Regulating the Regulators: The Future Prospects for Transcription-Factor-Based Agricultural Biotechnology Products

Abstract

It is now more than a decade since the first commercially successful genetically engineered agricultural crops were launched (Castle et al., 2006). These first products were based in large part on simple monogenic traits, such as herbicide tolerance or insect resistance, which did not require manipulation of complex molecular pathways in the transgenic plant. Since then, there has been a growing expectation that the biotechnology industry will deliver a second generation of transgenic products for more challenging traits relating to yield and yield stability, which are under complex polygenic control (Gutterson and Zhang, 2004; Salmeron and Herrera-Estrella, 2006). Advances in plant genomics and systems biology, including the availability of the complete genome sequences of both Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa), have offered an unprecedented opportunity to identify regulatory genes and networks that control these important traits. Because transcription factors (TFs) naturally act as master regulators of cellular processes, they are expected to be excellent candidates for modifying complex traits in crop plants, and TF-based technologies are likely to be a prominent part of the next generation of successful biotechnology crops. In this article, we review the prospects for modification of these target traits by TF regulation, including some of the challenges associated with such a strategy. Although transcriptional regulators are being proposed as the wave of the future for agricultural biotechnology, there is strong evidence that TFs have already played a major role in the origin of agriculture through the domestication of various crop plants. This subject is covered in detail in a review by Doebley et al. (2006), with some important examples for three major crops presented here. Probably the most well-known example of a TF with a critical role in a crop domestication trait is that of Teosinte branched1 (Tb1) of maize (Zea mays). Tb1 is a member of the TCP family of TFs, which are generally involved in the regulation of cell proliferation (Cubas et al., 1999). When Tb1 is expressed, it represses the outgrowth of lateral branches; maize plants carrying loss of function alleles produce numerous lateral branches (tillers; Doebley et al., 1997). During the domestication of teosinte to produce maize, an allele was selected that altered the regulation of Tb1, increasing its expression in primary auxiliary meristems. A survey of maize and teosinte alleles at this locus revealed alterations in the regulatory region of the gene and not in the coding sequence (Wang et al., 1999). Thus, selection for cis-regulatory changes that caused a change in expression of a single TF led to the dramatic shift in architecture that underlies the productivity of the most extensively grown crop in modern day North America. Shifts in expression patterns and TF activity have generated important characteristics in other grain crops. In rice, a key trait required for domestication is reduced grain shattering, which prevents the seeds from dropping off the panicles and allows for efficient harvesting of the grain. Two TFs have been identified as playing a major role in reducing grain shattering in domesticated rice plants. One of these was isolated as a quantitative trait locus (QTL) in a cross between a shattering-type ‘Indica’ cultivar and a nonshattering-type ‘Japonica’ (Konishi et al., 2006). This gene, qSH1, encodes a BEL1-type homeodomain protein that is orthologous to Arabidopsis REPLUMLESS (RPL), which is involved in the formation of an abscission zone in the Arabidopsis silique. The QTL is linked to a single nucleotide polymorphism in the regulatory sequences of the gene, not to any change in the coding sequence itself. qSH1 from the shattering cultivar is expressed in the developing abscission layer at the base of the seed, but in the nonshattering cultivar it is not expressed in this location. The expression pattern in other tissues is similar, suggesting that the selection was specifically for loss of expression in the abscission layer. This allele is thought to be an early mutation in the Japonica lineage that was selected during domestication. The other TF affecting this trait is shattering4 (sh4), allelic to sha1 (Li et al., 2006; Lin et al., 2007). SH4 is a member of the trihelix family of plant-specific TFs and was isolated as a major QTL for shattering in a cross between O. sativa and Oryza rufipogon. During domestication, a mutant allele was selected with a single amino acid substitution. Sh4 seems to be necessary for normal separation of the abscission layer that releases the grain from the panicle. For a detailed review of the history of rice domestication, including the role of TFs, the reader is referred to Kovach et al. (2007). Some of the major yield gains achieved by previous generations of conventional crop breeders have subsequently been attributed to alterations in TF activity. Beginni