<i>DWARF</i> and <i>SMALL SEED1</i>, a Novel Allele of <i>OsDWARF</i>, Controls Rice Plant Architecture,Seed Size,and Chlorophyll Biosynthesis

Plant architecture is a vital agronomic trait to control yield in rice (Oryza sativa L.). A dwarf and small seed 1 (dss1) mutant were obtained from the ethyl methanesulfonate (EMS) mutagenized progeny of a Guizhou glutinous landrace cultivar, Lipingzabianhe. The dss1 mutant displayed phenotypes similar to those of brassinosteroid (BR) deficient mutants, such as dwarfing, dark green and rugose erect leaves, small seeds, and loner neck internode panicles with primary branching. In our previous study, the underlying DSS1 gene was isolated, a novel allele of OsDWARF (OsBR6ox) that encodes a cytochrome P450 protein involved in the BR biosynthetic pathway by MutMap technology. In this work, we confirmed that a Thr335Ile amino acid substitution residing in DSS1/OsDWARF was responsible for the dwarf, panicle architecture, and small seed phenotypes in the dss1 mutants by genetic transformation experiments. The overexpression of OsDWARF in the dss1 mutant background could not only recover dss1 to the normal plant height and panicle architecture but also rescued normal leaf angles, seed size, and leaf color. Thus, the specific mutation in DSS1/OsDWARF influenced plant architecture, seed size, and chlorophyll biosynthesis.     

First Report of <i>Fusarium striatum</i> Causing Root Rot Disease of <i>Panax notoginseng</i> in Yunnan,China

Panax notoginseng is a traditional Chinese medicinal plant. Root rot of P. notoginseng is one of the most serious diseases affecting P. notoginseng growth and causes wilted leaves, fewer lateral roots and rotten roots. Root rot is a soil-borne disease, and mainly occurs from June to August in Yunnan Province when the temperatures are high and the air is humid. In this study, the endophytic fungal genus Fusarium isolate E-2018.1.22-#3.2 was obtained from a P. notoginseng embryo. Fusarium isolate E-2018.1.22-#3.2 was identified as Fusarium striatum based on morphological characteristics and molecular analysis. The fungus was found to have conidiophores and macroconidia, and its ITS, LSU and TEF-1α genes shared 100%, 99.2% and 99% identities with the homologous genes of Fusarium striatum, respectively. Isolate F. striatum E-2018.1.22-#3.2 can cause root rot symptoms, including black, soft roots, fewer lateral roots and leaf wilt, in 93% of the experimental P. notoginseng plants, and could be re-isolated, fulfilling Koch’s postulates. When the P. notoginseng plants were treated with the fungicide pyraclostrobin, isolate F. striatum E-2018.1.22-#3.2 was unable to cause root rot. We have therefore demonstrated that F. striatum E-2018.1.22-#3.2 is able to cause root rot disease in P. notoginseng. This is the first report of root rot disease caused by F. striatum on P. notoginseng in China.     

Cloning and Characterization of <i>EuGID1</i> in <i>Eucommia ulmoides</i> Oliver

Gibberellic acid controlled the key developmental processes of the life cycle of landing plants, and regulated the growth and development of plants. In this study, a novel gibberellin receptor gene EuGID1 was obtained from Eucommia ulmoides Oliver. The cDNA of EuGID1 was 1556 bp, and the open reading frame was 1029 bp, which encoded 343 amino acids. EuGID1 had the homology sequence with the hormone-sensitive lipase family. Amino acid sequence alignment confirmed EuGID1 protein had the highest homology with the GID1 protein of Manihot esculenta. EuGID1 was located in the nucleus and cell membrane and had expression in four plant organs. Overexpression of EuGID1 in transgenic Arabidopsis plants promoted plant elongation and increased siliques yield.     

Identification and Genetic Analysis of a Novel Allelic Variation of <i>Brittle-1</i> with Endosperm Mutant in Maize

Endosperm mutants are critical to the studies on both starch synthesis and metabolism and genetic improvement of starch quality in maize. In the present study, a novel maize endosperm mutant A0178 of natural variation was used as the experimental material and identified and then characterized. Through phenotypic identification, genetic analysis, main ingredients measurement and embryo rescue, development of genetic mapping population from A0178, the endosperm mutant gene was located. The results showed that the mutant exhibited extremely low germination ability as attributed to the inhibited embryo development, and amounts of sugars were accumulated in the mutant seeds and more sugars content was detected at 23 days after pollination (DAP) in A0178 than B73. Employing genetic linkage analysis, the mutant trait was mapped in the bin 5.04 on chromosome 5. Sequence analysis showed that two sites of base transversion and insertion presented in the protein coding region and non-coding region of the mutant brittle-1 (bt1), the adenylate translocator encoding gene involved in the starch synthesis. The single base insertion in the coding region cause frameshift mutation, early termination and lose of function of Brittle-1 (BT1). All results suggested that bt1 is a novel allelic gene and the causal gene of this endosperm mutant, providing insights on the mechanism of endosperm formation in maize.     

