Adaptive significance of flowering time variation across natural seasonal environments in Arabidopsis thaliana.

The relevance of flowering time variation and plasticity to climate adaptation requires a comprehensive empirical assessment. We investigated natural selection and the genetic architecture of flowering time in Arabidopsis through field experiments in Europe across multiple sites and seasons. We estimated selection for flowering time, plasticity and canalization. Loci associated with flowering time, plasticity and canalization by genome-wide association studies were tested for a geographic signature of climate adaptation. Selection favored early flowering and increased canalization, except at the northernmost site, but was rarely detected for plasticity. Genome-wide association studies revealed significant associations with flowering traits and supported a substantial polygenic inheritance. Alleles associated with late flowering, including functional FRIGIDA variants, were more common in regions experiencing high annual temperature variation. Flowering time plasticity to fall vs spring and summer environments was associated with GIGANTEA SUPPRESSOR 5, which promotes early flowering under decreasing day length and temperature. The finding that late flowering genotypes and alleles are associated with climate is evidence for past adaptation. Real-time phenotypic selection analysis, however, reveals pervasive contemporary selection for rapid flowering in agricultural settings across most of the species range. The response to this selection may involve genetic shifts in environmental cuing compared to the ancestral state.

Large-effect flowering time mutations reveal conditionally adaptive paths through fitness landscapes in Arabidopsis thaliana.

Contrary to previous assumptions that most mutations are deleterious, there is increasing evidence for persistence of large-effect mutations in natural populations. A possible explanation for these observations is that mutant phenotypes and fitness may depend upon the specific environmental conditions to which a mutant is exposed. Here, we tested this hypothesis by growing large-effect flowering time mutants of Arabidopsis thaliana in multiple field sites and seasons to quantify their fitness effects in realistic natural conditions. By constructing environment-specific fitness landscapes based on flowering time and branching architecture, we observed that a subset of mutations increased fitness, but only in specific environments. These mutations increased fitness via different paths: through shifting flowering time, branching, or both. Branching was under stronger selection, but flowering time was more genetically variable, pointing to the importance of indirect selection on mutations through their pleiotropic effects on multiple phenotypes. Finally, mutations in hub genes with greater connectedness in their regulatory networks had greater effects on both phenotypes and fitness. Together, these findings indicate that large-effect mutations may persist in populations because they influence traits that are adaptive only under specific environmental conditions. Understanding their evolutionary dynamics therefore requires measuring their effects in multiple natural environments.

Fluctuating, warm temperatures decrease the effect of a key floral repressor on flowering time in Arabidopsis thaliana.

The genetic basis of growth and development is often studied in constant laboratory environments; however, the environmental conditions that organisms experience in nature are often much more dynamic. We examined how daily temperature fluctuations, average temperature, day length and vernalization influence the flowering time of 59 genotypes of Arabidopsis thaliana with allelic perturbations known to affect flowering time. For a subset of genotypes, we also assessed treatment effects on morphology and growth. We identified 17 genotypes, many of which have high levels of the floral repressor FLOWERING LOCUS C (FLC), that bolted dramatically earlier in fluctuating - as opposed to constant - warm temperatures (mean = 22°C). This acceleration was not caused by transient VERNALIZATION INSENSITIVE 3-mediated vernalization, differential growth rates or exposure to high temperatures, and was not apparent when the average temperature was cool (mean = 12°C). Further, in constant temperatures, contrary to physiological expectations, these genotypes flowered more rapidly in cool than in warm environments. Fluctuating temperatures often reversed these responses, restoring faster bolting in warm conditions. Independently of bolting time, warm fluctuating temperature profiles also caused morphological changes associated with shade avoidance or 'high-temperature' phenotypes. Our results suggest that previous studies have overestimated the effect of the floral repressor FLC on flowering time by using constant temperature laboratory conditions.

Paths to selection on life history loci in different natural environments across the native range of Arabidopsis thaliana.

