De novo Short Read Assembly and Functional Annotation of Eleocharis vivipara , a C 3 /C 4 Interconvertible Sedge Plant

Abstract

More than 90% of terrestrial plant species possess the C3 photosynthetic system, while only about 3% undergo C4 photosynthesis. Nevertheless, C4 plants are responsible for 25% of the annual total terrestrial plant biomass production (Langdale, 2011; Sage and Zhu, 2011). Compared with C3 plants, C4 plants generally exhibit higher CO2 assimilation abilities (about 1.5 2-fold higher in individual blades), higher yields per growing area (approximately 2 3-fold) and higher water and nitrogen use efficiencies (Black, 1979). The productivity of major C3 crops, such as rice, wheat and soybean, could be greatly increased if the CO2 concentrating mechanism of C4 photosynthesis could be introduced into these C3 plants through genetic engineering. Such attempts have been made by various investigators (Häusler et al., 2002; Miyao et al., 2011), but no promising results have been obtained thus far. For example, several genes encoding C4 photosynthesis-related enzymes were successfully overexpressed individually or in combination in mesophyll cells of rice plants, but the transgenic plants did not perform C4-like photosynthesis and their growth rate was not accelerated appreciably (Taniguchi et al., 2008). These observations imply the introduction of genes for CO2-concentrating proteins is not sufficient to confer the C4 photosynthetic framework to C3 plants. In fact, two functionally differentiated cell types mesophyll cells and bundle-sheath cells and their structural arrangement called Kranz anatomy are required to concentrate CO2 and deliver it to ribulose-1,5-bisphosphate carboxylase/oxygenase. This CO2 concentration system has been thought to be essential for C4 photosynthesis. To convert C3 plants into C4 ones, all genes that are related to cellular and functional C4 differentiation, including those associated with C4 metabolism, transport of C4-related compounds and development of Kranz anatomy, must be identified and introduced into C3 plants (Covshoff and Hibberd, 2012). To identify the genes required for C4 photosynthesis, a straightforward approach was comparative analysis of C4 and C3 plants. However, identification of the key C4 photosynthetic genes is difficult because many of them are species-specific even among C4 model plants (e.g., Zea mays and Sorghum bicolor). Although the genera Flaveria and Cleome include both C3 and C4 species and have attracted much attention as alternative systems (Brown et al., 2005; Külahoglu et al., 2014), there are still speciesspecific genes, which makes it difficult to identify the core functional set of C4 photosynthetic genes (Gowik et al., 2011). Eleocharis vivipara (Cyperaceae), first investigated by Ueno et al. (1988) is an amphibious leafless sedge. This plant develops Kranz anatomy and shows C4 biochemical traits under terrestrial conditions, and performs NADdependent malic enzyme (NAD-ME)-type C4 photosynthesis. Interestingly, under submerged conditions, it grows without Kranz anatomy and exhibits C3 biochemical traits. When the submerged plants are exposed to air, they develop new shoots with C4 traits within about a week. This species is thus suitable for screening genes indispensable to the biochemical, cellular and structural transition from the C3 to the C4 photosynthetic system; however, its relevant genomic and transcriptomic information is currently lim-