Differential Responses of <i>NHX1</i> and <i>SOS1</i> Gene Expressions to Salinity in two <i>Miscanthus sinensis</i> Anderss. Accessions with Different Salt Tolerance

The lignocellulosic crop Miscanthus spp. has been identified as a good candidate for biomass production. The responses of Miscanthus sinensis Anderss. to salinity were studied to satisfy the needs for high yields in marginal areas and to avoid competition with food production. The results indicated that the relative advantages of the tolerant accession over the sensitive one under saline conditions were associated with restricted Na⁺ accumulation in shoots. Seedlings of two accessions (salt-tolerant ‘JM0119’ and salt-sensitive ‘JM0099’) were subjected to 0 (control), 100, 200, and 300 mM NaCl stress to better understand the salt-induced biochemical responses of genes involved in Na⁺ accumulation in M. sinensis. The adaptation responses of genes encoding for Na⁺ /H⁺ antiporters, NHX1 and SOS1 to NaCl stress were examined in JM0119 and JM0099.The cDNA sequences of genes examined were highly conserved among the relatives of M. sinensis based on the sequencing on approximate 600 bp-long cDNA fragments obtained from degenerate PCR. These salt-induced variations of gene expression investigated by quantitative real-time PCR provided evidences for insights of the molecular mechanisms of salt tolerance in M. sinensis. The expression of NHX1 was up-regulated by salt stress in JM0119 shoot and root tissues. However, it was hardly affected in JM0099 shoot tissue except for a significant increase at the 100 mM salt treatment, and it was salt-suppressed in the JM0099 root tissue. In the root tissue, the expression of SOS1 was induced by the high salt treatment in JM0119 but repressed by all salt treatments in JM0099. Thus, the remarkably higher expression of NHX1 and SOS1 were associated with the resistance to Na⁺ toxicity by regulation of the Na⁺ influx, efflux, and sequestration under different salt conditions.     

Effects of <i>Piriformospora indica</i> on the Respiration of <i>Taxus chinensis</i> var. <i>mairei</i> under Water Stress

Seedlings of Taxus chinensis var. mairei were used as experimental materials to study the adaptation of Piriformospora indica to this plant under water stress. The materials were divided into two groups, namely, with or without inoculation with P. indica. Each group was subjected to four different levels of water stress. Vitality and physiological and biochemical indexes of the roots of T. chinensis var. mairei were regularly measured. Under water stress, T. chinensis var. mairei had significantly decreased root vitality; root vitality was higher in inoculated roots than in uninoculated roots. Under intense water stress, the inoculated roots had a higher soluble sugar content than the uninoculated roots. Under water stress, T. chinensis var. mairei experienced decreased activity of aerobic respiratory metabolic enzymes. The activity of anaerobic respiratory metabolic enzymes and alcohol dehydrogenase initially increased and then decreased, whereas that of lactate dehydrogenase increased. The inoculated roots had a higher activity of respiratory metabolic enzymes than the uninoculated roots. As water stress was further intensified, the roots had significantly decreased activity of aerobic respiratory metabolic enzymes and significantly increased activity of anaerobic respiratory metabolic enzymes. The activity of respiratory metabolic enzymes decreased faster in the uninoculated roots than in the inoculated roots. This study demonstrated that Piriformospora indica plays a positive role in enhancing the antihypoxic ability of T. chinensis var. mairei, thereby alleviating plant damage due to water stress.     

