The DNA damage response pathway contributes to the stability of chromosome III derivatives lacking efficient replicators.

In eukaryotic chromosomes, DNA replication initiates at multiple origins. Large inter-origin gaps arise when several adjacent origins fail to fire. Little is known about how cells cope with this situation.

The genetic landscape of a cell.

A genome-scale genetic interaction map was constructed by examining 5.4 million gene-gene pairs for synthetic genetic interactions, generating quantitative genetic interaction profiles for approximately 75% of all genes in the budding yeast, Saccharomyces cerevisiae. A network based on genetic interaction profiles reveals a functional map of the cell in which genes of similar biological processes cluster together in coherent subsets, and highly correlated profiles delineate specific pathways to define gene function.

Ubiquitin-related modifier Urm1 acts as a sulphur carrier in thiolation of eukaryotic transfer RNA.

Ubiquitin-like proteins (UBLs) can change protein function, localization or turnover by covalent attachment to lysine residues. Although UBLs achieve this conjugation through an intricate enzymatic cascade, their bacterial counterparts MoaD and ThiS function as sulphur carrier proteins. Here we show that Urm1p, the most ancient UBL, acts as a sulphur carrier in the process of eukaryotic transfer RNA (tRNA) modification, providing a possible evolutionary link between UBL and sulphur transfer.

Rex1p deficiency leads to accumulation of precursor initiator tRNAMet and polyadenylation of substrate RNAs in Saccharomyces cerevisiae.

A synthetic genetic array was used to identify lethal and slow-growth phenotypes produced when a mutation in TRM6, which encodes a tRNA modification enzyme subunit, was combined with the deletion of any non-essential gene in Saccharomyces cerevisiae. We found that deletion of the REX1 gene resulted in a slow-growth phenotype in the trm6-504 strain. Previously, REX1 was shown to be involved in processing the 3' ends of 5S rRNA and the dimeric tRNA(Arg)-tRNA(Asp).

Genetic and biochemical analysis of yeast and human cap trimethylguanosine synthase: functional overlap of 2,2,7-trimethylguanosine caps, small nuclear ribonucleoprotein components, pre-mRNA splicing factors, and RNA decay pathways.

Trimethylguanosine synthase (Tgs1) is the enzyme that converts standard m(7)G caps to the 2,2,7-trimethylguanosine (TMG) caps characteristic of spliceosomal small nuclear RNAs. Fungi and mammalian somatic cells are able to grow in the absence of Tgs1 and TMG caps, suggesting that an essential function of the TMG cap might be obscured by functional redundancy. A systematic screen in budding yeast identified nonessential genes that, when deleted, caused synthetic growth defects with tgs1Delta.

Trans-Golgi network and endosome dynamics connect ceramide homeostasis with regulation of the unfolded protein response and TOR signaling in yeast.

Synthetic genetic array analyses identify powerful genetic interactions between a thermosensitive allele (sec14-1(ts)) of the structural gene for the major yeast phosphatidylinositol transfer protein (SEC14) and a structural gene deletion allele (tlg2Delta) for the Tlg2 target membrane-soluble N-ethylmaleimide-sensitive factor attachment protein receptor. The data further demonstrate Sec14 is required for proper trans-Golgi network (TGN)/endosomal dynamics in yeast. Paradoxically, combinatorial depletion of Sec14 and Tlg2 activities elicits trafficking defects from the endoplasmic reticulum, and these defects are accompanied by compromise of the unfolded protein response (UPR).

Genetic interactions of MAF1 identify a role for Med20 in transcriptional repression of ribosomal protein genes.

Transcriptional repression of ribosomal components and tRNAs is coordinately regulated in response to a wide variety of environmental stresses. Part of this response involves the convergence of different nutritional and stress signaling pathways on Maf1, a protein that is essential for repressing transcription by RNA polymerase (pol) III in Saccharomyces cerevisiae. Here we identify the functions buffering yeast cells that are unable to down-regulate transcription by RNA pol III.

The interaction network of the chaperonin CCT.

The eukaryotic cytosolic chaperonin containing TCP-1 (CCT) has an important function in maintaining cellular homoeostasis by assisting the folding of many proteins, including the cytoskeletal components actin and tubulin. Yet the nature of the proteins and cellular pathways dependent on CCT function has not been established globally. Here, we use proteomic and genomic approaches to define CCT interaction networks involving 136 proteins/genes that include links to the nuclear pore complex, chromatin remodelling, and protein degradation.

