Comparative transcriptomic analysis of maize ear heterosis during the inflorescence meristem differentiation stage.

BACKGROUND: Heterosis is widely used in many crops and is important for global food safety, and maize is one of the most successful crops to take advantage of heterosis. Gene expression patterns control the development of the maize ear, but the mechanisms by which heterosis affects transcriptional-level control are not fully understood. RESULTS: In this study, we sampled ear inflorescence meristems (IMs) from the single-segment substitution maize (Zea mays) line lx9801hlEW2b, which contains the heterotic locus hlEW2b associated with ear width, as well as the receptor parent lx9801, the test parent Zheng58, and their corresponding hybrids Zheng58 × lx9801hlEW2b (HY) and Zheng58 × lx9801 (CK). After RNA sequencing and transcriptomic analysis, 2531 unique differentially expressed genes (DEGs) were identified between the two hybrids (HY vs. CK). Our results showed that approximately 64% and 48% of DEGs exhibited additive expression in HY and CK, whereas the other genes displayed a non-additive expression pattern. The DEGs were significantly enriched in GO functional categories of multiple metabolic processes, plant organ morphogenesis, and hormone regulation. These essential processes are potentially associated with heterosis performance during the maize ear developmental stage. In particular, 125 and 100 DEGs from hybrids with allele-specific expression (ASE) were specifically identified in HY and CK, respectively. Comparison between the two hybrids suggested that ASE genes were involved in different development-related processes that may lead to the hybrid vigor phenotype during maize ear development. In addition, several critical genes involved in auxin metabolism and IM development were differentially expressed between the hybrids and showed various expression patterns (additive, non-additive, and ASE). Changes in the expression levels of these genes may lead to differences in auxin homeostasis in the IM, affecting the transcription of core genes such as WUS that control IM development. CONCLUSIONS: Our research suggests that additive, non-additive, and allele-specific expression patterns may fine-tune the expression of crucial DEGs that modulate carbohydrate and protein metabolic processes, nitrogen assimilation, and auxin metabolism to optimal levels, and these transcriptional changes may play important roles in maize ear heterosis. The results provide new information that increases our understanding of the relationship between transcriptional variation and heterosis during maize ear development, which may be helpful for clarifying the genetic and molecular mechanisms of heterosis.

Global transcriptional profiling between inbred parents and hybrids provides comprehensive insights into ear-length heterosis of maize (Zea mays).

BACKGROUND: Maize (Zea mays) ear length, which is an important yield component, exhibits strong heterosis. Understanding the potential molecular mechanisms of ear-length heterosis is critical for efficient yield-related breeding. RESULTS: Here, a joint netted pattern, including six parent-hybrid triplets, was designed on the basis of two maize lines harboring long (T121 line) and short (T126 line) ears. Global transcriptional profiling of young ears (containing meristem) was performed. Multiple comparative analyses revealed that 874 differentially expressed genes are mainly responsible for the ear-length variation between T121 and T126 lines. Among them, four key genes, Zm00001d049958, Zm00001d027359, Zm00001d048502 and Zm00001d052138, were identified as being related to meristem development, which corroborated their roles in the superior additive genetic effects on ear length in T121 line. Non-additive expression patterns were used to identify candidate genes related to ear-length heterosis. A non-additively expressed gene (Zm00001d050649) was associated with the timing of meristematic phase transition and was determined to be the homolog of tomato SELF PRUNING, which assists SINGLE FLOWER TRUSS in driving yield-related heterosis, indicating that Zm00001d050649 is a potential contributor to drive heterotic effect on ear length. CONCLUSION: Our results suggest that inbred parents provide genetic and heterotic effects on the ear lengths of their corresponding F1 hybrids through two independent pathways. These findings provide comprehensive insights into the transcriptional regulation of ear length and improve the understanding of ear-length heterosis in maize.

Maize brachytic2 (br2) suppresses the elongation of lower internodes for excessive auxin accumulation in the intercalary meristem region.

BACKGROUND: Short internodes contribute to plant dwarfism, which is exceedingly beneficial for crop production. However, the underlying mechanisms of internode elongation are complicated and have been not fully understood. RESULTS: Here, we report a maize dwarf mutant, dwarf2014 (d2014), which displays shortened lower internodes. Map-based cloning revealed that the d2014 gene is a novel br2 allele with a splicing variation, resulting in a higher expression of BR2-T02 instead of normal BR2-T01. Then, we found that the internode elongation in d2014/br2 exhibited a pattern of inhibition-normality-inhibition (transient for the ear-internode), correspondingly, at the 6-leaf, 12-leaf and 14-leaf stages. Indeed, BR2 encodes a P-glycoprotein1 (PGP1) protein that functions in auxin efflux, and our in situ hybridization assay showed that BR2 was mainly expressed in vascular bundles of the node and internode. Furthermore, significantly higher auxin concentration was detected in the stem apex of d2014 at the 6-leaf stage and strictly in the node region for the ear-internode at the 14-leaf stage. In such context, we propose that BR2/PGP1 transports auxin from node to internode through the vascular bundles, and excessive auxin accumulation in the node (immediately next to the intercalary meristem) region suppresses internode elongation of d2014. CONCLUSIONS: These findings suggest that low auxin levels mediated by BR2/PGP1 in the intercalary meristem region are crucial for internode elongation.

