Zhiyong Zhang, Jiaqiang Dong, Chen Ji, Yongrui Wu, and Joachim Messing

PNAS June 4, 2019 116 (23) 11223-11228;

Agricultural Sciences

Significance

World food supply depends on improving grain yield and quality, which are determined by accumulation of starch and proteins in maize endosperm, respectively. Because initiation of synthesis of the two compounds occurs 8 to 10 d after pollination in starchy endosperm cells, regulatory factors must control their coordinated accumulation. This work shows that two maize endosperm-specific transcription factors can coordinate the accumulation of starch and proteins by regulating the expression of key starch biosynthetic enzymes and the major seed proteins.

Abstract

Grain starch and protein are synthesized during endosperm development, prompting the question of what regulatory mechanism underlies the synchronization of the accumulation of secondary and primary gene products. We found that two endosperm-specific NAC transcription factors, ZmNAC128 and ZmNAC130, have such a regulatory function. Knockdown of expression of ZmNAC128 and ZmNAC130 with RNA interference (RNAi) caused a shrunken kernel phenotype with significant reduction of starch and protein. We could show that ZmNAC128 and ZmNAC130 regulate the transcription of Bt2 and then reduce its protein level, a rate-limiting step in starch synthesis of maize endosperm. Lack of ZmNAC128 and ZmNAC130 also reduced accumulation of zeins and nonzeins by 18% and 24% compared with nontransgenic siblings, respectively. Although ZmNAC128 and ZmNAC130 affected expression of zein genes in general, they specifically activated transcription of the 16-kDa γ-zein gene. The two transcription factors did not dimerize with each other but exemplified redundancy, whereas individual discovery of their function was not amenable to conventional genetics but illustrated the power of RNAi. Given that both the Bt2 and the 16-kDa γ-zein genes were activated by ZmNAC128 or ZmNAC130, we could identify a core binding site ACGCAA contained within their target promoter regions by combining Dual-Luciferase Reporter and Electrophoretic Mobility Shift assays. Consistent with these properties, transcriptomic profiling uncovered that lack of ZmNAC128 and ZmNAC130 had a pleiotropic effect on the utilization of carbohydrates and amino acids.

See https://www.pnas.org/content/116/23/11223

Figure 1:

The phenotypes of nacRNAi. (A) Kernel phenotypes of nacRNAi#4 in the B73 background. In the Upper, the white arrowheads point out the transgenic nacRNAi kernels in the cob. The Lower Left and Right represent the nontransgenic (NT) and nacRNAi kernels from the same cob, respectively. (B) Kernel weight (Upper) and test weight (Lower) of NT and nacRNAi seeds. The data are measured from the three mature cobs of nacRNAi#4 in the B73 background with ±SD. g, gram; g/mL, gram per milliliter. (C) The content of starch, zeins, and nonzeins in the mature kernels of NT and nacRNAi siblings. Percent represents mg per 100 mg mature dry kernels. In B and C, the asterisk represents a significant difference from NT (Student’s t test, P < 0.05).