DREB1 and DREB2 Genes in Garlic (Allium sativum L.): Genome-Wide Identification, Characterization, and Stress Response.

Dehydration-responsive element-binding (DREB) transcription factors (TFs) of the A1 and A2 subfamilies involved in plant stress responses have not yet been reported in Allium species. In this study, we used bioinformatics and comparative transcriptomics to identify and characterize DREB A1 and A2 genes redundant in garlic (Allium sativum L.) and analyze their expression in A. sativum cultivars differing in the sensitivity to cold and Fusarium infection. Eight A1 (AsaDREB1.1-1.8) and eight A2 (AsaDREB2.1-2.8) genes were identified. AsaDREB1.1-1.8 genes located in tandem on chromosome 1 had similar expression patterns, suggesting functional redundancy. AsaDREB2.1-2.8 were scattered on different chromosomes and had organ- and genotype-specific expressions. AsaDREB1 and AsaDREB2 promoters contained 7 and 9 hormone- and stress-responsive cis-regulatory elements, respectively, and 13 sites associated with TF binding and plant development. In both Fusarium-resistant and -sensitive cultivars, fungal infection upregulated the AsaDREB1.1-1.5, 1.8, 2.2, 2.6, and 2.8 genes and downregulated AsaDREB2.5, but the magnitude of response depended on the infection susceptibility of the cultivar. Cold exposure strongly upregulated the AsaDREB1 genes, but downregulated most AsaDREB2 genes. Our results provide the foundation for further functional analysis of the DREB TFs in Allium crops and could contribute to the breeding of stress-tolerant varieties.

Genome-Wide Identification, Expression, and Response to Fusarium Infection of the SWEET Gene Family in Garlic (Allium sativum L.).

Proteins of the SWEET (Sugar Will Eventually be Exported Transporters) family play an important role in plant development, adaptation, and stress response by functioning as transmembrane uniporters of soluble sugars. However, the information on the SWEET family in the plants of the Allium genus, which includes many crop species, is lacking. In this study, we performed a genome-wide analysis of garlic (Allium sativum L.) and identified 27 genes putatively encoding clade I-IV SWEET proteins. The promoters of the A. sativum (As) SWEET genes contained hormone- and stress-sensitive elements associated with plant response to phytopathogens. AsSWEET genes had distinct expression patterns in garlic organs. The expression levels and dynamics of clade III AsSWEET3, AsSWEET9, and AsSWEET11 genes significantly differed between Fusarium-resistant and -susceptible garlic cultivars subjected to F. proliferatum infection, suggesting the role of these genes in the garlic defense against the pathogen. Our results provide insights into the role of SWEET sugar uniporters in A. sativum and may be useful for breeding Fusarium-resistant Allium cultivars.

ZmDREB2.9 Gene in Maize (Zea mays L.): Genome-Wide Identification, Characterization, Expression, and Stress Response.

Dehydration-responsive element-binding (DREB) transcription factors of the A2 subfamily play key roles in plant stress responses. In this study, we identified and characterized a new A2-type DREB gene, ZmDREB2.9, in the Zea mays cv. B73 genome and compared its expression profile with those of the known A2-type maize genes ZmDREB2.1-2.8. ZmDREB2.9 was mapped to chromosome 8, contained 18 predicted hormone- and stress-responsive cis-elements in the promoter, and had two splice isoforms: short ZmDREB2.9-S preferentially expressed in the leaves, embryos, and endosperm and long ZmDREB2.9-L expressed mostly in the male flowers, stamens, and ovaries. Phylogenetically, ZmDREB2.9 was closer to A. thaliana DREB2A than the other ZmDREB2 factors. ZmDREB2.9-S, ZmDREB2.2, and ZmDREB2.1/2A were upregulated in response to cold, drought, and abscisic acid and may play redundant roles in maize stress resistance. ZmDREB2.3, ZmDREB2.4, and ZmDREB2.6 were not expressed in seedlings and could be pseudogenes. ZmDREB2.7 and ZmDREB2.8 showed similar transcript accumulation in response to cold and abscisic acid and could be functionally redundant. Our results provide new data on Z. mays DREB2 factors, which can be used for further functional studies as well as in breeding programs to improve maize stress tolerance.

Thaumatin-like Protein (TLP) Genes in Garlic (Allium sativum L.): Genome-Wide Identification, Characterization, and Expression in Response to Fusarium proliferatum Infection.

