Mutation of the gene encoding the PHD-type transcription factor SAB23 confers submergence tolerance in rice.

Submergence is a major constraint on rice production in South and Southeast Asia. In this study, we determined that a gene of the Sub1A-binding protein family, SAB23, encodes a plant homeodomain (PHD)-type transcription factor that has a novel function of negatively regulating submergence tolerance in rice. The T-DNA insertion mutant sab23 displayed reduced plant height, delayed seed maturation, and lower percentage seed set. Importantly, this mutant also exhibited enhanced submergence tolerance. In addition, CRISPR/Cas9 knock out of SAB23 resulted in a significant reduction in the content of the gibberellin GA4 and a dramatic increase in the content of GA1 in the plants. SAB23 binds to the promoter of CYTOCHROME P450 714B2 (CYP714B2), which encodes a GA13-oxidase that catalyses the conversion of GA53 to GA19. Disruption of SAB23 function led to increased CYP714B2 transcription, and overexpression of CYP714B2 produced phenotypes similar to those of the SAB23-knockout plants. Taken together, our results reveal that SAB23 negatively regulates rice submergence tolerance by modulating CYP714B2 expression, which has significant potential for use in future breeding.

Ubiquitin E3 ligase AtCHYR2 functions in glucose regulation of germination and post-germinative growth in Arabidopsis thaliana.

KEY MESSAGE: Cytoplasm-localized RING ubiquitin E3 ligase AtCHYR2 involved in plant glucose responses during germination and post-germinative growth. CHY ZINC FINGER AND RING PROTEIN (CHYR) containing both a CHY zinc finger and a C3H2C3-type RING domain plays important roles in plant drought tolerance and the abscisic acid (ABA) response; however, their functions in sugar signaling pathways are less studied. Here, we report a glucose (Glc) response gene AtCHYR2, a homolog of RZFP34/CHYR1, which is induced by various abiotic stresses, ABA, and sugar treatments. In vitro, we demonstrated that AtCHYR2 is a cytoplasm-localized RING ubiquitin E3 ligase. Overexpression of AtCHYR2 led to hypersensitivity to Glc and enhanced Glc-mediated inhibition of cotyledon greening and post-germinative growth. Contrastingly, AtCHYR2 loss-of-function plants were insensitive to Glc-regulated seed germination and primary root growth, suggesting that AtCHYR2 is a positively regulator of the plant glucose response. Additionally, physiological analyses showed that overexpression AtCHYR2 increased stomata aperture and photosynthesis under normal condition, and promoted accumulation of endogenous soluble sugar and starch in response to high Glc. Genome-wide RNA sequencing analysis showed that AtCHYR2 affects a major proportion of Glc-responsive genes. Particularly, sugar marker gene expression analysis suggested that AtCHYR2 enhances the Glc response via a signaling pathway dependent on glucose metabolism. Taken together, our findings show that a novel RING ubiquitin E3 ligase, AtCHYR2, plays an important role in glucose responses in Arabidopsis.

GIF1 controls ear inflorescence architecture and floral development by regulating key genes in hormone biosynthesis and meristem determinacy in maize.

BACKGROUND: Inflorescence architecture and floral development in flowering plants are determined by genetic control of meristem identity, determinacy, and maintenance. The ear inflorescence meristem in maize (Zea mays) initiates short branch meristems called spikelet pair meristems, thus unlike the tassel inflorescence, the ears lack long branches. Maize growth-regulating factor (GRF)-interacting factor1 (GIF1) regulates branching and size of meristems in the tassel inflorescence by binding to Unbranched3. However, the regulatory pathway of gif1 in ear meristems is relatively unknown. RESULT: In this study, we found that loss-of-function gif1 mutants had highly branched ears, and these extra branches repeatedly produce more branches and florets with unfused carpels and an indeterminate floral apex. In addition, GIF1 interacted in vivo with nine GRFs, subunits of the SWI/SNF chromatin-remodeling complex, and hormone biosynthesis-related proteins. Furthermore, key meristem-determinacy gene RAMOSA2 (RA2) and CLAVATA signaling-related gene CLV3/ENDOSPERM SURROUNDING REGION (ESR) 4a (CLE4a) were directly bound and regulated by GIF1 in the ear inflorescence. CONCLUSIONS: Our findings suggest that GIF1 working together with GRFs recruits SWI/SNF chromatin-remodeling ATPases to influence DNA accessibility in the regions that contain genes involved in hormone biosynthesis, meristem identity and determinacy, thus driving the fate of axillary meristems and floral organ primordia in the ear-inflorescence of maize.