Identification and Characterization of a Novel Yellow Leaf Mutant <i>yl1</i> in Rice

Leaf-color mutants play an important role in the study of chlorophyll metabolism, chloroplast development, and photosynthesis system. In this study, the yellow leaf 1 (yl1) rice mutant was identified from the ethyl methane sulfonate-treated mutant progeny of Lailong, a glutinous japonica rice landrace cultivated in Guizhou Province, China. Results showed that yl1 exhibited yellow leaves with decreased chlorophyll content throughout the growth period. Chloroplast development in the yl1 mutant was disrupted, and the grana lamellae was loosely packed and disordered. RNA sequencing and real-time quantitative polymerase chain reaction (qRT-PCR) analysis revealed that the chlorophyll synthesis-related genes OsCHLH, OsCHLM, OsCHLG, PORB, and YGL8, as well as the chloroplast development-related genes FtsZ, OsRpoTp, and RbcL, were down-regulated in the yl1 mutant. Genetic analysis revealed that the yellow leaf phenotype of yl1 was controlled by recessive nuclear gene. By employing the MutMap method, the mutation responsible for the phenotype was mapped to a 6.17 Mb region between 17.34 and 23.51 Mb on chromosome 3. Two non-synonymous single-nucleotide polymorphisms (SNPs) located in the gene locus LOC_Os03g31210 and LOC_Os03g36760 were detected in this region. The two SNPs were further confirmed by PCR and Sanger sequencing. The expression patterns of the two candidate genes indicated that LOC_Os03g36760 showed greater potential for functional verification. Subcellular protein localization revealed that the encoded product of LOC_Os03g36760 was localized in the nucleus, cytoplasm, and plasma membrane. These results will be useful for further characterization and cloning of the yl1 gene, and for research on the molecular mechanisms controlling biogenesis and chloroplast biochemical processes.     

Characterization and Candidate Gene Analysis of the Yellow-Green Leaf Mutant <i>ygl16</i> in Rice (<i>Oryza sativa</i> L.)

Leaf color mutants are ideal materials for studying many plant physiological and metabolic processes such as photosynthesis, photomorphogenesis, hormone physiology and disease resistance. In this study, the genetically stable yellow-green leaf mutant ygl16 was identified from mutated “Xinong 1B”. Compared with the wild type, the pigment concentration and photosynthetic capacity of the ygl16 decreased significantly. The ultrastructural observation showed that the distribution of thylakoid lamellae was irregular in ygl16 chloroplasts, and the grana and matrix lamellae were blurred and loose in varied degrees, and the chloroplast structure was disordered, while the osmiophilic corpuscles increased. The results of the genetic analysis and mapping showed that the phenotype of ygl16 was controlled by a pair of recessive nuclear gene. The gene located in the 56Kb interval between RM25654 and R3 on the long arm of chromosome 10. The sequencing results showed that the 121st base of the first intron of the candidate gene OsPORB/FGL changed from A to T in the interval. qRT-PCR results showed that the expression of chlorophyll synthase-related genes in the mutant decreased.     

Development of a New Cold-Tolerant Maize (<i>Zea mays</i> L.) Germplasm Using the <i>ICE1</i> Gene from <i>Arabidopsis thaliana</i>

To develop cold-tolerant maize germplasms and identify the activation of INDUCER OF CRT/DRE-BINDING FACTOR EXPRESSION (ICE1) expression in response to cold stress, RT-PCR was used to amplify the complete open reading frame sequence of the ICE1 gene and construct the plant expression vector pCAMBIA3301-ICE1-Bar. Immature maize embryos and calli were transformed with the recombinant vector using Agrobacterium tumefaciens-mediated transformations. From the regenerated plantlets, three T1 lines were screened and identified by PCR. A Southern blot analysis showed that a single copy of the ICE1 gene was integrated into the maize (Zea mays L.) genomes of the three T1 generations. Under low temperature-stress conditions (4°C), the relative conductivity levels decreased by 27.51%–31.44%, the proline concentrations increased by 12.50%–17.50%, the malondialdehyde concentrations decreased by 16.78%–18.37%, and the peroxidase activities increased by 19.60%–22.89% in the T1 lines compared with those of the control. A real-time quantitative PCR analysis showed that the ICE1 gene was ectopically expressed in the roots, stems, and leaves of the T1 lines. ICE1 positively regulates the expression of the CBF genes in response to cold stress. Thus, this study showed the successful transformation of maize with the ICE1 gene, resulting in the generation of a new maize germplasm that had increased tolerance to cold stress.     

