Predicting phenotypic variation in yeast from individual genome sequences

A central challenge in genetics is to predict phenotypic variation from individual genome sequences. Here we construct and evaluate phenotypic predictions for 19 strains of Saccharomyces cerevisiae. We use conservation-based methods to predict the impact of protein-coding variation within genes on protein function. We then rank strains using a prediction score that measures the total sum of function-altering changes in different sets of genes reported to influence over 100 phenotypes in genome-wide loss-of-function screens. We evaluate our predictions by comparing them with the observed growth rate and efficiency of 15 strains tested across 20 conditions in quantitative experiments. The median predictive performance, as measured by ROC AUC, was 0.76, and predictions were more accurate when the genes reported to influence a trait were highly connected in a functional gene network.     

Birth of a metabolic gene cluster in yeast by adaptive gene relocation

Although most eukaryotic genomes lack operons, they contain some physical clusters of genes that are related in function despite being unrelated in sequence. How these clusters are formed during evolution is unknown. The DAL cluster is the largest metabolic gene cluster in yeast and consists of six adjacent genes encoding proteins that enable Saccharomyces cerevisiae to use allantoin as a nitrogen source. We show here that the DAL cluster was assembled, quite recently in evolutionary terms, through a set of genomic rearrangements that happened almost simultaneously. Six of the eight genes involved in allantoin degradation, which were previously scattered around the genome, became relocated to a single subtelomeric site in an ancestor of S. cerevisiae and Saccharomyces castellii. These genomic rearrangements coincided with a biochemical reorganization of the purine degradation pathway, which switched to importing allantoin instead of urate. This change eliminated urate oxidase, one of several oxygen-consuming enzymes that were lost by yeasts that can grow vigorously in anaerobic conditions. The DAL cluster is located in a domain of modified chromatin involving both H2A.Z histone exchange and Hst1-Sum1-mediated histone deacetylation, and it may be a coadapted gene complex formed by epistatic selection.     

Population genomics of human gene expression

Genetic variation influences gene expression, and this variation in gene expression can be efficiently mapped to specific genomic regions and variants. Here we have used gene expression profiling of Epstein-Barr virus-transformed lymphoblastoid cell lines of all 270 individuals genotyped in the HapMap Consortium to elucidate the detailed features of genetic variation underlying gene expression variation. We find that gene expression is heritable and that differentiation between populations is in agreement with earlier small-scale studies. A detailed association analysis of over 2.2 million common SNPs per population (5% frequency in HapMap) with gene expression identified at least 1,348 genes with association signals in cis and at least 180 in trans. Replication in at least one independent population was achieved for 37% of cis signals and 15% of trans signals, respectively. Our results strongly support an abundance of cis-regulatory variation in the human genome. Detection of trans effects is limited but suggests that regulatory variation may be the key primary effect contributing to phenotypic variation in humans. We also explore several methodologies that improve the current state of analysis of gene expression variation.     

MicroRNAs can generate thresholds in target gene expression

MicroRNAs (miRNAs) are short, highly conserved noncoding RNA molecules that repress gene expression in a sequence-dependent manner. We performed single-cell measurements using quantitative fluorescence microscopy and flow cytometry to monitor a target gene's protein expression in the presence and absence of regulation by miRNA. We find that although the average level of repression is modest, in agreement with previous population-based measurements, the repression among individual cells varies dramatically. In particular, we show that regulation by miRNAs establishes a threshold level of target mRNA below which protein production is highly repressed. Near this threshold, protein expression responds sensitively to target mRNA input, consistent with a mathematical model of molecular titration. These results show that miRNAs can act both as a switch and as a fine-tuner of gene expression.     

Computer-based three-dimensional visualization of developmental gene expression

A broad understanding of the relationship between gene activation, pattern formation and morphogenesis will require adequate tools for three-dimensional and, perhaps four-dimensional, representation and analysis of molecular developmental processes. We present a novel, computer-based method for the 3D visualization of embryonic gene expression and morphological structures from serial sections. The information from these automatically aligned 3D reconstructions exceeds that from single-section and whole-mount visualizations of in situ hybridizations. In addition, these 3D models of gene-expression patterns can become a central component of a future developmental database designed for the collection and presentation of digitized, morphological and gene-expression data. This work is accompanied by a web site (http://www.univie.ac.at/GeneEMAC).     

Identification of foreign gene sequences by transcript filtering against the human genome.

We have developed a computational subtraction approach to detect microbial causes for putative infectious diseases by filtering a set of human tissue-derived sequences against the human genome. We demonstrate the potential of this method by identifying sequences from known pathogens in established expressed-sequence tag libraries.     

Action of BTN1, the yeast orthologue of the gene mutated in Batten disease.

Neuronal ceroid-lipofuscinoses (NCL) are autosomal recessive disorders that form the most common group of progressive neurodegenerative diseases in children, with an incidence as high as 1 in 12,500 live births, and with approximately 440,000 carriers in the United States. Disease progression is characterized by a decline in mental abilities, increased severity of untreatable seizures, blindness, loss of motor skills and premature death. The CLN3 gene, which is responsible for Batten disease, has been positionally cloned. The yeast gene, denoted BTN1, encodes a non-essential protein that is 39% identical and 59% similar to human CLN3. Strains lacking Btn1p, btn1-delta, are resistant to D-(-)-threo-2-amino-1-[p-nitrophenyl]-1,3-propanediol (ANP) in a pH-dependent manner. This phenotype was complemented by expression of human CLN3, demonstrating that yeast Btn1p and human CLN3 share the same function. Here, we report that btn1-delta yeast strains have an abnormally acidic vacuolar pH in the early phases of growth. Furthermore, DNA microarray analysis of BTN1 and btn1-delta strains revealed differential expression of two genes, with at least one, HSP30, involved in pH control. Because Btn1p is located in the vacuole, we suggest that Batten disease is caused by a defect in vacuolar (lysosomal) pH control. Our findings draw parallels between fundamental biological processes in yeast and previously observed characteristics of neurodegeneration in humans.