Positional cloning and characterization reveal the role of a miRNA precursor gene ZmLRT in the regulation of lateral root number and drought tolerance in maize.

Lateral roots play essential roles in drought tolerance in maize (Zea mays L.). However, the genetic basis for the variation in the number of lateral roots in maize remains elusive. Here, we identified a major quantitative trait locus (QTL), qLRT5-1, controlling lateral root number using a recombinant inbred population from a cross between the maize lines Zong3 (with many lateral roots) and 87-1 (with few lateral roots). Fine-mapping and functional analysis determined that the candidate gene for qLRT5-1, ZmLRT, expresses the primary transcript for the microRNA miR166a. ZmLRT was highly expressed in root tips and lateral root primordia, and knockout and overexpression of ZmLRT increased and decreased lateral root number, respectively. Compared with 87-1, the ZmLRT gene model of Zong3 lacked the second and third exons and contained a 14 bp deletion at the junction between the first exon and intron, which altered the splicing site. In addition, ZmLRT expression was significantly lower in Zong3 than in 87-1, which might be attributed to the insertions of a transposon and over large DNA fragments in the Zong3 ZmLRT promoter region. These mutations decreased the abundance of mature miR166a in Zong3, resulting in increased lateral roots at the seedling stage. Furthermore, miR166a post-transcriptionally repressed five development-related class-III homeodomain-leucine zipper genes. Moreover, knockout of ZmLRT enhanced drought tolerance of maize seedlings. Our study furthers our understanding of the genetic basis of lateral root number variation in maize and highlights ZmLRT as a target for improving drought tolerance in maize.

A C-terminal encoded peptide, ZmCEP1, is essential for kernel development in maize.

C-terminal encoded peptides (CEPs) are peptide hormones which act as mobile signals coordinating important developmental programs. Previous studies have unraveled that CEPs are able to regulate plant growth and abiotic stress via cell-to-cell communication in Arabidopsis and rice; however, little is known about their roles in maize. Here, we examined the spatiotemporal expression pattern of ZmCEP1 and showed that ZmCEP1 is highly expressed in young ears and tassels of maize, particularly in the vascular bundles of ears. Heterologous expression of ZmCEP1 in Arabidopsis results in smaller plants and seed size. Similarly, overexpression of ZmCEP1 in maize decreased the plant and ear height, ear length, kernel size, and 100-kernel weight. Consistently, exogenous application of the synthesized ZmCEP1 peptide to the roots of Arabidopsis and maize inhibited root elongation. Knock-out of ZmCEP1 through CRISPR/Cas9 significantly increased plant and ear height, kernel size and 100-kernel weight. Transcriptome analysis revealed that knock-out of ZmCEP1 up-regulated a subset of genes involved in nitrogen metabolism, nitrate transport, sugar transport and auxin response. Thus, these results provide new insights into the genetic and molecular function of ZmCEP1 in regulating kernel development and plant growth, providing novel opportunities for maize breeding.

Up-regulating the abscisic acid inactivation gene ZmABA8ox1b contributes to seed germination heterosis by promoting cell expansion.

Heterosis has been widely used in agriculture, but the underlying molecular principles are still largely unknown. During seed germination, we observed that maize (Zea mays) hybrid B73/Mo17 was less sensitive than its parental inbred lines to exogenous abscisic acid (ABA), and endogenous ABA content in hybrid embryos decreased more rapidly than in the parental inbred lines. ZmABA8ox1b, an ABA inactivation gene, was consistently more highly up-regulated in hybrid B73/Mo17 than in its parental inbred lines at early stages of seed germination. Moreover, ectopic expression of ZmABA8ox1b obviously promoted seed germination in Arabidopsis Remarkably, microscopic observation revealed that cell expansion played a major role in the ABA-mediated maize seed germination heterosis, which could be attributed to the altered expression of cell wall-related genes.

Overexpression of Peptide-Encoding OsCEP6.1 Results in Pleiotropic Effects on Growth in Rice (O. sativa).

Plant peptide hormones play an important role in regulating plant developmental programs via cell-to-cell communication in a non-cell autonomous manner. To characterize the biological relevance of C-TERMINALLY ENCODED PEPTIDE (CEP) genes in rice, we performed a genome-wide search against public databases using a bioinformatics approach and identified six additional CEP members. Expression analysis revealed a spatial-temporal pattern of OsCEP6.1 gene in different tissues and at different developmental stages of panicle. Interestingly, the expression level of the OsCEP6.1 was also significantly up-regulated by exogenous cytokinin. Application of a chemically synthesized 15-amino acid OsCEP6.1 peptide showed that OsCEP6.1 had a negative role in regulating root and seedling growth, which was further confirmed by transgenic lines. Furthermore, the constitutive expression of OsCEP6.1 was sufficient to lead to panicle architecture and grain size variations. Scanning electron microscopy analysis revealed that the phenotypic variation of OsCEP6.1 overexpression lines resulted from decreased cell size but not reduced cell number. Moreover, starch accumulation was not significantly affected. Taken together, these data suggest that the OsCEP6.1 peptide might be involved in regulating the development of panicles and grains in rice.

Ectopic expression of a maize hybrid up-regulated gene, ErbB-3 binding Protein 1 (ZmEBP1), increases organ size by promoting cell proliferation in Arabidopsis.

The alteration of gene expression in hybrids may be an important factor promoting phenotypic variation and plasticity. To provide insight into the underlying molecular basis of maize heterosis in terms of the kernel number per ear, we established DGE profiles for the immature ears of maize hybrid Zong3/87-1 and its parental lines at the floral organ differentiation stage. Among 4,337 identified differentially expressed genes, 4,021 (92%) exhibited nonadditive expression patterns in the hybrid. Notably, the maize homolog of Arabidopsis EBP1, designated ZmEBP1, displayed an overdominant expression pattern in the Zong3/87-1 hybrid. Moreover, the results of qRT-PCR revealed that the ZmEBP1 gene was upregulated in the immature ears of the reciprocal hybrids Zong3/87-1 and 87-1/Zong3 at different developmental stages. Additionally, ectopic expression of ZmEBP1 in Arabidopsis increased organ size, which was mainly attributed to an increase in cell numbers, rather than cell size. Considering all of these findings together, we speculate that upregulation of ZmEBP1 in maize hybrids may accelerate cell proliferation and promote ear development.