Genetic dissection of yield-related traits and mid-parent heterosis for those traits in maize (Zea mays L.).

BACKGROUND: Utilization of heterosis in maize could be critical in maize breeding for boosting grain yield. However, the genetic architecture of heterosis is not fully understood. To dissect the genetic basis of yield-related traits and heterosis in maize, 301 recombinant inbred lines derived from 08 to 641 × YE478 and 298 hybrids from the immortalized F2 (IF2) population were used to map quantitative trait loci (QTLs) for nine yield-related traits and mid-parent heterosis. RESULTS: We observed 156 QTLs, 28 pairs of loci with epistatic interaction, and 10 significant QTL × environment interactions in the inbred and hybrid mapping populations. The high heterosis in F1 and IF2 populations for kernel weight per ear (KWPE), ear weight per ear (EWPE), and kernel number per row (KNPR) matched the high percentages of QTLs (over 50%) for those traits exhibiting overdominance, whereas a notable predominance of loci with dominance effects (more than 70%) was observed for traits that show low heterosis such as cob weight per ear (CWPE), rate of kernel production (RKP), ear length (EL), ear diameter (ED), cob diameter, and row number (RN). The environmentally stable QTL qRKP3-2 was identified across two mapping populations, while qKWPE9, affecting the trait mean and the mid-parent heterosis (MPH) level, explained over 18% of phenotypic variations. Nine QTLs, qEWPE9-1, qEWPE10-1, qCWPE6, qEL8, qED2-2, qRN10-1, qKWPE9, qKWPE10-1, and qRKP4-3, accounted for over 10% of phenotypic variation. In addition, QTL mapping identified 95 QTLs that were gathered together and integrated into 33 QTL clusters on 10 chromosomes. CONCLUSIONS: The results revealed that (1) the inheritance of yield-related traits and MPH in the heterotic pattern improved Reid (PA) × Tem-tropic I (PB) is trait-dependent; (2) a large proportion of loci showed dominance effects, whereas overdominance also contributed to MPH for KNPR, EWPE, and KWPE; (3) marker-assisted selection for markers at genomic regions 1.09-1.11, 2.04, 3.08-3.09, and 10.04-10.05 contributed to hybrid performance per se and heterosis and were repeatedly reported in previous studies using different heterotic patterns is recommended.

Dissecting Heterosis During the Ear Inflorescence Development Stage in Maize via a Metabolomics-based Analysis.

Heterosis can increase the yield of many crops and has been extensively applied in agriculture. In maize, female inflorescence architecture directly determines grain yield. Thus, exploring the relationship between early maize ear inflorescence development and heterosis regarding yield-related traits may be helpful for characterizing the molecular mechanisms underlying heterotic performance. In this study, we fine mapped the overdominant heterotic locus (hlEW2b), associated with ear width, in an approximately 1.98-Mb region based on analyses of chromosome segment substitution lines and the corresponding testcross population. Maize ear inflorescences at the floral meristem stage were collected from two inbred lines, one chromosome segment substitution line that carried hlEW2b (sub-CSSL16), the receptor parent lx9801, and the Zheng58 × sub-CSSL16 and Zheng58 × lx9801 hybrid lines. A total of 256 metabolites were identified, including 31 and 24 metabolites that were differentially accumulated between the two hybrid lines and between the two inbred lines, respectively. Most of these metabolites are involved in complex regulatory mechanisms important for maize ear development. For example, nucleotides are basic metabolites affecting cell composition and carbohydrate synthesis. Additionally, nicotinate and nicotinamide metabolism is important for photosynthesis, plant stress responses, and cell expansion. Moreover, flavonoid and phenolic metabolites regulate auxin transport and cell apoptosis. Meanwhile, phytohormone biosynthesis and distribution influence the cell cycle and cell proliferation. Our results revealed that changes in metabolite contents may affect the heterotic performance related to ear width and yield in maize hybrid lines. This study provides new clues in heterosis at the metabolomics level and implies that differentially accumulated metabolites made distinct contributions to the heterosis at an early stage of ear inflorescences development.

The miR164-dependent regulatory pathway in developing maize seed.

MicroRNA164 (miR164) plays a key role in leaf and flower development, lateral root initiation, and stress responses. However, little is known about the regulatory roles of miR164 during seed development, particularly in maize. The aim of this study was to discover the developmental function of miR164 in maize seed. Small RNA sequencing (sRNA-seq) was performed at two key stages. The results indicated that miR164 was down-regulated during maize seed development. In addition, degradome library sequencing and transient expression assays identified the target genes for miR164. Two microRNA (miRNA) pairs, miR164-NAM, ATAF, and CUC32 (NAC32) and miR164-NAC40, were isolated. The developmental function of miR164 was determined by analyzing the differentially expressed genes (DEGs) between the wild-type and miR164 transgenic lines using RNA sequencing (RNA-seq) and by screening the DEGs related to NAC32 and NAC40 via co-expression and transient expression analysis. These results identified two beta-expansin genes, EXPB14 and EXPB15, which were located downstream of the NAC32 and NAC40 genes. This study revealed, for the first time, a miR164-dependent regulatory pathway, miR164-NAC32/NAC40-EXPB14/EXPB15, which participates in maize seed expansion. These findings highlight the significance of miR164 in maize seed development, and can be used to explore the role of miRNA in seed development.

