Results 1 - 10
of
98
Exploration of the function and organization of the yeast early secretory pathway through an epistatic miniarray profile. Cell 123: 507–519
- Cell
, 2005
"... We present a strategy for generating and analyzing comprehensive genetic-interaction maps, termed E-MAPs (epistatic miniarray profiles), comprising quantitative measures of aggravating or alleviating interactions between gene pairs. Crucial to the interpretation of E-MAPs is their high-density natur ..."
Abstract
-
Cited by 88 (8 self)
- Add to MetaCart
(Show Context)
We present a strategy for generating and analyzing comprehensive genetic-interaction maps, termed E-MAPs (epistatic miniarray profiles), comprising quantitative measures of aggravating or alleviating interactions between gene pairs. Crucial to the interpretation of E-MAPs is their high-density nature made possible by focusing on logically connected gene subsets and including essential genes. Described here is the analysis of an E-MAP of genes acting in the yeast early secretory pathway. Hierarchical clustering, together with novel analytical strategies and experimental verification, revealed or clarified the role of many proteins involved in extensively studied processes such as sphingolipid metabolism and retention of HDEL proteins. At a broader level, analysis of the E-MAP delineated pathway organization and components of physical complexes and illustrated the interconnection between the various secretory processes. Extension of this strategy to other logically connected gene subsets in yeast and higher eukaryotes should provide critical insights into the functional/organizational principles of biological systems.
Sgs1 helicase and two nucleases Dna2 and Exo1 resect DNA doublestrand break ends
- Downloaded from http://nar.oxfordjournals.org/ at Pennsylvania State University on February 21, 2013 Nucleic Acids Research, 2009
, 2008
"... Formation of single-strand DNA (ssDNA) tails at a double-strand break (DSB) is a key step in homologous recombination and DNA-damage signaling. The enzyme(s) producing ssDNA at DSBs in eukaryotes remain unknown. We monitored 5 0-strand resection at inducible DSB ends in yeast and identified proteins ..."
Abstract
-
Cited by 83 (1 self)
- Add to MetaCart
(Show Context)
Formation of single-strand DNA (ssDNA) tails at a double-strand break (DSB) is a key step in homologous recombination and DNA-damage signaling. The enzyme(s) producing ssDNA at DSBs in eukaryotes remain unknown. We monitored 5 0-strand resection at inducible DSB ends in yeast and identified proteins required for two stages of resection: initiation and long-range 5 0-strand resection. We show that the Mre11-Rad50-Xrs2 complex (MRX) initiates 5 0 degradation, whereas Sgs1 and Dna2 degrade 5 0 strands exposing long 3 0 strands. Deletion of SGS1 or DNA2 reduces resection and DSB repair by single-strand annealing between distant repeats while the remaining long-range resection activity depends on the exonuclease Exo1. In exo1D sgs1D double mutants, the MRX complex together with Sae2 nuclease generate, in a stepwise manner, only few hundred nucleotides of ssDNA at the break, resulting in inefficient gene conversion and G2/M damage checkpoint arrest. These results provide important insights into the early steps of DSB repair in eukaryotes.
Controlling gene expression in mycobacteria with anhydrotetracycline and Tet repressor. Nucleic Acids Res
, 2005
"... Gene expression systems that allow the regula-tion of bacterial genes during an infection are valuable molecular tools but are lacking for myco-bacterial pathogens. We report the development of mycobacterial gene regulation systems that allow controlling gene expression in fast and slow-growing myco ..."
Abstract
-
Cited by 45 (5 self)
- Add to MetaCart
(Show Context)
Gene expression systems that allow the regula-tion of bacterial genes during an infection are valuable molecular tools but are lacking for myco-bacterial pathogens. We report the development of mycobacterial gene regulation systems that allow controlling gene expression in fast and slow-growing mycobacteria, including Mycobacterium tuberculosis, using anhydrotetracycline (ATc) as inducer. The systems are based on the Escherichia coli Tn10-derived tet regulatory system and con-sist of a strong tet operator (tetO)-containing myco-bacterial promoter, expression cassettes for the repressor TetR and the chemical inducer ATc. These systems allow gene regulation over two orders of magnitude in Mycobacterium smegmatis and M.tuberculosis. TetR-controlled gene expression was inducer concentration-dependent and maximal with ATc concentrations at least 10- and 20-fold below the minimal inhibitory concentration for M.smegmatis and M.tuberculosis, respectively. Using the essential mycobacterial gene ftsZ, we showed that these expression systems can be used to construct conditional knockouts and to analyze the function of essential mycobacterial genes. Finally, we demonstrated that these systems allow gene regulation in M.tuberculosis within the macro-phage phagosome.
Mitochondrial protein import: from proteomics to functional mechanisms
- Nat. Rev. Mol. Cell Biol
, 2010
"... Abstract | Mitochondria contain ~1,000 different proteins, most of which are imported from the cytosol. Two import pathways that direct proteins into the mitochondrial inner membrane and matrix have been known for many years. The identification of numerous new transport components in recent proteom ..."