Selection on quantitative trait loci (QTL) may vary among natural environments due to differences in the genetic architecture of traits, environment-specific allelic effects or changes in the direction and magnitude of selection on specific traits. To dissect the environmental differences in selection on life history QTL across climatic regions, we grew a panel of interconnected recombinant inbred lines (RILs) of Arabidopsis thaliana in four field sites across its native European range. For each environment, we mapped QTL for growth, reproductive timing and development. Several QTL were pleiotropic across environments, three colocalizing with known functional polymorphisms in flowering time genes (CRY2, FRI and MAF2-5), but major QTL differed across field sites, showing conditional neutrality. We used structural equation models to trace selection paths from QTL to lifetime fitness in each environment. Only three QTL directly affected fruit number, measuring fitness. Most QTL had an indirect effect on fitness through their effect on bolting time or leaf length. Influence of life history traits on fitness differed dramatically across sites, resulting in different patterns of selection on reproductive timing and underlying QTL. In two oceanic field sites with high prereproductive mortality, QTL alleles contributing to early reproduction resulted in greater fruit production, conferring selective advantage, whereas alleles contributing to later reproduction resulted in larger size and higher fitness in a continental site. This demonstrates how environmental variation leads to change in both QTL effect sizes and direction of selection on traits, justifying the persistence of allelic polymorphism at life history QTL across the species range.

Candidate gene association mapping of Arabidopsis flowering time.

The pathways responsible for flowering time in Arabidopsis thaliana comprise one of the best characterized genetic networks in plants. We harness this extensive molecular genetic knowledge to identify potential flowering time quantitative trait genes (QTGs) through candidate gene association mapping using 51 flowering time loci. We genotyped common single nucleotide polymorphisms (SNPs) at these genes in 275 A. thaliana accessions that were also phenotyped for flowering time and rosette leaf number in long and short days. Using structured association techniques, we find that haplotype-tagging SNPs in 27 flowering time genes show significant associations in various trait/environment combinations. After correction for multiple testing, between 2 and 10 genes remain significantly associated with flowering time, with CO arguably possessing the most promising associations. We also genotyped a subset of these flowering time gene SNPs in an independent recombinant inbred line population derived from the intercrossing of 19 accessions. Approximately one-third of significant polymorphisms that were associated with flowering time in the accessions and genotyped in the outbred population were replicated in both mapping populations, including SNPs at the CO, FLC, VIN3, PHYD, and GA1 loci, and coding region deletions at the FRI gene. We conservatively estimate that approximately 4-14% of known flowering time genes may harbor common alleles that contribute to natural variation in this life history trait.

Effects of genetic perturbation on seasonal life history plasticity.

Like many species, the model plant Arabidopsis thaliana exhibits multiple different life histories in natural environments. We grew mutants impaired in different signaling pathways in field experiments across the species' native European range in order to dissect the mechanisms underlying this variation. Unexpectedly, mutational loss at loci implicated in the cold requirement for flowering had little effect on life history except in late-summer cohorts. A genetically informed photothermal model of progression toward flowering explained most of the observed variation and predicted an abrupt transition from autumn flowering to spring flowering in late-summer germinants. Environmental signals control the timing of this transition, creating a critical window of acute sensitivity to genetic and climatic change that may be common for seasonally regulated life history traits.

SKS6, a multicopper oxidase-like gene, participates in cotyledon vascular patterning during Arabidopsis thaliana development.

SKU5-Similar 6 (SKS6) is a one of a large gene family of 19 members in Arabidopsis thaliana (L.) Heynh that encode multicopper oxidase-like proteins that are related to ferroxidases, ascorbate oxidases and laccases. Only one member of the family has been previously studied; Skewed5 (SKU5) is involved in the control of root growth. The encoded SKS6 protein, like SKU5 appears to lack a functional copper-binding site and is most closely related to Bp10 from Brassica napus and Ntp303 from Nicotiana tobacum. The SKS6 promoter contains many putative regulatory sites and differential expression of an SKS6::GUS reporter gene revealed selective induction in several seedling tissues including guard cells, root cortex cells, and leaf margin hydathodes. It was also expressed later in flower development in flower primordia, ovules, and the abscission zones of seeds and siliques. Furthermore, SKS6 was upregulated in roots in response to treatment of seedlings with the hormones abscisic acid, indole-3 acetic acid, 2,4-dichlorophenoxyacetic acid and aminocyclopropane-1-carboxylate. A loss-of function sks6-1 T-DNA insertion allele revealed that cotyledon vascular patterning is affected in the mutant, suggesting a role for the protein in metabolism of nutrients or hormones in the hydathodes, the sites of auxin synthesis and chemical recycling.

