MULTIPARENTAL POPULATIONS The Genetic Basis of Natural Variation in Seed Size and Seed Number and Their Trade-Off Using Arabidopsis thaliana MAGIC Lines

Offspring number and size are key traits determining an individual’s fitness and a crop’s yield. Yet, extensive natural variation within species is observed for these traits. Such variation is typically explained by trade-offs between fecundity and quality, for which an optimal solution is environmentally dependent. Understanding the genetic basis of seed size and number, as well as any possible genetic constraints preventing the maximization of both, is crucial from both an evolutionary and applied perspective. We investigated the genetic basis of natural variation in seed size and number using a set of Arabidopsis thaliana multiparent advanced generation intercross (MAGIC) lines. We also tested whether life history affects seed size, number, and their trade-off. We found that both seed size and seed number are affected by a large number of mostly nonoverlapping QTL, suggesting that seed size and seed number can evolve independently. The allele that increases seed size at most identified QTL is from the same natural accession, indicating past occurrence of directional selection for seed size. Although a significant trade-off between seed size and number is observed, its expression depends on life-history characteristics, and generally explains little variance. We conclude that the trade-off between seed size and number might have a minor role in explaining the maintenance of variation in seed size and number, and that seed size could be a valid target for selection. THE reproductive output of an organism is a critical lifehistory trait defining its fitness and is the result of both offspring number and quality. In the case of cereal crops, the number and size of seeds are also the main constituents of yield. Thus, understanding the genetic architecture of seed size and number, and any possible genetic constraints to maximizing them, is crucial from both an evolutionary and applied perspective (Sadras 2007; Van Daele et al. 2012; Kesavan et al. 2013). Despite its importance, the genetic basis of natural variation in seed size and number and their interaction with life-history traits remain poorly understood. Previous studies on the genetic basis of seed traits have predominantly used mutant screens and identified genes in key pathways involved in seed development (Garcia et al. 2003; Tzafrir et al. 2004; Adamski et al. 2009; Fang et al. 2012). However, since this approach only allows for the comparison of phenotypic effects of genes that are “on” or “off” (Koornneef et al. 2004), genes’ contribution to natural continuous variation in seed size or seed number remain largely uncharacterized. Because the effects of mutants are often dependent on the genetic background (Tonsor et al. 2005; Chou et al. 2011), a QTL mapping approach using multiple parents is ideal to identify genetic factors that can contribute to natural variation in these traits in a heterogeneous genetic background. Identification of genetic factors affecting seed traits is further complicated by potential trade-offs between them. Life-history theory suggests that if there are finite resources to be invested in reproduction, a trade-off between seed size and seed number must occur (Venable 1992). Although the seed size/number trade-off is well accepted on theoretical grounds, empirical evidence for its existence is still contentious and dependent upon the context under which it is evaluated (Venable 1992; Sadras 2007; Paul-Victor and Turnbull 2009; House et al. 2010). One possible explanation for context dependency in trade-offs is that the resources available for reproduction are not discrete from the whole plant budget. With many competing allocations within the organism, Copyright © 2014 by the Genetics Society of America doi: 10.1534/genetics.114.170746 Manuscript received July 22, 2014; accepted for publication September 30, 2014; published Early Online October 13, 2014. Supporting information is available online at http://www.genetics.org/lookup/suppl/ doi:10.1534/genetics.114.170746/-/DC1. Corresponding author: Department of Biology and Biochemistry, University of Bath, Claverton Down, Bath, BA2 5DW, United Kingdom. E-mail: [email protected] Genetics, Vol. 198, 1751–1758 December 2014 1751 trade-offs may arise not just between seed size and seed number, but also with other competing sources such as defense against biotic and abiotic stress (Bazzaz et al. 1987; Züst et al. 2011; Wituszyńska et al. 2013). Alternatively, the expression of the trade-off might be