Extensive cross-environment fitness variation lies along few axes of genetic variation in the model alga, Chlamydomonas reinhardtii.

Variation is essential to ecological and evolutionary dynamics, but genetic variation of quantitative traits may be concentrated in a limited number of dimensions, constraining ecoevolutionary dynamics. We describe high-dimension variation in natural accessions of the model alga, Chlamydomonas reinhardtii, and test the hypothesis that extensive fitness variation across 30 environments is constrained to a small number of axes. We used high-throughput phenotyping to investigate morphological, fitness, and genotype × environment (G × E) variation in 18 natural C. reinhardtii accessions in 30 environments. The organismal phenotypes of cell cycle, cell size, and phototactic behavior exhibited substantial genetic variation between lines, and we found up to 74-fold fitness variation across accessions and environments. Approximately 47% of the extensive G × E variation is accounted for by the first two principal components (PCs) of the G-matrix corresponding to covariation in metals response, nitrogen availability, or salt and nutrient response. The natural variation of C. reinhardtii accessions supports the hypothesis that, despite abundant genetic variation across single environments, the species' adaptive response should be constrained along few major axes of selection. These results highlight the utility of natural accessions for integrating ecoevolutionary and genetic research.

Physiological genomics of response to soil drying in diverse Arabidopsis accessions.

Arabidopsis thaliana, like many species, is characterized by abundant genetic variation. This variation is rapidly being cataloged at the sequence level, but careful dissection of genetic variation in whole-organism responses to stresses encountered in the natural environment are lacking; this functional variation can be exploited as a natural mutant screen to determine gene function. Here, we document physiological and transcriptomic response to soil drying in 17 natural accessions of Arabidopsis. By imposing ecologically realistic stress conditions, we found that acclimation in Arabidopsis involved a strong signature of increased investment in photosynthesis, carbohydrate turnover, and root growth. Our results extend previous work in the Columbia accession suggesting that abscisic acid signaling pathways play an important role in drought stress response. We also identified several mechanisms, including an increase in leaf nitrogen concentration and upregulation of two-component signaling relays, that were common to most natural accessions but had not been identified in studies using only the Columbia accession. Principal component analysis reveals strong correlations between suites of genes and specific physiological responses to stress. The functional variants we identified may represent adaptive mutations in natural habitats and useful variants for agronomic improvement of crop species.

Exploring genetic and expression differences between physiologically extreme ecotypes: comparative genomic hybridization and gene expression studies of Kas-1 and Tsu-1 accessions of Arabidopsis thaliana.

Recent studies have documented remarkable genetic variation among Arabidopsis thaliana accessions collected from diverse habitats. Of particular interest are accessions with putatively locally adapted phenotypes - that is, accessions with attributes that are likely adaptive at their sites of origin. These genotypes may provide insight into the genetic basis of adaptive evolution as well as allow the discovery of genes of ecological importance. We studied the physiology, genome content and gene expression of two physiologically extreme accessions (Tsu-1 from Tsushima, Japan and Kas-1 from Kashmir, India). Our study was conducted under two levels of soil moisture and accompanied by physiological measurements to characterize early responses to soil drying. Genomic hybridizations identified 42,503 single feature polymorphisms (SFP) between accessions, providing an initial screen for genetic differences. Transcript profiling identified a large number (5996) of genes exhibiting constitutive differences in expression including genes involved in many biological pathways. Mild soil drying resulted in only subtle physiological responses but resulted in gene expression changes in hundreds of transcripts, including 352 genes exhibiting differential responses between accessions. Our results highlight the value of genomic studies of natural accessions as well as identify a number of candidate genes underlying physiological differences between Tsu-1 and Kas-1.

Contributions of ATM mutations to familial breast and ovarian cancer.

This study addresses the prevalence of ATM mutations and the association with breast cancer in Austrian families selected for a history of breast or ovarian cancer or both [hereditary breast and ovarian cancer (HBOC)]. In 270 HBOC families previously screened for BRCA1 and BRCA2 mutations, 137 different sequence alterations of ATM were identified. Seven of these were mutations presumed to cause ataxia telangiectasia based on their effect on the ATM protein, including five that caused a protein truncation and two missense mutations in the catalytic kinase domain of the highly conserved COOH terminus of the protein. The seven mutations were found in 10 families (3.7%). In addition, one missense variant, L1420F, was observed in 13 HBOC families (4.8%) but was not observed in any of the 122 healthy volunteers with no history of breast cancer. In addition, the variant segregated with breast cancer in some of the families, suggesting that it may be pathogenic for breast cancer. Sixty-two additional variants of potential significance were observed in 65 HBOC families, but not in healthy controls. These variants included 24 sequence alterations with possible effects on splicing or protein-protein interactions. This study indicates that there is a significant prevalence of ATM mutations in breast and ovarian cancer families and adds to a growing body of evidence that ATM mutations confer increased susceptibility to breast cancer.

Two ATM variants and breast cancer risk.

