Interactions of C₄ Subtype Metabolic Activities and Transport in Maize Are Revealed through the Characterization of DCT2 Mutants

Abstract

C 4 photosynthesis in grasses requires the coordinated movement of metabolites through two specialized leaf cell types, mesophyll (M) and bundle sheath (BS), to concentrate CO 2 around Rubisco. Despite the importance of transporters in this process, few have been identified or rigorously characterized. In maize (Zea mays), DCT2 has been proposed to function as a plastid-localized malate transporter and is preferentially expressed in BS cells. Here, we characterized the role of DCT2 in maize leaves using Activator-tagged mutant alleles. Our results indicate that DCT2 enables the transport of malate into the BS chloroplast. Isotopic labeling experiments show that the loss of DCT2 results in markedly different metabolic network operation and dramatically reduced biomass production. In the absence of a functioning malate shuttle, dct2 lines survive through the enhanced use of the phosphoenolpyruvate carboxykinase carbon shuttle pathway that in wild-type maize accounts for ;25% of the photosynthetic activity. The results emphasize the importance of malate transport during C 4 photosynthesis, define the role of a primary malate transporter in BS cells, and support a model for carbon exchange between BS and M cells in maize.

Characterization of maize leaf pyruvate orthophosphate dikinase using high throughput sequencing

“…Comparing our study with others (Eastmond et al ; Weissmann et al ) allows for the integration of additional insights. C 4 photosynthesis in maize occurs through two shuttles, one linked to NADP‐ME, one depending on PPDK to supply PEP for PEPC, and an aspartate shuttle linked to PEPCK (Furbank ).…”

DCT4- a new member of the dicarboxylate transporter family in C4grasses

“…In C 4 species, the expression of many photosynthetic genes is enriched in either BS and M cells (Li et al, 2010; John et al, 2014; Tausta et al, 2014; Weissmann et al, 2015; Rao et al, 2016). In NADP-ME species, two transporters, one within the BS cells and another in M cells, move malate in and out of the chloroplast during C 4 photosynthesis (Brautigam et al ., 2008; Weissmann and Brutnell, 2012; John et al ., 2014; Tausta et al ., 2014; Wang et al ., 2014a).…”

“…This supposition is in agreement with the view that the three subtypes of C 4 photosynthesis are mixed rather than exclusive (Hatch, 1971; Chapman and Hatch, 1979; Furbank, 2011; Pick et al ., 2011; Wang et al ., 2014b). For example, Z. mays utilizes both the NADP-ME (75%) and PEPCK (25%) pathways to fix carbon (Chapman and Hatch, 1979; Wingler et al, 1999; Weissmann et al, 2015), and has similar expression levels of DCT2 and OMT1 and low expression of DCT1. S. bicolor moves carbon through both malate and aspartate, although no PEPCK activity was detected in its leaves (Chapman and Hatch, 1979).…”

“…Interestingly, both ZmOMT1 and SbOMT1 are only slightly differentially expressed in the M cells (Figure 2). As the loss of DCT2 in Z. mays prevents movement of malate into the BS chloroplast (Weissmann et al, 2015), OMT1 cannot be moving malate into the BS chloroplast alongside DCT2 . But OMT1 may also have a role in organic acid metabolism in both cell types, such as shuttling reducing equivalents in organelles other than the chloroplast (Pleite et al ., 2005).…”

Understanding metabolite transport and metabolism in C4 plants through RNA-seq

“…DiT2 is moderately upregulated in conjunction with NADP-ME [6][7][8] and DiT2 is strongly expressed in maize bundle sheath cells (Table 1) and S. bicolor bundle sheath cells [25]. Mutant analysis of maize showed symptoms consistent with a role in malate import in the C 4 cycle [37 ] raising the question whether the original assay was unsuitable for detecting the exchange [36] or whether Flaveria and maize may use different transporters (Figure 1d). The molecular identity of the pyruvate exporter is currently unknown but it cannot be excluded that pyruvate may rely on diffusion to pass the chloroplast membrane in its electroneutral form, pyruvic acid [38,39], especially given the high concentrations of pyruvate generated by NADP-ME.…”