TOUSLED kinase activity oscillates during the cell cycle and interacts with chromatin regulators.

The TOUSLED (TSL)-like nuclear protein kinase family is highly conserved in plants and animals. tsl loss of function mutations cause pleiotropic defects in both leaf and flower development, and growth and initiation of floral organ primordia is abnormal, suggesting that basic cellular processes are affected. TSL is more highly expressed in exponentially growing Arabidopsis culture cells than in stationary, nondividing cells. While its expression remains constant throughout the cell cycle in dividing cells, TSL kinase activity is higher in enriched late G2/M-phase and G1-phase populations of Arabidopsis suspension culture cells compared to those in S-phase. tsl mutants also display an aberrant pattern and increased expression levels of the mitotic cyclin gene CycB1;1, suggesting that TSL represses CycB1;1 expression at certain times during development or that cells are delayed in mitosis. TSL interacts with and phosphorylates one of two Arabidopsis homologs of the nucleosome assembly/silencing protein Asf1 and histone H3, as in humans, and a novel plant SANT/myb-domain protein, TKI1, suggesting that TSL plays a role in chromatin metabolism.

Rice phospholipase D isoforms show differential cellular location and gene induction.

Phospholipase D (PLD) has emerged as an important enzyme involved in signal transduction, stress responses, protein trafficking, and membrane metabolism. This report describes the cloning and characterization of three novel PLD genes from rice, designated RPLD3, RPLD4 and RPLD5. The rice PLDs, including the previously isolated RPLD1 and RPLD2, are similar to PLD subfamilies of Arabidopsis: Based on sequence homology and domain conservation, RPLD1 is most similar to the PLDalpha subfamily of PLDs while RPLD5 most closely resembles the PLDdelta type. RPLD2, 3 and 4 represent a unique subfamily, although they are most similar to PLDalpha. RPLD1 is located on chromosome 1, RPLD5 on chromosome 3, and RPLD2, RPLD3, and RPLD4 are tandemly arrayed on chromosome 5. Transcriptional analysis reveals that RPLD1, present in healthy rice vegetative tissues, is induced rapidly but transiently in wounded leaf tissues. RPLD2, also induced by wounding, is present at lower levels but for a more prolonged duration than RPLD1. Immunolocalization with peptide specific antibodies to each of the five PLDs was used to demonstrate that the isoforms have overlapping but distinct patterns of distribution in healthy rice cells. RPLD1 was detected in mesophyll cell wall, membranes, and chloroplasts, whereas RPLD3 and RPLD4 were located predominantly in the chloroplasts. Labeling of RPLD2 and RPLD5 was sparse, and was most concentrated in the secondary walls of xylem (RPLD2) and guard cells (RPLD2 and RPLD5). This combined information on structural features, expression profiles, and cellular localization will assist the basis for dissection of PLD isoform function in rice.

The F-box-containing protein UFO and AGAMOUS participate in antagonistic pathways governing early petal development in Arabidopsis.

The UNUSUAL FLORAL ORGANS (UFO) gene is required for multiple processes in the developing Arabidopsis flower, including the proper patterning and identity of both petals and stamens. The gene encodes an F-box-containing protein, UFO, which interacts physically and genetically with the Skp1 homolog, ASK1. In this report, we describe four ufo alleles characterized by the absence of petals, which uncover another role for UFO in promoting second whorl development. This UFO-dependent pathway is required regardless of the second whorl organ to be formed, arguing that it affects a basic process acting in parallel with those establishing organ identity. However, the pathway is dispensable in the absence of AGAMOUS (AG), a known inhibitor of petal development. In situ hybridization results argue that AG is not transcribed in the petal region, suggesting that it acts non-cell-autonomously to inhibit second whorl development in ufo mutants. These results are combined into a genetic model explaining early second whorl initiation/proliferation, in which UFO functions to inhibit an AG-dependent activity.