The ATM gene is mutated in ataxia-telangiectasia (AT). Heterozygote female relatives of AT cases have a 2-7fold increased risk of breast cancer. We previously reported high risks of breast cancer associated with certain ATM variants. To estimate the risks more precisely, we have examined two ATM variants, c.1066-6T>G (IVS10-6T>G) and c.4258C>T (p.Leu1420Phe), in additional cases and controls from the same Australian cohorts previously used to estimate the risk of breast cancer associated with c.1066-6T>G. A total of 775 and 84 population-based controls were genotyped for the c.1066-6T>G and c.4258C>T ATM variants respectively, as were index cases from 378 and 373 non-BRCA1/2 breast cancer families. Penetrance was estimated by Bayes factor analysis. The allele frequencies of ATM c.1066-6T>G and c.4258C>T estimated from controls were 0.005 (95% CI=0.002 to 0.009) and 0.012 (95% CI=0.001 to 0.042), respectively. We identified three new breast cancer families with c.1066-6T>G, and seven families with c.4258C>T. Combining with the two c.1066-6T>G families previously reported, the estimated penetrance to age 70 of c.1066-6T>G was 17.2% (95% CI=4.7% to 37.5%). For c.4258C>T, the estimated average penetrance was 4.8% (95% CI 1.7% to 10.1%). In conclusion, we found no evidence that the ATM c.4258C>T variant increases breast cancer risk, and little evidence that c.1066-6T>G confers an elevated risk. Analysis of additional families will be necessary to define more precisely the risk, if any, associated with c.1066-6T>G.

Mutation of the ATM gene is not involved in the pathogenesis of either follicle center lymphoma or its transformation to higher-grade lymphoma.

Loss of function of the ataxia-telangiectasia mutated (ATM) gene, located on human chromosome 11q22-23, is the cause of ataxia-telangiectasia (A-T), which is associated with an extremely high risk for lymphoma. Abnormalities in 11q22-23, including deletions and mutations of the ATM gene, have been reported in T-cell prolymphocytic leukemias, B-CLL and in mantle cell lymphoma. In a survey of gene expression in follicle center lymphomas (FCL) and diffuse large B-cell lymphomas (DLBCL), almost all FCL expressed significant levels of ATM and the majority of DLBCL expressed low levels of ATM. This finding raised the possibility that the transformation of some FCL to DLBCL might be associated with inactivation of the ATM gene. Therefore, we analyzed biopsy specimens of 17 patients with FCL obtained at the time of diagnosis, four subsequent biopsies obtained at the time of FCL relapse and seven subsequent biopsies at the time of transformation to DLBCL. A comprehensive analysis of the ATM gene was performed by denaturing high performance liquid chromatography and sequencing. The analysis covered all of the 66 exons including the 9168 base pairs of ATM coding sequence as well as 16,676 base pairs of non-coding sequence. Twenty-eight known polymorphisms and rare sequence variants were observed, but no classic A-T mutations were detected. In 11 tumors, both tumor B-cells and normal T-cells were sorted for separate examination, and in each case, polymorphisms and rare variants were present in both tumor and normal cells. No new ATM gene mutations were associated with transformation from FCL to DLBCL. Thus, ATM gene mutations do not play a pivotal role either in the pathogenesis of FCL or in its transformation to DLBCL.

Genetics of drought adaptation in Arabidopsis thaliana II. QTL analysis of a new mapping population, KAS-1 x TSU-1.

Despite compelling evidence that adaptation to local climate is common in plant populations, little is known about the evolutionary genetics of traits that contribute to climatic adaptation. A screen of natural accessions of Arabidopsis thaliana revealed Tsu-1 and Kas-1 to be opposite extremes for water-use efficiency and climate at collection sites for these accessions differs greatly. To provide a tool to understand the genetic basis of this putative adaptation, Kas-1 and Tsu-1 were reciprocally crossed to create a new mapping population. Analysis of F(3) families showed segregating variation in both delta(13)C and transpiration rate, and as expected these traits had a negative genetic correlation (r(g)=- 0.3). 346 RILs, 148 with Kas-1 cytoplasm and 198 with Tsu-1 cytoplasm, were advanced to the F(9) and genotyped using 48 microsatellites and 55 SNPs for a total of 103 markers. This mapping population was used for QTL analysis of delta(13)C using F(9) RIL means. Analysis of this reciprocal cross showed a large effect of cytoplasmic background, as well as two QTL for delta(13)C. The Kas-1 x Tsu-1 mapping population provides a powerful new resource for mapping QTL underlying natural variation and for dissecting the genetic basis of water-use efficiency differences.

Natural genetic variation in whole-genome expression in Arabidopsis thaliana: the impact of physiological QTL introgression.

A long-standing and fundamental question in biology is how genes influence complex phenotypes. Combining near-isogenic line mapping with genome expression profiling offers a unique opportunity for exploring the functional relationship between genotype and phenotype and for generating candidate genes for future study. We used a whole-genome microarray produced with ink-jet technology to measure the relative expression level of over 21,500 genes from an Arabidopsis thaliana near-isogenic line (NIL) and its recurrent parent. The NIL material contained two introgressions (bottom of chromosome II and top of chromosome III) of the Cvi-1 ecotype in a Ler-2 ecotype genome background. Each introgression 'captures' a Cvi allele of a physiological quantitative trait loci (QTL) that our previous studies have shown increases transpiration and reduces water-use efficiency at the whole-plant level. We used a mixed model anova framework for assessing sources of expression variability and for evaluating statistical significance in our array experiment. We discovered 25 differentially expressed genes in the introgression at a false-discovery rate (FDR) cut-off of 0.20 and identified new candidate genes for both QTL regions. Several differentially expressed genes were confirmed with QRT-PCR (quantitative reverse transcription-polymerase chain reaction) assays. In contrast, we found no statistically significant differentially expressed genes outside of the QTL introgressions after controlling for multiple tests. We discuss these results in the context of candidate genes, cloning QTL, and phenotypic evolution.