Abstract
-
Cited by 29 (3 self)
- Add to MetaCart
Abstract | Mitochondria contain ~1,000 different proteins, most of which are imported from the cytosol. Two import pathways that direct proteins into the mitochondrial inner membrane and matrix have been known for many years. The identification of numerous new transport components in recent proteomic studies has led to novel mechanistic insight into these pathways and the discovery of new import pathways into the outer membrane and intermembrane space. Protein translocases do not function as independent units but are integrated into dynamic networks and are connected to machineries that function in bioenergetics, mitochondrial morphology and coupling to the endoplasmic reticulum.
Role of essential genes in mitochondrial morphogenesis in Saccharomyces cerevisiae
- Mol. Biol. Cell
, 2006
"... Mitochondria are essential organelles of eukaryotic cells. Inheritance and maintenance of mitochondrial structure depend on cytoskeleton-mediated organelle transport and continuous membrane fusion and fission events. However, in Saccha-romyces cerevisiae most of the known components involved in thes ..."
Abstract
-
Cited by 24 (2 self)
- Add to MetaCart
Mitochondria are essential organelles of eukaryotic cells. Inheritance and maintenance of mitochondrial structure depend on cytoskeleton-mediated organelle transport and continuous membrane fusion and fission events. However, in Saccha-romyces cerevisiae most of the known components involved in these processes are encoded by genes that are not essential for viability. Here we asked which essential genes are required for mitochondrial distribution and morphology. To address this question, we performed a systematic screen of a yeast strain collection harboring essential genes under control of a regulatable promoter. This library contains 768 yeast mutants and covers approximately two thirds of all essential yeast genes. A total of 119 essential genes were found to be required for maintenance of mitochondrial morphology. Among these, genes were highly enriched that encode proteins involved in ergosterol biosynthesis, mitochondrial protein import, actin-dependent transport processes, vesicular trafficking, and ubiquitin/26S proteasome-dependent protein degradation. We conclude that these cellular pathways play an important role in mitochondrial morphogenesis and inheritance.
2007. Analysis of transcriptional activation at a distance in Saccharomyces cerevisiae
"... This article cites 78 articles, 45 of which can be accessed free ..."
Abstract
-
Cited by 21 (0 self)
- Add to MetaCart
(Show Context)
This article cites 78 articles, 45 of which can be accessed free
Esf2p, a U3-associated factor required for small-subunit processome assembly and compaction
- Mol. Cell. Biol
, 2005
"... Esf2p is the Saccharomyces cerevisiae homolog of mouse ABT1, a protein previously identified as a putative partner of the TATA-element binding protein. However, large-scale studies have indicated that Esf2p is primarily localized to the nucleolus and that it physically associates with pre-rRNA proc ..."
Abstract
-
Cited by 15 (7 self)
- Add to MetaCart
(Show Context)
Esf2p is the Saccharomyces cerevisiae homolog of mouse ABT1, a protein previously identified as a putative partner of the TATA-element binding protein. However, large-scale studies have indicated that Esf2p is primarily localized to the nucleolus and that it physically associates with pre-rRNA processing factors. Here, we show that Esf2p-depleted cells are defective for pre-rRNA processing at the early nucleolar cleavage sites A 0 through A 2 and consequently are inhibited for 18S rRNA synthesis. Esf2p was stably associated with the 5 external transcribed spacer (ETS) and the box C؉D snoRNA U3, as well as additional box C؉D snoRNAs and proteins enriched within the small-subunit (SSU) processome/90S preribosomes. Esf2p colocalized on glycerol gradients with 90S preribosomes and slower migrating particles containing 5 ETS fragments. Strikingly, upon Esf2p depletion, chromatin spreads revealed that SSU processome assembly and compaction are inhibited and glycerol gradient analysis showed that U3 remains associated within 90S preribosomes. This suggests that in the absence of proper SSU processome assembly, early pre-rRNA processing is inhibited and U3 is not properly released from the 35S pre-rRNAs. The identification of ABT1 in a large-scale analysis of the human nucleolar proteome indicates that its role may also be conserved in mammals. Ribosome biogenesis is a complicated pathway that requires in eukaryotes an excess of 200 protein trans-acting factors and just as many small nucleolar RNAs (snoRNAs) Most ribosome biogenesis factors are localized to the nucleolus, are essential for cell viability, and are components of large preribosomal protein complexes. Consequently, proteomic analyses have resulted in the identification of many potential new rRNA processing factors (RRPs) (reviewed in references 10 and 19), and the use of affinity purification has delineated distinct and presumably successive preribosomal complexes (reviewed in references 10, 12, and 46). Among these is the small-subunit (SSU) processome (3, 4, 9, 14, 35), a U3-containing ribonucleoprotein complex primarily involved in the three initial pre-rRNA processing reactions (cleavage at sites A 0 through A 2 ) (see As 35S pre-rRNA molecules emerge from the Pol I transcription machinery, 5Ј pre-rRNA termini are bound by U3 and early RRPs, generating the so-called "terminal knobs." These are visualized by electron microscopy on chromatin spreads at the ends of the rRNA gene "Christmas tree" branches
Non-redundant roles of mitochondria-associated Fbox proteins
- Mfb1 and Mdm30, in maintenance of mitochondrial morphology in
, 2006
"... Mitochondria constantly fuse and divide to adapt organellar morphology to the cell’s ever-changing physiological conditions. Little is known about the molecular mechanisms regulating mitochondrial dynamics. F-box proteins are subunits of both Skp1-Cullin-F-box (SCF) ubiquitin ligases and non-SCF com ..."
