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COMATOSE (CTS), the Arabidopsis homologue of human Adrenoleukodystrophy protein (ALDP), is required for import of substrates for peroxisomal beta-oxidation. A new allelic series and a homology model based on the bacterial ABC transporter, Sav1866, provide novel insights into structure-function relations of ABC subfamily D proteins. In contrast to ALDP, where the majority of mutations result in protein absence from the peroxisomal membrane, all CTS mutants produced stable protein. Mutation of conserved residues in the Walker A and B motifs in CTS nucleotide-binding domain (NBD) 1 resulted in a null phenotype but had little effect in NBD2, indicating that the NBDs are functionally distinct in vivo. Two alleles containing mutations in NBD1 outside the Walker motifs (E617K and C631Y) exhibited resistance to auxin precursors 2,4-dichlorophenoxybutyric acid (2,4-DB) and indole butyric acid (IBA) but were wild type in all other tests. The homology model predicted that the transmission interfaces are domain-swapped in CTS, and the differential effects of mutations in the conserved "EAA motif" of coupling helix 2 supported this prediction, consistent with distinct roles for each NBD. Our findings demonstrate that CTS functions can be separated by mutagenesis and the structural model provides a framework for interpretation of phenotypic data.

Despite Arabidopsis thaliana's pre-eminence as a model organism, major questions remain regarding the geographic structure of its genetic variation due to the geographically incomplete sample set available for previous studies. Many of these questions are addressed here with an analysis of genome-wide variation at 10 loci in 475 individuals from 167 globally distributed populations, including many from critical but previously un-sampled regions. Rooted haplotype networks at three loci suggest that A. thaliana arose in the Caucasus region. Identification of large-scale metapopulations indicates clear east-west genetic structure, both within proposed Pleistocene refugia and post-Pleistocene colonized regions. The refugia themselves are genetically differentiated from one another and display elevated levels of within-population genetic diversity relative to recolonized areas. The timing of an inferred demographic expansion coincides with the Eemian interglacial (approximately 120,000 years ago). Taken together, these patterns are strongly suggestive of Pleistocene range dynamics. Spatial autocorrelation analyses indicate that isolation by distance is pervasive at all hierarchical levels, but that it is reduced in portions of Europe.

Population-based methods for the genetic mapping of adaptive traits and the analysis of natural selection require that the population structure and demographic history of a species are taken into account. We characterized geographic patterns of genetic variation in the model plant Arabidopsis thaliana by genotyping 115 genome-wide single nucleotide polymorphism (SNP) markers in 351 accessions from the whole species range using a matrix-assisted laser desorption/ionization time-of-flight assay, and by sequencing of nine unlinked short genomic regions in a subset of 64 accessions. The observed frequency distribution of SNPs is not consistent with a constant-size neutral model of sequence polymorphism due to an excess of rare polymorphisms. There is evidence for a significant population structure as indicated by differences in genetic diversity between geographic regions. Accessions from Central Asia have a low level of polymorphism and an increased level of genome-wide linkage disequilibrium (LD) relative to accessions from the Iberian Peninsula and Central Europe. Cluster analysis with the structure program grouped Eurasian accessions into K = 6 clusters. Accessions from the Iberian Peninsula and from Central Asia constitute distinct populations, whereas Central and Eastern European accessions represent admixed populations in which genomes were reshuffled by historical recombination events. These patterns likely result from a rapid postglacial recolonization of Eurasia from glacial refugial populations. Our analyses suggest that mapping populations for association or LD mapping should be chosen from regional rather than a species-wide sample or identified genetically as sets of individuals with similar average genetic distances.

Background and Aims: Germination and establishment of seeds are complex traits affected by a wide range of internal and external influences. The effects of parental temperature preconditioning and temperature during germination on germination and establishment of Arabidopsis thaliana were examined., Methods: Seeds from parental plants grown at 14 and at 22 degrees C were screened for germination (protrusion of radicle) and establishment (greening of cotyledons) at three different temperatures (10, 18 and 26 degrees C). Seventy-three accessions from across the entire distribution range of A. thaliana were included., Key Results: Multifactorial analyses of variances revealed significant differences in the effects of genotypes, preconditioning, temperature treatment, and their interactions on duration of germination and establishment. Reaction norms showed an enormous range of plasticity among the preconditioning and different germination temperatures. Correlations of percentage total germination and establishment after 38 d with the geographical origin of accessions were only significant for 14 degrees C preconditioning but not for 22 degrees C preconditioning. Correlations with temperature and precipitation on the origin of the accessions were mainly found at the lower germination temperatures (10 and 18 degrees C) and were absent at higher germination temperatures (26 degrees C)., Conclusions: Overall, the data show huge variation of germination and establishment among natural accessions of A. thaliana and might serve as a valuable source for further germination and plasticity studies.