Identification of major QTL for waterlogging tolerance in maize using genome-wide association study and bulked sample analysis.

Waterlogging has increasingly become one of the major constraints to maize (Zea mays L.) production in some maize growing areas as it seriously decreases the yield. Waterlogging tolerance in maize germplasm provides a basis for maize waterlogging improvement. In this study, nine seedling traits, plant height (PH), root length (RL), shoot dry weight (SDW), root dry weight (RDW), adventitious root number (ARN), node number of brace root (BRNN), brace root number (BRN), brace root dry weigh (BRDW), survival rate (SR), and the secondary traits that were defined as relative phenotypic value of seedling traits under waterlogging and control treatments were used in a natural population that contain 365 inbred lines to evaluate the waterlogging tolerance of tropical maize. The result showed that maize waterlogging tolerance was genetically controlled and seedling traits were significantly different between the control and waterlogging treatments. PH, RL, SDW, and RDW are important seedling traits for waterlogging tolerance identification. Some tropical maize inbred lines were identified with extreme waterlogging tolerance that can provide an important germplasm resource for breeding. Population structure analysis showed that two major phylogenetic subgroups in tropical maize could be identified. Genome-wide association study (GWAS) using 39,266 single nucleotide polymorphisms (SNPs) across the whole genome identified 49 trait-SNPs distributed on over all 10 chromosomes excluding chromosome 10. Seventy-one significant SNPs, distributed on all 10 chromosomes excluding chromosome 5, were identified by extend bulked sample analysis (Ext-BSA) based on the inbred lines with extreme phenotypes. GWAS and Ext-BSA identified the same loci on bin1.07, bin6.01, bin2.09, bin6.04, bin7.02, and bin7.03. Nine genes were proposed as potential candidate genes. Cloning and functional validation of these genes would be helpful for understanding the molecular mechanism of waterlogging tolerance in maize.

A chlorophyll a oxygenase 1 gene ZmCAO1 contributes to grain yield and waterlogging tolerance in maize.

Chlorophylls function in photosynthesis, and are critical to plant developmental processes and responses to environmental stimuli. Chlorophyll b is synthesized from chlorophyll a by chlorophyll a oxygenase (CAO). Here, we characterize a yellow-green leaf (ygl) mutant and identify the causal gene which encodes a chlorophyll a oxygenase in maize (ZmCAO1). A 51 bp Popin transposon insertion in ZmCAO1 strongly disrupts its transcription. Low enzyme activity of ZmCAO1 leads to reduced concentrations of chlorophyll a and chlorophyll b, resulting in the yellow-green leaf phenotype of the ygl mutant. The net photosynthetic rate, stomatal conductance, and transpiration rate are decreased in the ygl mutant, while concentrations of δ-aminolevulinic acid (ALA), porphobilinogen (PBG) and protochlorophyllide (Pchlide) are increased. In addition, a ZmCAO1 mutation results in down-regulation of key photosynthetic genes, limits photosynthetic assimilation, and reduces plant height, ear size, kernel weight, and grain yield. Furthermore, the zmcao1 mutant shows enhanced reactive oxygen species production leading to sensitivity to waterlogging. These results demonstrate the pleiotropy of ZmCAO1 function in photosynthesis, grain yield, and waterlogging tolerance in maize.

Graphene oxide and indole-3-acetic acid cotreatment regulates the root growth of Brassica napus L. via multiple phytohormone pathways.