Molecular chaperone Hsp90 stabilizes Pih1/Nop17 to maintain R2TP complex activity that regulates snoRNA accumulation.

Hsp90 is a highly conserved molecular chaperone that is involved in modulating a multitude of cellular processes. In this study, we identify a function for the chaperone in RNA processing and maintenance. This functionality of Hsp90 involves two recently identified interactors of the chaperone: Tah1 and Pih1/Nop17.

The Shwachman-Bodian-Diamond syndrome protein mediates translational activation of ribosomes in yeast.

The autosomal recessive disorder Shwachman-Diamond syndrome, characterized by bone marrow failure and leukemia predisposition, is caused by deficiency of the highly conserved Shwachman-Bodian-Diamond syndrome (SBDS) protein. Here, we identify the function of the yeast SBDS ortholog Sdo1, showing that it is critical for the release and recycling of the nucleolar shuttling factor Tif6 from pre-60S ribosomes, a key step in 60S maturation and translational activation of ribosomes. Using genome-wide synthetic genetic array mapping, we identified multiple TIF6 gain-of-function alleles that suppressed the pre-60S nuclear export defects and cytoplasmic mislocalization of Tif6 observed in sdo1Delta cells.

Exploring the mode-of-action of bioactive compounds by chemical-genetic profiling in yeast.

Discovering target and off-target effects of specific compounds is critical to drug discovery and development. We generated a compendium of "chemical-genetic interaction" profiles by testing the collection of viable yeast haploid deletion mutants for hypersensitivity to 82 compounds and natural product extracts. To cluster compounds with a similar mode-of-action and to reveal insights into the cellular pathways and proteins affected, we applied both a hierarchical clustering and a factorgram method, which allows a gene or compound to be associated with more than one group.

Identification of a bacterial type III effector family with G protein mimicry functions.

Many bacterial pathogens use the type III secretion system to inject "effector" proteins into host cells. Here, we report the identification of a 24 member effector protein family found in pathogens including Salmonella, Shigella, and enteropathogenic E. coli.

The lipopeptide antibiotic A54145 biosynthetic gene cluster from Streptomyces fradiae.

Ca(2+)-dependent cyclic lipodepsipeptides are an emerging class of antibiotics for the treatment of infections caused by Gram-positive pathogens. These compounds are synthesized by nonribosomal peptide synthetase (NRPS) complexes encoded by large gene clusters. The gene cluster encoding biosynthetic pathway enzymes for the Streptomyces fradiae A54145 NRP was cloned from a cosmid library and characterized.

The synthetic genetic interaction spectrum of essential genes.

The nature of synthetic genetic interactions involving essential genes (those required for viability) has not been previously examined in a broad and unbiased manner. We crossed yeast strains carrying promoter-replacement alleles for more than half of all essential yeast genes to a panel of 30 different mutants with defects in diverse cellular processes. The resulting genetic network is biased toward interactions between functionally related genes, enabling identification of a previously uncharacterized essential gene (PGA1) required for specific functions of the endoplasmic reticulum.

Daptomycin biosynthesis in Streptomyces roseosporus: cloning and analysis of the gene cluster and revision of peptide stereochemistry.

Daptomycin is a 13 amino acid, cyclic lipopeptide produced by a non-ribosomal peptide synthetase (NRPS) mechanism in Streptomyces roseosporus. A 128 kb region of S. roseosporus DNA was cloned and verified by heterologous expression in Streptomyces lividans to contain the daptomycin biosynthetic gene cluster (dpt).

Global mapping of the yeast genetic interaction network.

A genetic interaction network containing approximately 1000 genes and approximately 4000 interactions was mapped by crossing mutations in 132 different query genes into a set of approximately 4700 viable gene yeast deletion mutants and scoring the double mutant progeny for fitness defects. Network connectivity was predictive of function because interactions often occurred among functionally related genes, and similar patterns of interactions tended to identify components of the same pathway. The genetic network exhibited dense local neighborhoods; therefore, the position of a gene on a partially mapped network is predictive of other genetic interactions.

Integration of chemical-genetic and genetic interaction data links bioactive compounds to cellular target pathways.

Bioactive compounds can be valuable research tools and drug leads, but it is often difficult to identify their mechanism of action or cellular target. Here we investigate the potential for integration of chemical-genetic and genetic interaction data to reveal information about the pathways and targets of inhibitory compounds. Taking advantage of the existing complete set of yeast haploid deletion mutants, we generated drug-hypersensitivity (chemical-genetic) profiles for 12 compounds.