Heterotic loci identified for maize kernel traits in two chromosome segment substitution line test populations.

Heterosis has been widely used to increase grain quality and yield, but its genetic mechanism remains unclear. In this study, the genetic basis of heterosis for four maize kernel traits was examined in two test populations constructed using a set of 184 chromosome segment substitution lines (CSSLs) and two inbred lines (Zheng58 and Xun9058) in two environments. 63 and 57 different heterotic loci (HL) were identified for four kernel traits in the CSSLs × Zheng58 and CSSLs × Xun9058 populations, respectively. Of these, nine HL and six HL were identified for four kernel traits in the CSSLs × Zheng58 and CSSLs × Xun9058 populations, at the two locations simultaneously. Comparative analysis of the HL for the four kernel traits identified only 21 HL in the two test populations simultaneously. These results showed that most HL for the four kernel traits differed between the two test populations. The common HL were important loci from the Reid × Tangsipingtou heterotic model, and could be used to predict hybrid performance in maize breeding.

Comparative mapping of quantitative trait loci for tassel-related traits of maize in F2:3 and RIL populations.

Tassel architecture is an important trait in maize breeding and hybrid seed production. In this study, we investigated total tassel length (TTL) and tassel branch number (TBN) in 266 F2:3 families across six environments and in 301 recombinant inbred lines (RILs) across three environments, where all the plants were derived from a cross between 08-641 and Ye478. We compared the genetic architecture of the two traits across two generations through combined analysis. In total, 27 quantitative trait loci (QTLs) (15 in F2:3; 16 in RIL), two QTL × environment interactions (both in F2:3), 11 pairs of epistatic interactions (seven in F2:3; four in RIL) and four stable QTLs in both the F2:3 and RILs were detected. The RIL population had higher detection power than the F2:3 population. Nevertheless, QTL × environment interactions and epistatic interactions could be more easily detected in the F2:3 population than in the RILs. Overall, the QTL mapping results in the F2:3 and RILs were greatly influenced by genetic generations and environments. Finally, fine mapping for a novel and major QTL, qTTL-2-3 (bin 2.07), which accounted for over 8.49% of the phenotypic variation across different environments and generations, could be useful in marker-assisted breeding.

Identification of heterotic loci associated with grain yield and its components using two CSSL test populations in maize.

Heterosis has widely been used to increase grain yield and quality. In this study, the genetic basis of heterosis on grain yield and its main components in maize were examined over 2 years in two locations in two test populations constructed from a set of 184 chromosome segment substitution lines (CSSLs) and two inbred lines (Zheng58 and Xun9058). Of the 169 heterotic loci (HL) associated with grain yield and its five components identified in CSSL × Zheng58 and CSSL × Xun9058 test populations, only 25 HL were detected in both populations. The comparison of quantitative trait loci (QTLs) detected in the CSSL population with HL detected in the two test populations revealed that only 15.46% and 17.35% of the HL in the given populations respectively, shared the same chromosomal regions as that of the corresponding QTLs and showed dominant effects as well as pleiotropism with additive and dominant effects. In addition, most of the HL (74.23% and 74.49%) had overdominant effects. These results suggest that overdominance is the main contributor to the effects of heterosis on grain yield and its components in maize, and different HL are associated with heterosis for different traits in different hybrids.

Sucrose and ABA regulate starch biosynthesis in maize through a novel transcription factor, ZmEREB156.

Sucrose is not only the carbon source for starch synthesis, but also a signal molecule. Alone or in coordination with ABA, it can regulate the expression of genes involved in starch synthesis. To investigate the molecular mechanisms underlying this effect, maize endosperms were collected from Zea mays L. B73 inbred line 10 d after pollination and treated with sucrose, ABA, or sucrose plus ABA at 28 °C in the dark for 24 h. RNA-sequence analysis of the maize endosperm transcriptome revealed 47 candidate transcription factors among the differentially expressed genes. We therefore speculate that starch synthetic gene expression is regulated by transcription factors induced by the combination of sucrose and ABA. ZmEREB156, a candidate transcription factor, is induced by sucrose plus ABA and is involved in starch biosynthesis. The ZmEREB156-GFP-fused protein was localized in the nuclei of onion epidermal cells, and ZmEREB156 protein possessed strong transcriptional activation activity. Promoter activity of the starch-related genes Zmsh2 and ZmSSIIIa increased after overexpression of ZmEREB156 in maize endosperm. ZmEREB156 could bind to the ZmSSIIIa promoter but not the Zmsh2 promoter in a yeast one-hybrid system. Thus, ZmEREB156 positively modulates starch biosynthetic gene ZmSSIIIa via the synergistic effect of sucrose and ABA.