Identification and characterization of a novel stay-green QTL that increases yield in maize.

Functional stay-green is a valuable trait that extends the photosynthetic period, increases source capacity and biomass and ultimately translates to higher grain yield. Selection for higher yields has increased stay-green in modern maize hybrids. Here, we report a novel QTL controlling functional stay-green that was discovered in a mapping population derived from the Illinois High Protein 1 (IHP1) and Illinois Low Protein 1 (ILP1) lines, which show very different rates of leaf senescence. This QTL was mapped to a single gene containing a NAC-domain transcription factor that we named nac7. Transgenic maize lines where nac7 was down-regulated by RNAi showed delayed senescence and increased both biomass and nitrogen accumulation in vegetative tissues, demonstrating NAC7 functions as a negative regulator of the stay-green trait. More importantly, crosses between nac7 RNAi parents and two different elite inbred testers produced hybrids with prolonged stay-green and increased grain yield by an average 0.29 megagram/hectare (4.6 bushel/acre), in 2 years of multi-environment field trials. Subsequent RNAseq experiments, one employing nac7 RNAi leaves and the other using leaf protoplasts overexpressing Nac7, revealed an important role for NAC7 in regulating genes in photosynthesis, chlorophyll degradation and protein turnover pathways that each contribute to the functional stay-green phenotype. We further determined the putative target of NAC7 and provided a logical extension for the role of NAC7 in regulating resource allocation from vegetative source to reproductive sink tissues. Collectively, our findings make a compelling case for NAC7 as a target for improving functional stay-green and yields in maize and other crops.

A transposable element insertion within ZmGE2 gene is associated with increase in embryo to endosperm ratio in maize.

Most of the maize kernel oil is located in the embryo while the majority of starch is located in the endosperm. Maize kernel composition and value are affected significantly by the ratio of the embryo size to the endosperm size; however, the genetic regulation of embryo to endosperm ratio (EER) in maize is unknown. Here we identified ZmGE2 gene, which encodes a cytochrome p450 protein, as a gene associated with EER variation in maize. We first expressed rice Giant Embryo (GE) gene driven by oleosin promoter in maize and detected a 23.2 % reduction in EER in transgenic seeds, demonstrating the existence of evolutionarily conserved mechanisms for EER determination in rice and maize. We next identified maize GE2, a homolog of rice GE sharing 70 % identity in amino sequence, as a candidate based on the similar expression pattern and co-localization with a previously detected QTL for EER. Followed by linkage and association mapping, a 247-bp transposable element (TE) insertion in 3'-untranslated region of ZmGE2 gene was identified to be associated with increase in EER and kernel oil content. Expression level of the favorable ZmGE2 allele containing the 247-bp TE insertion was strongly reduced. In addition, the 247-bp TE insertion site was a selection target during the artificial long-term selection for the high EER trait in a high oil population. This is the first report that demonstrates an association of ZmGE2 with EER variation in maize and identifies ZmGE2 gene as a promising target for manipulation of EER and grain composition by either transgenic approach or molecular breeding in maize.

Expression of ZmLEC1 and ZmWRI1 increases seed oil production in maize.

Increasing seed oil production is a major goal for global agriculture to meet the strong demand for oil consumption by humans and for biodiesel production. Previous studies to increase oil synthesis in plants have focused mainly on manipulation of oil pathway genes. As an alternative to single-enzyme approaches, transcription factors provide an attractive solution for altering complex traits, with the caveat that transcription factors may face the challenge of undesirable pleiotropic effects. Here, we report that overexpression of maize (Zea mays) LEAFY COTYLEDON1 (ZmLEC1) increases seed oil by as much as 48% but reduces seed germination and leaf growth in maize. To uncouple oil increase from the undesirable agronomic traits, we identified a LEC1 downstream transcription factor, maize WRINKLED1 (ZmWRI1). Overexpression of ZmWRI1 results in an oil increase similar to overexpression of ZmLEC1 without affecting germination, seedling growth, or grain yield. These results emphasize the importance of field testing for developing a commercial high-oil product and highlight ZmWRI1 as a promising target for increasing oil production in crops.

A phenylalanine in DGAT is a key determinant of oil content and composition in maize.

Plant oil is an important renewable resource for biodiesel production and for dietary consumption by humans and livestock. Through genetic mapping of the oil trait in plants, studies have reported multiple quantitative trait loci (QTLs) with small effects, but the molecular basis of oil QTLs remains largely unknown. Here we show that a high-oil QTL (qHO6) affecting maize seed oil and oleic-acid contents encodes an acyl-CoA:diacylglycerol acyltransferase (DGAT1-2), which catalyzes the final step of oil synthesis. We further show that a phenylalanine insertion in DGAT1-2 at position 469 (F469) is responsible for the increased oil and oleic-acid contents. The DGAT1-2 allele with F469 is ancestral, whereas the allele without F469 is a more recent mutant selected by domestication or breeding. Ectopic expression of the high-oil DGAT1-2 allele increases oil and oleic-acid contents by up to 41% and 107%, respectively. This work provides insights into the molecular basis of natural variation of oil and oleic-acid contents in plants and highlights DGAT as a promising target for increasing oil and oleic-acid contents in other crops.