BACKGROUND: Studies have indicated that graphene oxide (GO) could regulated Brassica napus L. root growth via abscisic acid (ABA) and indole-3-acetic acid (IAA). To study the mechanism and interaction between GO and IAA further, B. napus L (Zhongshuang No. 9) seedlings were treated with GO and IAA accordance with a two factor completely randomized design. RESULTS: GO and IAA cotreatment significantly regulated the root length, number of adventitious roots, and contents of IAA, cytokinin (CTK) and ABA. Treatment with 25 mg/L GO alone or IAA (> 0.5 mg/L) inhibited root development. IAA cotreatment enhanced the inhibitory role of GO, and the inhibition was strengthened with increased in IAA concentration. GO treatments caused oxidative stress in the plants. The ABA and CTK contents decreased; however, the IAA and gibberellin (GA) contents first increased but then decreased with increasing IAA concentration when IAA was combined with GO compared with GO alone. The 9-cis-epoxycarotenoid dioxygenase (NCED) transcript level strongly increased when the plants were treated with GO. However, the NCED transcript level and ABA concentration gradually decreased with increasing IAA concentration under GO and IAA cotreatment. GO treatments decreased the transcript abundance of steroid 5-alpha-reductase (DET2) and isochorismate synthase 1 (ICS), which are associated with brassinolide (BR) and salicylic acid (SA) biosynthesis, but increased the transcript abundance of brassinosteroid insensitive 1-associated receptor kinase 1 (BAK1), cam-binding protein 60-like G (CBP60) and calmodulin binding protein-like protein 1, which are associated with BR and SA biosynthesis. Last, GO treatment increased the transcript abundance of 1-aminocyclopropane-1-carboxylic acid synthase 2 (ACS2), which is associated with the ethylene (ETH) pathway. CONCLUSIONS: Treatment with 25 mg/L GO or IAA (> 0.5 mg/L) inhibited root development. However, IAA and GO cotreatment enhanced the inhibitory role of GO, and this inhibition was strengthened with increased IAA concentration. IAA is a key factor in the response of B. napus L to GO and the responses of B. napus to GO and IAA cotreatment involved in multiple pathways, including those involving ABA, IAA, GA, CTK, BR, SA. Specifically, GO and IAA cotreatment affected the GA content in the modulation of B. napus root growth.

Cytoplasmic ribosomal protein L14B is essential for fertilization in Arabidopsis.

Plant cytoplasmic ribosomal proteins not only participate in protein synthesis, but also have specific roles in developmental regulation. However, the high heterogeneity of plant ribosome makes our understanding of these proteins very limited. Here we reported that RPL14B, a component of the ribosome large subunit, is critical for fertilization in Arabidopsis. RPL14B is existed in a majority of organs and tissues. No homozygous rpl14b mutant is available, indicating that RPL14B is irreplaceable for sexual reproduction. Smaller-sized rpl14b pollens could germinate normally, but pollen tube competitiveness is grievously weakened. Beside, cell fate specification is impaired in female gametophytes from heterozygous rpl14b/RPL14B ovules, resulting in defect of micropylar pollen tube attraction. However, this defect could be restored by restricted expression of RPL14B in synergid cells. Successful fertilization requires normal pollen tube growth and precise pollen tube guidance. Thus our results show a novel role of RPL14B in fertilization and shed new light on regulatory mechanism of pollen tube growth and precise pollen tube guidance.

Identification of a candidate gene underlying qKRN5b for kernel row number in Zea mays L.

KEY MESSAGE: A quantitative trait locus for kernel row number, qKRN5, was dissected into two tightly linked loci, qKRN5a and qKRN5b. Fine mapping, comparative analysis of nucleotide sequences and gene expression established the endonuclease/exonuclease/phosphatase family protein-encoding gene Zm00001d013603 as a causal gene of qKRN5b. Maize grain yield is determined by agronomically important traits that are controlled by interactions among and between genes and environmental factors. Considerable efforts have been made to identify major quantitative trait loci (QTLs) for yield-related traits; however, few were previously isolated and characterized in maize. In this study, we divided a QTL for kernel row number (KRN), qKRN5, into two tightly linked loci, qKRN5a and qKRN5b, using advanced backcross populations derived from near-isogenic lines. KRN was greater in individuals that were homozygous for the NX531 allele, which showed coupling-phase linkage. The major QTL qKRN5b had an additive effect of approximately one kernel row. Furthermore, fine mapping narrowed qKRN5b within a 147.2-kb region. The upstream sequence Zm00001d013603 and its expression in the ear inflorescence showed obvious differences between qKRN5b near-isogenic lines. In situ hybridization located Zm00001d013603 on the primordia of the spikelet pair meristems and spikelet meristems, but not in the inflorescence meristem, which indicates a role in regulating the initiation of reproductive axillary meristems of ear inflorescences. Expression analysis and nucleotide sequence alignment revealed that Zm00001d013603, which encodes an endonuclease/exonuclease/phosphatase family protein that hydrolyzes phosphatidyl inositol diphosphates, is the causal gene of qKRN5b. These results provide insight into the genetic basis of KRN and have potential value for enhancing maize grain yield.