Abstract
-
Cited by 15 (1 self)
- Add to MetaCart
Mitochondria constantly fuse and divide to adapt organellar morphology to the cell’s ever-changing physiological conditions. Little is known about the molecular mechanisms regulating mitochondrial dynamics. F-box proteins are subunits of both Skp1-Cullin-F-box (SCF) ubiquitin ligases and non-SCF complexes that regulate a large number of cellular processes. Here, we analyzed the roles of two yeast F-box proteins, Mfb1 and Mdm30, in mitochondrial dynamics. Mfb1 is a novel mitochondria-associated F-box protein. Mitochondria in mutants lacking Mfb1 are fusion competent, but they form aberrant aggregates of interconnected tubules. In contrast, mitochondria in mutants lacking Mdm30 are highly fragmented due to a defect in mitochondrial fusion. Fragmented mitochondria are docked but nonfused in mdm30 cells. Mitochondrial fusion is also blocked during sporulation of homozygous diploid mutants lacking Mdm30, leading to a mitochondrial inheritance defect in ascospores. Mfb1 and Mdm30 exert nonredundant functions and likely have different target proteins. Because defects in F-box protein mutants could not be mimicked by depletion of SCF complex and proteasome core subunits, additional yet unknown factors are likely involved in regulating mitochondrial dynamics. We propose that mitochondria-associated F-box proteins Mfb1 and Mdm30 are key components of a complex machinery that regulates mitochondrial dynamics throughout yeast’s entire life cycle.
A survey of essential gene function in the yeast cell division cycle
- Mol. Biol. Cell
, 2006
"... Mutations impacting specific stages of cell growth and division have provided a foundation for dissecting mechanisms that underlie cell cycle progression. We have undertaken an objective examination of the yeast cell cycle through flow cytometric analysis of DNA content in TetO 7 promoter mutant str ..."
Abstract
-
Cited by 13 (1 self)
- Add to MetaCart
Mutations impacting specific stages of cell growth and division have provided a foundation for dissecting mechanisms that underlie cell cycle progression. We have undertaken an objective examination of the yeast cell cycle through flow cytometric analysis of DNA content in TetO 7 promoter mutant strains representing 75 % of all essential yeast genes. More than 65 % of the strains displayed specific alterations in DNA content, suggesting that reduced function of an essential gene in most cases impairs progression through a specific stage of the cell cycle. Because of the large number of essential genes required for protein biosynthesis, G1 accumulation was the most common phenotype observed in our analysis. In contrast, relatively few mutants displayed S-phase delay, and most of these were defective in genes required for DNA replication or nucleotide metabolism. G2 accumulation appeared to arise from a variety of defects. In addition to providing a global view of the diversity of essential cellular processes that influence cell cycle progression, these data also provided predictions regarding the functions of individual genes: we identified four new genes involved in protein trafficking (NUS1, PHS1, PGA2, PGA3), and we found that CSE1 and SMC4 are important for DNA replication.
Multiplicative mixture models for overlapping clustering
, 2008
"... The problem of overlapping clustering, where a point is allowed to belong to multiple clusters, is becoming increasingly important in a variety of applications. In this paper, we present an overlapping clustering algorithm based on multiplicative mixture models. We analyze a general setting where ea ..."
Abstract
-
Cited by 12 (3 self)
- Add to MetaCart
(Show Context)
The problem of overlapping clustering, where a point is allowed to belong to multiple clusters, is becoming increasingly important in a variety of applications. In this paper, we present an overlapping clustering algorithm based on multiplicative mixture models. We analyze a general setting where each component of the multiplicative mixture is from an exponential family, and present an efficient alternating maximization algorithm to learn the model and infer overlapping clusters. We also show that when each component is assumed to be a Gaussian, we can apply the kernel trick leading to non-linear cluster separators and obtain better clustering quality. The efficacy of the proposed algorithms is demonstrated using experiments on both UCI benchmark datasets and a microarray gene expression dataset. 1
