The most critical step in maize domestication (Zea mays ssp. mays) was the liberation of the kernel from the hardened, protective casing that envelops the kernel in the maize progenitor, teosinte 1 . This evolutionary step exposed the kernel on the surface of the ear such that it could be readily utilized as a food source by humans. Here, we show that this key event in maize domestication is controlled by a single gene (teosinte glume architecture; tga1) belonging to the SBP-domain family 2 of transcriptional regulators. The factor controlling the phenotypic difference between maize and teosinte maps to a 1 kilobase region within which maize and teosinte show only six fixed differences in their DNA sequences. One of these differences encodes a non-conservative amino acid substitution and may affect protein function, while the other five differences potentially affect gene regulation. Molecular evolution analyses show that this region was the target of selection during maize domestication. Our results demonstrate that modest genetic changes in single genes can induce dramatic changes in phenotype during domestication and evolution.The origin of the maize ear has been considered one of the greatest mysteries in both crop domestication 3 and plant evolution 4 . While a wealth of botanical and genetic information has identified the wild Mexican grass, teosinte (Zea mays ssp. parviglumis), as the direct progenitor of maize, the profound differences in the structure of the maize and teosinte female inflorescences (ears) has posed a challenge to formulating a compelling model for the developmental and genetic steps involved in this evolutionary transition 3 . At the heart of the problem is the fact that teosinte kernels are tightly encased in structures called cupulate fruitcases, while maize kernels are borne uncovered on the surface of the ear (Figure 1a, b). The strength with which the fruitcase envelops the teosinte kernel and the stony nature of this casing far exceed the relatively flimsy and loosely bound chaff that surrounds the kernels of the ancestors of the other domesticated cereals. Indeed, the stony fruitcase of teosinte had been considered such an obstacle to the use of teosinte as a grain that teosinte was dismissed by some as a possible progenitor of maize 5 . It was argued that the genetic steps to free the grain from this casing and thereby convert teosinte into a useful crop were too complex to have arisen under domestication.Each of the 5 to 12 cupulate fruitcases in a teosinte ear is formed from an invaginated internode (cupule) within which the kernel sits, and a glume that covers the opening of the cupule such that the kernel is completely hidden from view (Figure 1b, d). When mature, the teosinte ear

teosinte glume architecture1 (tga1), a member of the SBP-box gene family of transcriptional regulators, has been identified as the gene conferring naked kernels in maize vs. encased kernels in its wild progenitor, teosinte. However, the identity of the causative polymorphism within tga1 that produces these different phenotypes has remained unknown. Using nucleotide diversity data, we show that there is a single fixed nucleotide difference between maize and teosinte in tga1, and this difference confers a Lys (teosinte allele) to Asn (maize allele) substitution. This substitution transforms TGA1 into a transcriptional repressor. While both alleles of TGA1 can bind a GTAC motif, maize-TGA1 forms more stable dimers than teosinte-TGA1. Since it is the only fixed difference between maize and teosinte, this alteration in protein function likely underlies the differences in maize and teosinte glume architecture. We previously reported a difference in TGA1 protein abundance between maize and teosinte based on relative signal intensity of a Western blot. Here, we show that this signal difference is not due to tga1 but to a second gene, neighbor of tga1 (not1). Not1 encodes a protein that has 92% amino acid similarity to TGA1 and that is recognized by the TGA1 antibody. Genetic mapping and phenotypic data show that tga1, without a contribution from not1, controls the difference in covered vs. naked kernels. No trait differences could be associated with the maize vs. teosinte alleles of not1. Our results document how morphological evolution can be driven by a simple nucleotide change that alters protein function. KEYWORDS tga1; glume architecture; teosinte; maize; domestication A LTHOUGH the study of adaptive evolution through natural or artificial selection dates back to Darwin, the genetic mechanisms that drive changes in morphology remain strongly debated (Stern and Orgogozo 2008). Questions surrounding the number of genes (few or many) and types of mutations (regulatory or coding) in these genes have been a focus in this debate (Carroll 2005(Carroll , 2008Hoekstra and Coyne 2007). These questions can only be answered through the genetic and molecular dissection of genes that have undergone selective pressure between lineages or within populations. Crop species offer a powerful system for investigating these questions since crops are the products of continuous directional selection to adapt them to the human controlled environment and human needs (Meyer and Purugganan 2013). Research on crop models is further facilitated by the extensive genetic and genomic resources available for them. Moreover, because of the recent divergence of crop species from their wild progenitors, crop-progenitor pairs remain cross-compatible and amenable to genetic analysis.Maize was domesticated in the central Balsas valley of Mexico 9000 years ago from a wild relative called teosinte (Piperno et al. 2001;Matsuoka et al. 2002). Because the morphology of modern maize is drastically different from teosinte species, the progenitor of mai...

Selection during evolution, whether natural or artificial, acts through the phenotype. For multifaceted phenotypes such as plant and inflorescence architecture, the underlying genetic architecture is comprised of a complex network of interacting genes rather than single genes that act independently to determine the trait. As such, selection acts on entire gene networks. Here, we begin to define the genetic regulatory network to which the maize domestication gene, (), belongs. Using a combination of molecular methods to uncover either direct or indirect regulatory interactions, we identified a set of genes that lie downstream of in a gene network regulating both plant and inflorescence architecture. Additional genes, known from the literature, also act in this network. We observed that regulates both core cell cycle genes and another maize domestication gene, (). We show that several members of the MADS-box gene family are either directly or indirectly regulated by and/or, and that sits atop a cascade of transcriptional regulators controlling both plant and inflorescence architecture. Multiple members of the network appear to have been the targets of selection during maize domestication. Knowledge of the regulatory hierarchies controlling traits is central to understanding how new morphologies evolve.

Summary• Hardened floral bracts and modifications to the inflorescence axis of grasses have been hypothesized to protect seeds from predation and ⁄ or aid seed dispersal, and have evolved multiple times independently within the family. Previous studies have demonstrated that mutations in the maize (Zea mays ssp. mays) gene teosinte glume architecture (tga1) underlie a reduction in hardened structures, yielding free fruits that are easy to harvest. It remains unclear whether the causative mutation(s) occurred in the cis-regulatory or protein-coding regions of tga1, and whether similar mutations in TGA1-like genes can explain variation in the dispersal unit in related grasses.• To address these questions TGA1-like genes were cloned and sequenced from a number of grasses and analyzed phylogenetically in relation to morphology; protein expression was investigated by immunolocalization.• TGA1-like proteins were expressed throughout the spikelet in the early development of all grasses, and throughout the flower of the grass relative Joinvillea. Later in development, expression patterns differed between Tripsacum dactyloides, maize and teosinte (Z. mays ssp. parviglumis).• These results suggest an ancestral role for TGA1-like genes in early spikelet development, but do not support the hypothesis that TGA1-like genes have been repeatedly modified to affect glume and inflorescence axis diversification.