Identification of Maize CC-Type Glutaredoxins That Are Associated with Response to Drought Stress.

Global maize cultivation is often adversely affected by drought stress. The CC-type glutaredoxin (GRX) genes form a plant-specific subfamily that regulate plant growth and respond to environmental stresses. However, how maize CC-type GRX (ZmGRXCC) genes respond to drought stress remains unclear. We performed a TBLASTN search to identify ZmGRXCCs in the maize genome and verified the identified sequences using the NCBI conservative domain database (CDD). We further established a phylogenetic tree using Mega7 and surveyed known cis-elements in the promoters of ZmGRXCCs using the PlantCARE database. We found twenty-one ZmGRXCCs in the maize genome by a genome-wide investigation and compared their phylogenetic relationships with rice, maize, and Arabidopsis. The analysis of their redox active sites showed that most of the 21 ZmGRXCCs share similar structures with their homologs. We assessed their expression at young seedlings and adult leaves under drought stress and their expression profiles in 15 tissues, and found that they were differentially expressed, indicating that different ZmGRXCC genes have different functions. Notably, ZmGRXCC14 is up-regulated at seedling, V12, V14, V16, and R1 stages. Importantly, significant associations between genetic variation in ZmGRXCC14 and drought tolerance are found at the seedling stage. These results will help to advance the study of the function of ZmGRXCCs genes under drought stress and understand the mechanism of drought resistance in maize.

Genome-Wide Analysis of TCP Family Genes in Zea mays L. Identified a Role for ZmTCP42 in Drought Tolerance.

The Teosinte-branched 1/Cycloidea/Proliferating (TCP) plant-specific transcription factors (TFs) have been demonstrated to play a fundamental role in plant development and organ patterning. However, it remains unknown whether or not the TCP gene family plays a role in conferring a tolerance to drought stress in maize, which is a major constraint to maize production. In this study, we identified 46 ZmTCP genes in the maize genome and systematically analyzed their phylogenetic relationships and synteny with rice, sorghum, and Arabidopsis TCP genes. Expression analysis of the 46 ZmTCP genes in different tissues and under drought conditions, suggests their involvement in maize response to drought stress. Importantly, genetic variations in ZmTCP32 and ZmTCP42 are significantly associated with drought tolerance at the seedling stage. RT-qPCR results suggest that ZmTCP32 and ZmTCP42 RNA levels are both induced by ABA, drought, and polyethylene glycol treatments. Based on the significant association between the genetic variation of ZmTCP42 and drought tolerance, and the inducible expression of ZmTCP42 by drought stress, we selected ZmTCP42, to investigate its function in drought response. We found that overexpression of ZmTCP42 in Arabidopsis led to a hypersensitivity to ABA in seed germination and enhanced drought tolerance, validating its function in drought tolerance. These results suggested that ZmTCP42 functions as an important TCP TF in maize, which plays a positive role in drought tolerance.

Identification of AtHsp90.6 involved in early embryogenesis and its structure prediction by molecular dynamics simulations.

Heat-shock protein of 90 kDa (Hsp90) is a key molecular chaperone involved in folding the synthesized protein and controlling protein quality. Conformational dynamics coupled to ATPase activity in N-terminal domain is essential for Hsp90's function. However, the relevant process is still largely unknown in plant Hsp90s, especially those required for plant embryogenesis which is inextricably tied up with human survival. Here, AtHsp90.6, a member of Hsp90 family in Arabidopsis, was firstly identified as a protein essential for embryogenesis. Thus we modelled AtHsp90.6 in its functionally closed 'lid-down' and open 'lid-up' states, exploring the nucleotide binding mechanism in these two states. Free energy landscape and electrostatic potential analysis revealed the switching mechanism between these two states. Collectively, this study quantitatively analysed the conformational changes of AtHsp90.6 bound to ATP or ADP. This result may help us understand the mechanism of action of AtHsp90.6 in future.

Genome-wide analysis of maize OSCA family members and their involvement in drought stress.

