NAC-type transcription factors regulate accumulation of starch and protein in maize seeds

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. maize endosperm | gene regulation | starch synthesis | protein

TubZIP28, a novel bZIP family transcription factor from Triticum urartu, and TabZIP28, its homologue from Triticum aestivum, enhance starch synthesis in wheat

“…For example, O2 and PBF control both SSP and starch synthesis in maize endosperms (Zhang et al , ). Moreover, ZmNAC128 and ZmNAC130 regulate the accumulation of both SSPs and starch in maize seeds (Zhang et al , ). Apart from the activation on starch synthesis, whether TubZIP28 regulated SSP synthesis was of major interest.…”

“…Seeds of 10 randomly selected plants in each transgenic line and WT were used for flour grinding. In the total starch content determination, 100 mg flour was used and the content was measured using Megazyme (Irishtown, Ireland) Total Starch Assay Kit (catalog: K‐TSTA) according to the manufacturer's instructions (Botticella et al , ; Zhang et al , ). The percentage of milligrams per 100 mg of flour was used to represent the total starch content.…”

“…Thus far, there have been a few reports on the transcriptional regulation of starch synthesis in cereals, and only several transcription factors (TFs) have been identified as transcriptional regulators of SSRGs. Rice Starch Regulator 1 ( RSR1 ) and OsbZIP58 are starch synthesis regulators in rice (Fu & Xue, ; Wang et al , ); ZmNAC36 , ZmbZIP91 , ZmEREB156 , ZmNAC128 , and ZmNAC130 are regulators in maize, and SUSIBA2 is the regulator in barley (Sun et al , , Zhang et al , ; Chen et al , ; Huang et al , ; López‐González et al , ; Zhang et al , ). In common wheat, no regulator of starch synthesis has been identified owing to the complexity of its polyploid genome, except for the TaRSR1 characterized through homology‐based cloning of the RSR1 in rice (Liu et al , ).…”

An update on the maize zein-gene family in the post-genomics era

“…Besides interacting in-vivo with 27-kD γ-zein gene, ZmbZIP22 additionally targets OHPs, O2 and PBF, thus synergistically mediate the expression of 27-kD γ-zein (Li et al 2018, b). More recently, two new transcription factors ZmNAC128 and ZmNAC130 were shown to have regulatory functions, coordinating the accumulation of starch and proteins at filling stage from 10 to 35 days post pollination through the transcriptional regulation of the 16-kDa γ-zein genes and at least Bt2 (Zhang, Dong, Ji, Wu & Messing 2019). The current advances in genome sequencing technologies and availability of high quality reference genome of maize lines will further assist in hunting for more TFs that could possibly shed light on the molecular mechanism underlying zein gene expression and regulation.…”

A Heat Stress Responsive NAC Transcription Factor Heterodimer Plays Key Roles in Rice Grain Filling

“…We selected four of these genes, ONAC025 , ONAC127 , ONAC128 and ONAC129 for further analyses, which located in the linked regions of chromosome 11 and were identified as a gene cluster (Fang et al, 2008). As recent study implied two NAC transcription factors specifically expressed in maize seeds, ZmNAC128 and ZmNAC130 , playing critical roles in starch and protein accumulation in seeds filling (Zhang et al, 2019). We inferred that the four NAC transcriptional factors might also be involved in some important processes in rice caryopsis development.…”