Exit from Mitosis Is Triggered by Tem1-Dependent Release of the Protein Phosphatase Cdc14 from Nucleolar RENT Complex
TL;DR: A mutation is identified, net1-1, that bypasses the lethality of tem1 delta and is a key component of a multifunctional complex, denoted RENT (for regulator of nucleolar silencing and telophase), that also contains Cdc14 and the silencing regulator Sir2.
About: This article is published in Cell.The article was published on 1999-04-16 and is currently open access. It has received 758 citations till now. The article focuses on the topics: RENT complex & Mitotic exit.
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TL;DR: The construction and analysis of a collection of yeast strains expressing full-length, chromosomally tagged green fluorescent protein fusion proteins helps reveal the logic of transcriptional co-regulation, and provides a comprehensive view of interactions within and between organelles in eukaryotic cells.
Abstract: A fundamental goal of cell biology is to define the functions of proteins in the context of compartments that organize them in the cellular environment. Here we describe the construction and analysis of a collection of yeast strains expressing full-length, chromosomally tagged green fluorescent protein fusion proteins. We classify these proteins, representing 75% of the yeast proteome, into 22 distinct subcellular localization categories, and provide localization information for 70% of previously unlocalized proteins. Analysis of this high-resolution, high-coverage localization data set in the context of transcriptional, genetic, and protein-protein interaction data helps reveal the logic of transcriptional co-regulation, and provides a comprehensive view of interactions within and between organelles in eukaryotic cells.
4,310 citations
TL;DR: Although the nucleolus is primarily associated with ribosome biogenesis, several lines of evidence now show that it has additional functions, such as regulation of mitosis, cell-cycle progression and proliferation, many forms of stress response and biogenesis of multiple ribonucleoprotein particles.
Abstract: The nucleolus is a distinct subnuclear compartment that was first observed more than 200 years ago. Nucleoli assemble around the tandemly repeated ribosomal DNA gene clusters and 28S, 18S and 5.8S ribosomal RNAs (rRNAs) are transcribed as a single precursor, which is processed and assembled with the 5S rRNA into ribosome subunits. Although the nucleolus is primarily associated with ribosome biogenesis, several lines of evidence now show that it has additional functions. Some of these functions, such as regulation of mitosis, cell-cycle progression and proliferation, many forms of stress response and biogenesis of multiple ribonucleoprotein particles, will be discussed, as will the relation of the nucleolus to human diseases.
1,353 citations
TL;DR: It is proposed that binding of resveratrol to SIRT1 promotes a conformational change that better accommodates the attached coumarin group.
994 citations
TL;DR: Discovery of an intrinsic deacetylation activity for the conserved SIR2 family provides a mechanism for modifying histones and other proteins to regulate transcription and diverse biological processes.
Abstract: Homologs of the chromatin-bound yeast silent information regulator 2 (SIR2) protein are found in organisms from all biological kingdoms. SIR2 itself was originally discovered to influence mating-type control in haploid cells by locus-specific transcriptional silencing. Since then, SIR2 and its homologs have been suggested to play additional roles in suppression of recombination, chromosomal stability, metabolic regulation, meiosis, and aging. Considering the far-ranging nature of these functions, a major experimental goal has been to understand the molecular mechanism(s) by which this family of proteins acts. We report here that members of the SIR2 family catalyze an NAD–nicotinamide exchange reaction that requires the presence of acetylated lysines such as those found in the N termini of histones. Significantly, these enzymes also catalyze histone deacetylation in a reaction that absolutely requires NAD, thereby distinguishing them from previously characterized deacetylases. The enzymes are active on histone substrates that have been acetylated by both chromatin assembly-linked and transcription-related acetyltransferases. Contrary to a recent report, we find no evidence that these proteins ADP-ribosylate histones. Discovery of an intrinsic deacetylation activity for the conserved SIR2 family provides a mechanism for modifying histones and other proteins to regulate transcription and diverse biological processes.
971 citations
TL;DR: Key events in mitosis such as sister chromatid separation and subsequent inactivation of cyclin-dependent kinase 1 are regulated by ubiquitin- dependent proteolysis, mediated by the anaphase-promoting complex.
949 citations
Cites background from "Exit from Mitosis Is Triggered by T..."
...…leased, Cdc14 dephosphorylates at least three proteins, Cdh1, Sic1, and the transcription factor Swi5, all of which and cyclin A-Cdk2 have been reported as the major contribute to CDK inactivation (Shou et al., 1999; Visintin Cdh1 kinases, respectively (Lukas et al., 1999; Sigrist et al., 1998)....
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References
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Book•
01 Jan 1988
TL;DR: A second edition of Antibodies: A Laboratory Manual is being published in September 2013, Revised, extended and updated by Edward Greenfield of the Dana-Farber Cancer Center, the material has been recast with extensive new information and new chapters have been added.