BACKGROUND: Worldwide cultivation of maize is often impacted negatively by drought stress. Hyperosmolality-gated calcium-permeable channels (OSCA) have been characterized as osmosensors in Arabidopsis. However, the involvement of members of the maize OSCA (ZmOSCA) gene family in response to drought stress is unknown. It is furthermore unclear which ZmOSCA gene plays a major role in genetic improvement of drought tolerance in Maize. METHODS: We predicted the protein domain structure and transmembrane regions by using the NCBI Conserved Domain Database database and TMHMM server separately. The phylogeny tree was built by Mega7. We used the mixed linear model in TASSEL to perform the family-based association analysis. RESULTS: In this report, 12 ZmOSCA genes were uncovered in the maize genome by a genome-wide survey and analyzed systematically to reveal their synteny and phylogenetic relationship with the genomes of rice, maize, and sorghum. These analyses indicated a relatively conserved evolutionary history of the ZmOSCA gene family. Protein domain and transmembrane analysis indicated that most of the 12 ZmOSCAs shared similar structures with their homologs. The result of differential expression analysis under drought at various stages, as well as the expression profiles in 15 tissues, revealed a functional divergence of ZmOSCA genes. Notably, the expression level of ZmOSCA4.1 being up-regulated in both seedlings and adult leaves. Notably, the association analysis between genetic variations in these genes and drought tolerance was detected. Significant associations between genetic variation in ZmOSCA4.1 and drought tolerance were found at the seedling stage. Our report provides a detailed analysis of the ZmOSCAs in the maize genome. These findings will contribute to future studies on the functional characterization of ZmOSCA proteins in response to water deficit stress, as well as understanding the mechanism of genetic variation in drought tolerance in maize.

Drought stress responses in maize are diminished by Piriformospora indica.

As an endophytic fungus of Sebacinales, Piriformospora indica promotes plant growth and resistance to abiotic stress, including drought. Colonization of maize roots promoted the leaf size, root length and number of tap roots. Under drought stress, the maize seedlings profited from the presence of the fungus and performed visibly better than the uncolonized controls. To identify genes and biological processes involved in growth promotion and drought tolerance conferred by P. indica, the root transcriptome of colonized and uncolonized seedlings was analyzed 0, 6 and 12 h after drought stress (20% polyethylene glycol 6000). The number of P. indica-responsive genes increased from 464 (no stress at 0 h) to 1337 (6 h drought) and 2037 (12 h drought). Gene Ontology analyses showed that the carbon and sulfur metabolisms are major targets of the fungus. Furthermore, the growth promoting effect of P. indica is reflected by higher transcript levels for microtubule associated processes. Under drought stress, the fungus improved the oxidative potential of the roots, and stimulated genes for hormone functions, including those which respond to abscisic acid, auxin, salicylic acid and cytokinins. The comparative analyses of our study provides systematic insight into the molecular mechanism how P. indica promotes plant performance under drought stress, and presents a collection of genes which are specifically targeted by the fungus under drought stress in maize roots.

Bulked Segregant RNA-seq Reveals Differential Expression and SNPs of Candidate Genes Associated with Waterlogging Tolerance in Maize.

Waterlogging has increasingly become one of the major constraints to maize productivity in some maize production zones because it causes serious yield loss. Bulked segregant RNA-seq (BSR-seq) has been widely applied to profile candidate genes and map associated Single Nucleotide Polymorphism (SNP) markers in many species. In this study, 10 waterlogging sensitive and eight tolerant inbred lines were selected from 60 maize inbred lines with waterlogging response determined and preselected by the International Maize and Wheat Improvement Center (CIMMYT) from over 400 tropical maize inbred lines. BSR-seq was performed to identify differentially expressed genes and SNPs associated with waterlogging tolerance. Upon waterlogging stress, 354 and 1094 genes were differentially expressed in the tolerant and sensitive pools, respectively, compared to untreated controls. When tolerant and sensitive pools were compared, 593 genes were differentially expressed under untreated and 431 genes under waterlogged conditions, of which 122 genes overlapped. To validate the BSR-seq results, the expression levels of six genes were determined by qRT-PCR. The qRT-PCR results were consistent with BSR-seq results. Comparison of allelic polymorphism in mRNA sequences between tolerant and sensitive pools revealed 165 (normal condition) and 128 (waterlogged condition) high-probability SNPs. We found 18 overlapping SNPs with genomic positions mapped. Eighteen SNPs were contained in 18 genes, and eight and nine of 18 genes were responsive to waterlogging stress in tolerant and sensitive lines, respectively. Six alleles of the 18 originated from tolerant pool were significantly up-regulated under waterlogging, but not those from sensitive pool. Importantly, one allele (GRMZM2G055704) of the six genes was mapped between umc1619 and umc1948 on chromosome 1 where a QTL associated with waterlogging tolerance was identified in a previous research, strongly indicating that GRMZM2G055704 is a candidate gene responsive to waterlogging. Our research contributes to the knowledge of the molecular mechanism for waterlogging tolerance in maize.