Comparative physiological analysis of mutant Arabidopsis seeds under defined environmental conditions was used to analyse the relative contributions of components of peroxisomal beta-oxidation in the control of seed germination potential. The COMATOSE (CTS) and KAT2 loci were shown to play essential roles in regulating germination and establishment potentials, whereas LACS6 and LACS7 loci only influenced establishment following germination. The viability and desiccation tolerance of three different mutant alleles of CTS were shown to be intermediate between that of dormant and non-dormant wild-type seeds. Analysis of ttg-1 cts-1 double mutant seeds demonstrated that the cts lesion did not influence after-ripening capacity. These data demonstrate the importance of peroxisomal beta-oxidation in the control of germination potential, but suggest that breakdown of stored lipid is not an important prerequisite for germination. A function is suggested for CTS following after-ripening within pathways related to the progression of germination prior to radicle emergence.

Background and Aims: Estimates of the amount of nuclear DNA of Arabidopsis thaliana, known to be among the lowest within angiosperms, vary considerably. This study aimed to determine genome size of a range of accessions from throughout the entire Eurasian range of the species., Methods: Twenty accessions from all over Europe and one from Japan were examined using flow cytometry., Key Results: Significant differences in mean C-values were detected over a 1.1-fold range. Mean haploid (1C) genome size was 0.215 pg (211 Mbp) for all analysed accessions. Two accessions were tetraploid., Conclusions: A closer investigation of the DNA fractions involved in intraspecific genome size differences in this experimentally accessible species may provide information on the factors involved in stability and evolution of genome sizes.

Range expansion from Pleistocene refugia and anthropogenic influences contribute to the present distribution pattern of Arabidopsis thaliana. We scored a genome-wide set of CAPSs and found two markers with an east-west geographic distribution across the Eurasian range of the species. Regions around the two SNPs were sequenced in 98 accessions, including newly collected plants from Middle Asia and Western Siberia. These regions correspond to a gene ( approximately 1500 bp) and a non-coding region ( approximately 500 bp) 300 kbp apart on chromosome 2. Nucleotide diversities, pi, of the two sequenced fragments were 0.0032 and 0.0130. The haplotypes of both sequences belonged to one of two groups: a rather uniform "Asian" and a more variable "European" haplotype group, on the basis of non-disjunct clusters of SNPs. Recombination between "Asian" and "European" haplotypes occurs where they meet. Especially in the "European" haplotype, many rare SNP variants representing independent mutations are scattered among the shared haplotype-specific SNPs. This agrees with previous suggestions of two large haplotype groups in A. thaliana and the post-glacial colonization of central Europe from the east and the west. A clear correlation between climatic factors and the haplotype distribution may reflect the dispersal history rather than local climate adaptation. The pattern of SNP variation within the contiguous sequences explains why only a minority of SNPs selected across the genome show evidence of this geographic pattern.

A Geographical Information System (GIS) is used to analyse allelic information of 13 sequenced loci of natural populations of Arabidopsis thaliana and to identify geographical structures. GIS provides tools for visualization and analysis of geographical population structures using molecular data. The geographical distribution of the number of variable positions in the alignments, the distribution of recombinant sequence blocks, and the distribution of a newly defined measure, the differentiation index, are studied. The differentiation index is introduced to measure the sequence divergence among individual plants sampled from various geographical localities. The numbers of variable positions and the differentiation index are also used for a metadata analysis covering about 26 kb of the genome. This analysis reveals, for the first time, differences in DNA sequence structures of geographically different populations of A. thaliana. The broadly defined west Mediterranean region consists of accessions with the highest numbers of polymorphic positions followed by the west European region. The GIS technology Kriging is used to define Arabidopsis specific diversity zones in Europe. The highest genetic variability is observed along the Atlantic coast from the western Iberian Peninsula to southern Great Britain, while lowest variability is found in central Europe.