<i>VvAGAMOUS</i> Affect Development of Four Different Grape Species Ovary

Grape pistil has an important influence on fruit size and quality. However, there were few studies on grape ovary, and the development process of the ovary is still unclear. Therefore, in this paper, four different grape varieties with different lengths of small inflorescences, namely ‘Musct Hambourg’ grape (Vitis vinifera), ‘Concord’ grape (Vitis labrusca), ‘ShanPuTao’ grape (Vitis amurensis) and ‘GongNiang2Hao’ grape (Vitis amurensis × Vitis vinifera) were used as test materials. Four varieties ovary were significant differences by means of stereomicroscope, paraffin section. The expression of ovary determining gene VvAGAMOUS (VvAG) and its development related genes VvCRABS CLAW (VvCRC) andVvAGAMOUS-LIKE 11 (VvAGL11) with similar functions during the development of different grape varieties were preliminarily explored using fluorescence quantitative test. The relationship between VvAG and VvCRC, VvAG and VvAGL11 were analyzed using Y1H assay. Our results showed that there were obvious abdominal sutures on the surface of expect for ‘Musct Hambourg’ grape, and existing poly carpels. The ovary development of ‘ShanPuTao’ and ‘GongNiang2Hao’ grape was completed when the inflorescence length was less than 1 cm, while the ‘Concord’ and ‘Musct Hambourg’ grape were fully developed when the length of inflorescence was 3–4 and 4–5 cm, respectively. VvAG and VvCRC began to express in large quantities after the formation of stamen primordia, while VvAGL11 during the forming of ovule primordia. Therefore, VvAG and VvCRC mainly regulated the development of stamens and carpels and also promote the development of ovules, while VvAGL11 major regulated the development of ovules. The promoters of VvCRC and VvAGL11 were bound by VvAG. This study provides an important theoretical basis for further research on the molecular mechanism of grape ovary development.     

Identification of a Novel <i>OsCYP2</i> Allele that Was Involved in Rice Response to Low Temperature Stress

Cyclophilin (CYP) plays an important role in plant response to stress, and OsCYP2, one gene of cyclophlilin family, is involved in auxin signal transduction and stress signaling in rice. However, the mechanism that OsCYP2 is involved in rice response to low temperature is still unclear. We identified a new OsCYP2 allelic mutant, lrl3, with fewer lateral roots, and the differences in shoot height, primary root length and adventitious root length increased with the growth process compared to the wild-type plant. Auxin signaling pathway was also affected and became insensitive to gravity. The transgenic rice plants with over-expression of OsCYP2 were more tolerant to low temperature than the wild-type plants, suggesting that OsCYP2 was involved in the low temperature response in rice. In addition, OsCYP2 negatively regulated the expression of OsTPS38, a terpene synthase gene, and was dependent on the OsCDPK7-mediated pathway in response to low temperature stress. OsTPS38- overexpressed transgenic line ox-2 was more sensitive to low temperature. Therefore, OsCYP2 may negatively regulate OsTPS38 through an OsCDPK7-dependent pathway to mediate the response to low temperature in rice. These results provide a new basis for auxin signaling genes to regulate rice response to low temperature stress.     

Phytosulfokine-α Promotes Root Growth by Repressing Expression of <i>Pectin Methylesterase Inhibitor</i> (<i>PMEI</i>) Genes in <i>Medicago truncatula</i>

Phytosulfokine-α (PSK-α), a sulfated pentapeptide with the sequence YIYTQ, is encoded by a small precursor gene family in Arabidopsis. PSK-α regulates multiple growth and developmental processes as a novel peptide hormone. Despite its importance, functions of PSK-α in M. truncatula growth remains unknown. In this study, we identified five genes to encode PSK-α precursors in M. truncatula. All of these precursors possess conserved PSK-α signature motif. Expression pattern analysis of these MtPSK genes revealed that each gene was expressed in a tissue-specific or ubiquitous pattern and three of them were remarkably expressed in root. Treatment of M. truncatula seedlings with synthetic PSK- α peptide significantly promoted root elongation. In addition, expression analysis of downstream genes by RNA-seq and qRT-PCR assays suggested that PSK-α signaling might regulate cell wall structure via PMEI-PME module to promote root cell growth. Taken together, our results shed light on the mechanism by which PSK-α promotes root growth in M. truncatula, providing a new resource for improvement of root growth in agriculture.     

