ABSTRACT
PDE4 inhibitors have the potential to alleviate the symptoms and underlying inflammation associated with dry eye. Disclosed herein is the development of a novel series of water-soluble PDE4 inhibitors. Our studies led to the discovery of coumarin 18, which is effective in a rabbit model of dry eye and a tear secretion test in rats.
Subject(s)
4-Aminopyridine/analogs & derivatives , Anti-Inflammatory Agents/chemistry , Coumarins/chemistry , Phosphodiesterase 4 Inhibitors , Water/chemistry , 4-Aminopyridine/chemical synthesis , 4-Aminopyridine/chemistry , 4-Aminopyridine/therapeutic use , Animals , Anti-Inflammatory Agents/chemical synthesis , Anti-Inflammatory Agents/therapeutic use , Binding Sites , Computer Simulation , Coumarins/chemical synthesis , Coumarins/therapeutic use , Cyclic Nucleotide Phosphodiesterases, Type 4/metabolism , Disease Models, Animal , Dry Eye Syndromes/drug therapy , Rabbits , RatsABSTRACT
The mRNA and protein expression in Saccharomyces cerevisiae cultured in rich or minimal media was analyzed by oligonucleotide arrays and quantitative multidimensional protein identification technology. The overall correlation between mRNA and protein expression was weakly positive with a Spearman rank correlation coefficient of 0.45 for 678 loci. To place the data sets in a proper biological context, a clustering approach based on protein pathways and protein complexes was implemented. Protein expression levels were transcriptionally controlled for not only single loci but for entire protein pathways (e.g., Met, Arg, and Leu biosynthetic pathways). In contrast, the protein expression of loci in several protein complexes (e.g., SPT, COPI, and ribosome) was posttranscriptionally controlled. The coupling of the methods described provided insight into the biology of S. cerevisiae and a clustering strategy by which future studies should be based.
Subject(s)
RNA, Fungal/genetics , RNA, Fungal/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Saccharomyces cerevisiae Proteins/genetics , Saccharomyces cerevisiae Proteins/metabolism , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolism , Amino Acid Sequence , Cluster Analysis , Data Interpretation, Statistical , Gene Expression Profiling/statistics & numerical data , Genes, Fungal , Molecular Sequence Data , Oligonucleotide Array Sequence Analysis/statistics & numerical data , Peptides/genetics , Peptides/metabolism , Protein Array Analysis/statistics & numerical data , Saccharomyces cerevisiae/growth & developmentABSTRACT
The availability of a complete genome sequence allows the detailed study of intraspecies variability. Here we use high-density oligonucleotide arrays to discover 11,115 single-feature polymorphisms (SFPs) existing in one or more of 14 different yeast strains. We use these SFPs to define regions of genetic identity between common laboratory strains of yeast. We assess the genome-wide distribution of genetic variation on the basis of this yeast population. We find that genome variability is biased toward the ends of chromosomes and is more likely to be found in genes with roles in fermentation or in transport. This subtelomeric bias may arise through recombination between nonhomologous sequences because full-gene deletions are more common in these regions than in more central regions of the chromosome.
Subject(s)
Genetic Variation , Saccharomyces cerevisiae/genetics , Chromosome Mapping , Oligonucleotide Array Sequence Analysis , Phylogeny , Polymorphism, Genetic , TelomereABSTRACT
A competitive growth assay has been used to identify yeast genes involved in the repair of UV- or MMS-induced DNA damage. A collection of 2,827 yeast strains was analyzed in which each strain has a single ORF replaced with a cassette containing two unique sequence tags, allowing for its detection by hybridization to a high-density oligonucleotide array. The hybridization data identify a high percentage of the deletion strains present in the collection that were previously characterized as being sensitive to the DNA-damaging agents. The assay, and subsequent analysis, has been used to identify six genes not formerly known to be involved in the damage response, whose deletion renders the yeast sensitive to UV or MMS treatment. The recently identified genes include three uncharacterized ORFs, as well as genes that encode protein products implicated in ubiquitination, gene silencing, and transport across the mitochondrial membrane. Epistatsis analysis of four of the genes was performed to determine the DNA damage repair pathways in which the protein products function.
Subject(s)
DNA Damage , DNA, Fungal/drug effects , DNA, Fungal/radiation effects , Epistasis, Genetic , Methyl Methanesulfonate/pharmacology , Mutagens/pharmacology , Saccharomyces cerevisiae/drug effects , Saccharomyces cerevisiae/genetics , Ultraviolet RaysABSTRACT
Short open reading frames (ORFs) occur frequently in primary genome sequence. Distinguishing bona fide small genes from the tens of thousands of short ORFs is one of the most challenging aspects of genome annotation. Direct experimental evidence is often required. Here we use a combination of expression profiling and mass spectrometry to verify the independent transcription of 138 and the translation of 50 previously nonannotated genes in the Saccharomyces cerevisiae genome. Through combined evidence, we propose the addition of 62 new genes to the genome and provide experimental support for the inclusion of 10 previously identified genes.