Natural variations in stearoyl-acp desaturase genes affect the conversion of stearic to oleic acid in maize kernel.

KEY MESSAGE: We identified 11 SAD genes, and mined their natural variations associated with the conservation of stearic to oleic acid, especially ZmSAD1 supported by both the QTL and an expression QTL. Maize oil is generally regarded as a healthy vegetable oil owing to its low abundance of saturated fatty acids. Stearoyl-ACP desaturase (SAD) is a key rate-limiting enzyme for the conservation of stearic (C18:0) to oleic (C18:1) acid. Here, 11 maize SAD genes were identified to have more divergent functions than Arabidopsis SAD genes. The genomic regional associations in a maize panel including 508 inbred lines identified 6 SAD genes significantly associated (P < 0.01) with the C18:0/C18:1 ratio or the level of C18:0 or C18:1, one gene of which co-localized with a quantitative trait locus (QTL) and 5 of which co-localized with an expression QTL. ZmSAD1, supported by both the QTL and an expression QTL, had the largest effect on C18:0/C18:1. One nonsynonymous single-nucleotide polymorphism in exon 3 and one 5-bp insertion/deletion in the 3' untranslated region were further shown to contribute to the natural variation in C18:0/C18:1 according to ZmSAD1-based association mapping. Finally, selection tests of ZmSAD1 in teosinte, regular maize, and high-oil maize indicated that ZmSAD1 was not a selection target during the process of maize domestication and high-oil maize development. These results will guide the manipulation of the ratio between saturated and unsaturated fatty acids in maize.

Modification of the fatty acid composition in Arabidopsis and maize seeds using a stearoyl-acyl carrier protein desaturase-1 (ZmSAD1) gene.

BACKGROUND: Stearoyl-acyl carrier protein desaturase (SAD) is a key enzyme that catalyses the conversion of stearoyl-acyl carrier protein (ACP) to oleoyl-ACP, a precursor for the biosynthesis of polyunsaturated fatty acids. ZmSAD1 (GenBank: KU949326) is a major QTL for stearic acid content in maize seeds. To investigate the biological function and the application potential of maize ZmSAD1 in oil biosynthesis, we isolated the full-length ZmSAD1 cDNA from maize B73 and overexpressed it in Arabidopsis and maize. RESULTS: Under seed-specific overexpression of ZmSAD1 in Arabidopsis, the stearic acid content and the ratio of saturated to unsaturated fatty acids in the seeds were significantly decreased relative to those in the control. Conversely, in transgenic ZmSAD1 RNAi Arabidopsis seeds, the contents of stearic acid and long-chain saturated acids and the ratio of saturated to unsaturated fatty acids were significantly increased; in addition, the oleic acid content was significantly decreased. More importantly, transgenic ZmSAD1 maize that expressed high levels of ZmSAD1 in its mature seeds showed reduced stearic acid content (1.57 %) and a lower saturated to unsaturated fatty acid ratio (20.40 %) relative to those (1.64 % and 20.61 %, respectively) of the control. Conversely, down-regulation of ZmSAD1 in maize resulted in increased levels of stearic acid (1.78 %), long-chain saturated acids (0.85 %) and the ratio of saturated to unsaturated fatty acids (21.54 %) relative to those (1.64 %, 0.74 %, and 20.61 %, respectively) of the control, whereas the oleic acid (32.01 %) level was significantly decreased relative to that (32.68 %) of the control. CONCLUSIONS: Our work demonstrates that the contents of stearic acid, oleic acid, and long-chain saturated acids, and the ratio of saturated to unsaturated fatty acids, are modified in maize seeds by seed-specific overexpression or down-regulation of ZmSAD1. Therefore, the ZmSAD1 gene is a useful tool for engineering the seed oil composition in maize and other crops.

Overexpression of Vitreoscilla hemoglobin increases waterlogging tolerance in Arabidopsis and maize.