Abstract: ince its publication in 1988, Antibodies: A Laboratory Manual, by Harlow and Lane, has become a classic, an essential resource for molecular biology, immunology, and cell culture labs. In order to keep the book in print, Cold Spring Harbor Laboratory Press eventually produced the paperback edition currently available for sale. Now, after 25 years, a second edition is being published in September 2013. Revised, extended and updated by Edward Greenfield of the Dana-Farber Cancer Center, the material has been recast with extensive new information and new chapters have been added. The new edition provides clear, authoritative, current and up-to-date protocols with background information and troubleshooting advice. The book is an invaluable resource for all those engaged in antibody research and development.
22,254 citations
TL;DR: Silver staining allows a substantial shortening of sample preparation time and may, therefore, be preferable over Coomassie staining, and this work removes a major obstacle to the low-level sequence analysis of proteins separated on polyacrylamide gels.
Abstract: Proteins from silver-stained gels can be digested enzymatically and the resulting peptides analyzed and sequenced by mass spectrometry. Standard proteins yield the same peptide maps when extracted from Coomassie- and silver-stained gels, as judged by electrospray and MALDI mass spectrometry. The low nanogram range can be reached by the protocols described here, and the method is robust. A silver-stained one-dimensional gel of a fraction from yeast proteins was analyzed by nanoelectrospray tandem mass spectrometry. In the sequencing, more than 1000 amino acids were covered, resulting in no evidence of chemical modifications due to the silver staining procedure. Silver staining allows a substantial shortening of sample preparation time and may, therefore, be preferable over Coomassie staining. This work removes a major obstacle to the low-level sequence analysis of proteins separated on polyacrylamide gels.
8,437 citations
"Exit from Mitosis Is Triggered by T..." refers background or methods in this paper
...…CA59935, and R. J. D. was supported by Junior Investigatorracy MALDI peptide mapping on a modified REFLEX mass specAwards from the Searle/Chicago Community Trust and the Beckmantrometer (Bruker Daltonics, Bremen, Germany) essentially as deFoundation.scribed previously (Shevchenko et al., 1996b)....
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...W. S. is a Howard Hughesdescribed previously (Shevchenko et al., 1996b)....
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...Nevertheless, eventually Clb2 bined in a layered approach (Shevchenko et al., 1996a). was completely degraded and Sic1 accumulated to high One Net1-interacting protein (Cdc14) was identified by levels (Figure 1B, right panels)....
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TL;DR: A series of yeast shuttle vectors and host strains has been created to allow more efficient manipulation of DNA in Saccharomyces cerevisiae to perform most standard DNA manipulations in the same plasmid that is introduced into yeast.
Abstract: A series of yeast shuttle vectors and host strains has been created to allow more efficient manipulation of DNA in Saccharomyces cerevisiae. Transplacement vectors were constructed and used to derive yeast strains containing nonreverting his3, trp1, leu2 and ura3 mutations. A set of YCp and YIp vectors (pRS series) was then made based on the backbone of the multipurpose plasmid pBLUESCRIPT. These pRS vectors are all uniform in structure and differ only in the yeast selectable marker gene used (HIS3, TRP1, LEU2 and URA3). They possess all of the attributes of pBLUESCRIPT and several yeast-specific features as well. Using a pRS vector, one can perform most standard DNA manipulations in the same plasmid that is introduced into yeast.
8,364 citations
Journal Article•
7,587 citations
TL;DR: A simple and robust technique for the sequencing of proteins isolated by polyacrylamide gel electro-phoresis, using nano-electrospray3,4 tandem mass spectrometry5,6 and multiple-sequence stretches of up to 16 amino acids are obtained.
Abstract: Molecular analysis of complex biological structures and processes increasingly requires sensitive methods for protein sequencing. Electrospray mass spectrometry has been applied to the high-sensitivity sequencing of short peptides, but technical difficulties have prevented similar success with gel-isolated proteins. Here we report a simple and robust technique for the sequencing of proteins isolated by polyacrylamide gel electrophoresis, using nano-electrospray tandem mass spectrometry. As little as 5 ng protein starting material on Coomassie- or silver-stained gels can be sequenced. Multiple-sequence stretches of up to 16 amino acids are obtained, which identify the protein unambiguously if already present in databases or provide information to clone the corresponding gene. We have applied this method to the sequencing and cloning of a protein which inhibits the proliferation of capillary endothelial cells in vitro and thus may have potential antiangiogenic effects on solid tumours.
1,695 citations
"Exit from Mitosis Is Triggered by T..." refers background in this paper
...Identification and characterizationquadruple instrument (PE Sciex, Concord, ON, Canada) as deof a yeast nucleolar protein that is similar to a rat liver nucleolarscribed previously (Wilm et al., 1996)....
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