DDG1 and G Protein α Subunit RGA1 Interaction Regulates Plant Height and Senescence in Rice (<i>Oryza sativa</i>)

Many studies have already shown that dwarfism and moderate delayed leaf senescence positively impact rice yield, but the underlying molecular mechanism of dwarfism and leaf senescence remains largely unknown. Here, using map-based cloning, we identified an allele of DEP2, DDG1, which controls plant height and leaf senescence in rice. The ddg1 mutant displayed dwarfism, short panicles, and delayed leaf senescence. Compared with the wild-type, ddg1 was insensitive to exogenous gibberellins (GA) and brassinolide (BR). DDG1 is expressed in various organs, especially in stems and panicles. Yeast two-hybrid assay, bimolecular fluorescent complementation and luciferase complementation image assay showed that DDG1 interacts with the α-subunit of the heterotrimeric G protein. Disruption of RGA1 resulted in dwarfism, short panicles, and darker-green leaves. Furthermore, we found that ddg1 and the RGA1 mutant was more sensitive to salt treatment, suggesting that DDG1 and RGA1 are involved in regulating salt stress response in rice. Our results show that DDG1/DEP2 regulates plant height and leaf senescence through interacting with RGA1.     

Phenotype Analysis and Fine Mapping of the Male Sterile Mutant <i>ms10</i> in Rice (<i>Oryza sativa</i> L.)

There is a positive correlation between fertility and yield, and the decrease of fertility is bound to a greatly reduced crop yield. Male sterile mutants can be used in hybrid rice. Therefore, rice male sterility has an important value in research and application, and the study of related mutants is also very vital. The mutant ms10 (male sterile 10) reported in this study was induced by ethyl methane sulfonate (EMS) in the indica maintainer line Xinong 1B. There was no significant difference between the ms10 and wild type in the vegetative growth stage. However, in the reproductive growth stage, ms10 showed that the plant became shorter, the anther became smaller and the color became lighter, and finally showed the phenotype of male sterility in comparison to the wild type. I₂-KI staining showed that the pollen was malformed and only a little was active. Scanning electron microscopy observation showed that the exine waxy layer of the ms10 anther decreased, suggesting that the protective effect on pollen was decreased. This may be one of the reasons leading to the phenotype of male sterility. Finally, the pollen showed shrinkage and collapsed, and the structure of germinating pore cover disappeared. This may be the result of sterility. Genetic analysis showed that the male sterility phenotype of the mutant was controlled by a single recessive nuclear gene. MS10 was mapped between the molecular markers IND37 and IND51 on chromosome 4, with a physical distance of 178.6 kb. These results lay the foundation for further studies on MS10.     

Participation of Auxin Transport in the Early Response of the <i>Arabidopsis</i> Root System to Inoculation with <i>Azospirillum brasilense</i>

The potential of Plant Growth Promoting Rhizobacteria (PGPR) has been demonstrated in the case of plant inoculation with bacteria of the genus Azospirillum which improves yield. A. brasilense produces a wide variety of molecules, including the natural auxin indole-3-acetic acid (IAA), as well as other phytoregulators. However, several studies have suggested that auxin induces changes in plant development during their interaction with the bacteria. The effects of A. brasilense Sp245 on the development of Arabidopsis thaliana root were investigated to help explain the molecular basis of the interaction. The results obtained showed a decrease in primary root length from the first day and remained so throughout the exposure, accompanied by a stimulation of initiation and maturation of lateral root primordia and an increase of lateral roots. An enhanced auxin response was evident in the vascular tissue and lateral root meristems of inoculated plants. However, after five days of bacterization, the response disappeared in the primary root meristems. The role of polar auxin transport (PAT) in auxins relocation involved the PGP1, AXR4-1, and BEN2 proteins, which apparently mediated A. brasilense-induced root branching of Arabidopsis seedlings.     

<i>OsMAPK6</i> Affects Male Fertility by Reducing Microspore Number and Delaying Tapetum Degradation in <i>Oryza sativa</i> L.

The mitogen-activated protein kinase (MAPK) cascade is important in stress signal transduction and plant development. In the present study, we identified a rice (Oryza sativa L.) mutant with reduced fertility, Oryza sativa mitogen-activated protein kinase 6 (osmapk6), which harbored a mutated MAPK gene. Scanning and transmission electron microscopy, quantitative RT-PCR analysis, TUNEL assays, RNA in situ hybridization, longitudinal and transverse histological sectioning, and map-based cloning were performed to characterize the osmapk6 mutant. The gene OsMAPK6 was expressed throughout the plant but predominantly in the microspore mother cells, tapetal cells, and microspores in the anther sac. Compared with the wild type, the total number of microspores was reduced in the osmapk6 mutant. The formation of microspore mother cells was reduced in the osmapk6 anther sac at an early stage of anther development, which was the primary reason for the decrease in the total number of microspores. Programmed cell death of some tapetal cells was delayed in osmapk6 anthers and affected exine formation in neighboring microspores. These results suggest that OsMAPK6 plays pivotal roles in microspore mother cell formation and tapetal cell degradation.