BACKGROUND: Vitreoscilla hemoglobin (VHb) is a type of hemoglobin found in the Gram-negative aerobic bacterium Vitreoscilla that has been shown to contribute to the tolerance of anaerobic stress in multiple plant species. Maize (Zea mays L.) is susceptible to waterlogging, causing significant yield loss. In this study, we approached this problem with the introduction of an exogenous VHb gene. RESULTS: We overexpressed the VHb gene in Arabidopsis and maize under the control of the CaMV35S promoter. After 14 days of waterlogging treatment, the transgenic VHb Arabidopsis plants remained green, while the controls died. Under waterlogging, important plant growth traits of VHb plants, including seedling height, primary root length, lateral root number, and shoot dry weight were significantly improved relative to those of the controls. The VHb gene was also introduced into a maize line through particle bombardment and was then transferred to two elite maize inbred lines through marker-assisted backcrossing. The introduction of VHb significantly enhanced plant growth under waterlogging stress on traits, including seedling height, primary root length, lateral root number, root dry weight, and shoot dry weight, in both Zheng58 and CML50 maize backgrounds. Under the waterlogging condition, transgenic VHb maize seedlings exhibited elevated expression of alcohol dehydrogenase (ADH1) and higher peroxidase (POD) enzyme activity. The two VHb-containing lines, Zheng58 (VHb) and CML50 (VHb), exhibited higher tolerance to waterlogging than their negative control lines (Zheng58 and CML50). CONCLUSIONS: These results demonstrate that the exogenous VHb gene confers waterlogging tolerance to the transgenic maize line. In Maize in the place of to the transgenic maize line, the VHb gene is a useful molecular tool for the improvement of waterlogging and submergence-tolerance.

Isolation and functional characterization of a waterlogging-induced promoter from maize.

Using thermal asymmetric interlaced polymerase chain reaction (TAIL PCR), promoter sequences of 1,444 bp (HM241145) and 1,249 bp (HM241146) from the Zea mays zinc finger (zmzf) protein gene were isolated from the maize inbred lines Mo17 and Hz32, respectively. Sequence analysis demonstrated that the Mo17 zmzf promoter contained multiple cis-regulatory elements responding to anaerobic conditions. These included two GC-motifs, five anaerobic response elements (AREs), one GT-motif, and four G-boxes. Sequence alignment revealed that there was a 195 bp DNA deletion (-258 to -452 bp) in the Hz32 zmzf promoter compared with the Mo17 promoter. The deleted fragment contained three AREs. According to quantitative real time PCR analysis, the expression of the uidA gene driven by the Mo17 zmzf promoter increased after day 1 waterlogging treatment, peaked after day 2 and decreased at days 4-8. Staining for ß-glucuronidase (GUS) was observed in the roots of Mo17 zmzf transgenic lines under waterlogged conditions, but not in the leaves. GUS expression was not observed in the roots and leaves of Hz32 zmzf transgenic lines, even under submerged conditions, indicating that the Hz32 zmzf promoter was non-functional because of the deletion of three AREs. We propose that the Mo17 zmzf promoter was inducible by waterlogging, highly active, and root-specific, and might be useful for the genetic engineering of waterlogging tolerance.

Time course of expression of chalcone synthase gene in Ginkgo biloba.

Chalcone synthase (CHS) catalyses the first and key regulatory step of flavonoid biosynthetic pathway. A chalcone synthase gene was isolated from Ginkgo biloba leaves using the method of rapid amplification of the cDNA ends (RACE). The full-length cDNA, designated as GbCHS2, is 1,608 bp in length (GenBank accession No. DQ054841) and contains an open reading frame of 1,173 bp encoding a protein of 391 amino acids. Alignment of the predicted amino acid sequence of GbCHS2 has been shown to have high sequence similarity with GbCHS1. All the active sites and active site motifs in GbCHS1 protein were also found in GbCHS2. Correlation analysis between CHS activity and flavonoid accumulation during ginkgo leaf growth indicated that CHS might be the rate-limiting enzyme in the biosynthesis pathway of flavonoids in ginkgo leaves. Results of semi-quantitative RT-PCR analysis showed that flavonoid accumulation paralleled the transcription level of change in chs gene, suggesting chs gene as the specific key gene regulating flavonoid accumulation in ginkgo leaves.