dependent on the level of resources available (Noordwijk and Jong 1986; Venable 1992; Bennett et al. 2012). Variation in life-history is common within populations; and typically, later flowering plants are larger and have more resources to invest in reproduction, reducing the expression of trade-offs (Aarssen and Clauss 1992; Clauss and Aarssen 1994; Jakobsson and Eriksson 2000; Mendez-Vigo et al. 2013). Thus, variation in seed size might be due to genetic factors with direct effects on seed size, or genetic factors with indirect effects, such as through resource uptake or life-history determinants, as well as nongenetic variation. A better understanding of natural variation in seed size therefore requires simultaneous consideration of genetic variation and life-history strategies. It is important to determine the existence and mechanism behind trade-offs, because environmentally caused trade-offs can be modified by selection or genetic manipulation. However, for trade-offs that result from genetic pleiotropy or linkage disequilibrium, responses to selection will be constrained (Lande and Arnold 1983; Roff and Fairbairn 2007; Latta and Gardner 2009). In addition, the assumption of a trade-off between seed size and seed number has shaped breeding practices (Egli 1998; Sadras 2007): Seed number has been the main target for crop improvement because it is more variable than seed size (Harper et al. 1970; Venable 1992; Sadras 2007; Sadras and Egli 2008). However, if seed size shows less environmental variation and higher heritability than seed number, seed size might be a useful target for genetic crop improvement (Sadras and Slafer 2012), but only if the trade-off can be teased apart. Here, we investigate the genetic basis of natural variation in seed size and its correlation with seed number using a set of recombinant inbred lines of A. thaliana, derived from MAGIC lines (Kover et al. 2009b). A. thaliana is an ideal model organism for the study of natural variation in seed size and number, because there is extensive variation among worldwide accessions for both of these traits and for many life-history traits (Krannitz et al. 1991; Alonso-Blanco et al. 1999; Kover et al. 2009b). Few studies have addressed the issue of QTL for seed size and number, taking into account other life-history traits (Alonso-Blanco et al. 1999; Van Daele et al. 2012), and these only used mapping lines created from two parents. Multiparental lines are better for addressing genetic correlations and possible trade-offs than traditional mapping lines, due to the larger number of alleles and recombination events. This allows mapping to smaller intervals (Kover et al. 2009b), reducing overlap in positions due to large confidence intervals. In addition, the larger number of alleles improves our ability to determine whether the distributions of allelic effects are compatible with pleiotropy. With only two alleles present, the same allele may increase the value of any two traits 50% of the time by chance alone. With multiple alleles, a significant correlation in allelic effects provides stronger evidence of a common genetic mechanism. Specifically, the following questions are addressed: (i) What is the genetic architecture underlying seed size and seed number per fruit? (ii) Is there evidence for a genetically determined seed size/number trade-off? (iii) How are seed traits and the seed size/number trade-off affected by life-history traits? Materials and Methods Plant material and growth conditions The set of MAGIC lines used here was produced by advanced intermating of 19 parental accessions of A. thaliana for four generations, followed by seven generations of inbreeding (Kover et al. 2009b). These lines have been genotyped for 1260 single nucleotide polymorphisms (SNPs), distributed throughout the five chromosomes (Chr) at a spacing of 96 kb apart, using an Illumina Golden Gate assay (Kover et al. 2009b). We have previously shown that these lines allow for QTL mapping with high resolution to chromosomal intervals smaller than 1 Mb (Kover et al. 2009b). The genotypes for all MAGIC lines are listed in Supporting Information, File S1. Three replicates of each of 700 MAGIC lines were grown in the autumn of 2009 at the University of Bath greenhouse. The greenhouse was set at 21 day/18 night and 16/8 hr of light/dark. Seeds from each line were placed in three separate 5.5-cm diameter pots filled with F2+Sand Soil (Levington, The Scotts Company) and randomly allocated to trays that held 24 pots. Trays were rotated around the greenhouse at regular intervals to ensure uniform growth conditions and mitigate positional effects.