembly of a Y-shaped multiprotein complex from seven nucleoporins.

Now that it is likely that all yeast nucleoporins are known, one of the ultimate goals is the in vitro assembly of the entire nuclear pore complex from its approximately 30 individual components. Here, we report the reconstitution of seven proteins (Nup133p, Nup145p-C, Nup120p, Nup85p, Nup84p, Seh1p and Sec13p) into a heptameric 0.5 MDa nuclear pore subcomplex.

The nucle(ol)ar Tif6p and Efl1p are required for a late cytoplasmic step of ribosome synthesis.

Deletion of elongation factor-like 1 (Efl1p), a cytoplasmic GTPase homologous to the ribosomal translocases EF-G/EF-2, results in nucle(ol)ar pre-rRNA processing and pre-60S subunits export defects. Efl1p interacts genetically with Tif6p, a nucle(ol)ar protein stably associated with pre-60S subunits and required for their synthesis and nuclear exit. In the absence of Efl1p, 50% of Tif6p is relocated to the cytoplasm.

Complex formation between Tap and p15 affects binding to FG-repeat nucleoporins and nucleocytoplasmic shuttling.

Mammalian Tap-p15 and yeast Mex67p-Mtr2p are conserved and essential mRNA export factor complexes that transport mRNPs through the nuclear pore. Here, we report that the small subunit p15 affects the binding of the large subunit Tap to repeat nucleoporins. BIAcore measurements revealed that recombinant Tap binds with high affinity (K(d) in the nm range) to repeat nucleoporins and dissociates from them very slowly.

The intracellular location of two aminoacyl-tRNA synthetases depends on complex formation with Arc1p.

In yeast, two aminoacyl-tRNA synthetases, MetRS and GluRS, are associated with Arc1p. We have studied the mechanism of this complex formation and found that the non-catalytic N-terminally appended domains of MetRS and GluRS are necessary and sufficient for binding to Arc1p. Similarly, it is the N-terminal domain of Arc1p that contains distinct but overlapping binding sites for MetRS and GluRS.

The Nsp1p carboxy-terminal domain is organized into functionally distinct coiled-coil regions required for assembly of nucleoporin subcomplexes and nucleocytoplasmic transport.

Nucleoporin Nsp1p, which has four predicted coiled-coil regions (coils 1 to 4) in the essential carboxy-terminal domain, is unique in that it is part of two distinct nuclear pore complex (NPC) subcomplexes, Nsp1p-Nup57p-Nup49p-Nic96p and Nsp1p-Nup82p-Nup159p. As shown by in vitro reconstitution, coiled-coil region 2 (residues 673 to 738) is sufficient to form heterotrimeric core complexes and can bind either Nup57p or Nup82p. Accordingly, interaction of Nup82p with Nsp1p coil 2 is competed by excess Nup57p.

Splicing factor Sub2p is required for nuclear mRNA export through its interaction with Yra1p.

The yeast nuclear protein Yra1p is an essential export factor for mRNA. Yra1p interacts directly with the mRNA transport factor Mex67p/Mtr2p, which is associated with the nuclear pore. Here, we report a genetic interaction between YRA1 and SUB2, the gene for a DEAD box helicase involved in splicing.

Genomic-scale measurement of mRNA turnover and the mechanisms of action of the anti-cancer drug flavopiridol.

Background: Flavopiridol, a flavonoid currently in cancer clinical trials, inhibits cyclin-dependent kinases (CDKs) by competitively blocking their ATP-binding pocket. However, the mechanism of action of flavopiridol as an anti-cancer agent has not been fully elucidated.

Results: Using DNA microarrays, we found that flavopiridol inhibited gene expression broadly, in contrast to two other CDK inhibitors, roscovitine and 9-nitropaullone.

Targeting of Ran: variation on a common theme?

The Ran GTPase plays a key role in nucleocytoplasmic transport. In its GTP-bound form, it directly interacts with members of the importin beta family of nuclear transport receptors and modulates their association with cargo. Work in cell-free higher-eukaryote systems has demonstrated additional roles for Ran in spindle and nuclear envelope formation during mitosis.

Identification of a 60S preribosomal particle that is closely linked to nuclear export.

A nuclear GTPase, Nug1p, was identified in a genetic screen for components linked to 60S ribosomal subunit export. Nug1p cosedimented with nuclear 60S preribosomes and was required for subunit export to the cytoplasm. Tagged Nug1p coprecipitated with proteins of the 60S subunit, late precursors to the 25S and 5.