Plant antifungal proteins include the pathogenesis-related (PR)-5 family of fungi- and other stress-responsive thaumatin-like proteins (TLPs). However, the information on the TLPs of garlic (Allium sativum L.), which is often infected with soil Fusarium fungi, is very limited. In the present study, we identified 32 TLP homologs in the A. sativum cv. Ershuizao genome, which may function in the defense against Fusarium attack. The promoters of A. sativumTLP (AsTLP) genes contained cis-acting elements associated with hormone signaling and response to various types of stress, including those caused by fungal pathogens and their elicitors. The expression of AsTLP genes in Fusarium-resistant and -susceptible garlic cultivars was differently regulated by F. proliferatum infection. Thus, in the roots the mRNA levels of AsTLP7-9 and 21 genes were increased in resistant and decreased in susceptible A. sativum cultivars, suggesting the involvement of these genes in the garlic response to F. proliferatum attack. Our results provide insights into the role of TLPs in garlic and may be useful for breeding programs to increase the resistance of Allium crops to Fusarium infections.

Pathogenesis-Related Genes of PR1, PR2, PR4, and PR5 Families Are Involved in the Response to Fusarium Infection in Garlic (Allium sativum L.).

Plants of the genus Allium developed a diversity of defense mechanisms against pathogenic fungi of the genus Fusarium, including transcriptional activation of pathogenesis-related (PR) genes. However, the information on the regulation of PR factors in garlic (Allium sativum L.) is limited. In the present study, we identified AsPR genes putatively encoding PR1, PR2, PR4, and PR5 proteins in A. sativum cv. Ershuizao, which may be involved in the defense against Fusarium infection. The promoters of the AsPR1-5 genes contained jasmonic acid-, salicylic acid-, gibberellin-, abscisic acid-, auxin-, ethylene-, and stress-responsive elements associated with the response to plant parasites. The expression of AsPR1c, d, g, k, AsPR2b, AsPR5a, c (in roots), and AsPR4a(c), b, and AsPR2c (in stems and cloves) significantly differed between garlic cultivars resistant and susceptible to Fusarium rot, suggesting that it could define the PR protein-mediated protection against Fusarium infection in garlic. Our results provide insights into the role of PR factors in A. sativum and may be useful for breeding programs to increase the resistance of Allium crops to Fusarium infections.

Nepenthes × ventrata Transcriptome Profiling Reveals a Similarity Between the Evolutionary Origins of Carnivorous Traps and Floral Organs.

The emergence of the carnivory syndrome and traps in plants is one of the most intriguing questions in evolutionary biology. In the present study, we addressed it by comparative transcriptomics analysis of leaves and leaf-derived pitcher traps from a predatory plant Nepenthes ventricosa × Nepenthes alata. Pitchers were collected at three stages of development and a total of 12 transcriptomes were sequenced and assembled de novo. In comparison with leaves, pitchers at all developmental stages were found to be highly enriched with upregulated genes involved in stress response, specification of shoot apical meristem, biosynthesis of sucrose, wax/cutin, anthocyanins, and alkaloids, genes encoding digestive enzymes (proteases and oligosaccharide hydrolases), and flowering-related MADS-box genes. At the same time, photosynthesis-related genes in pitchers were transcriptionally downregulated. As the MADS-box genes are thought to be associated with the origin of flower organs from leaves, we suggest that Nepenthes species could have employed a similar pathway involving highly conserved MADS-domain transcription factors to develop a novel structure, pitcher-like trap, for capture and digestion of animal prey during the evolutionary transition to carnivory. The data obtained should clarify the molecular mechanisms of trap initiation and development and may contribute to solving the problem of its emergence in plants.

Genome-Wide Identification and Expression of Chitinase Class I Genes in Garlic (Allium sativum L.) Cultivars Resistant and Susceptible to Fusarium proliferatum.

Vegetables of the Allium genus are prone to infection by Fusarium fungi. Chitinases of the GH19 family are pathogenesis-related proteins inhibiting fungal growth through the hydrolysis of cell wall chitin; however, the information on garlic (Allium sativum L.) chitinases is limited. In the present study, we identified seven class I chitinase genes, AsCHI1-7, in the A. sativum cv. Ershuizao genome, which may have a conserved function in the garlic defense against Fusarium attack. The AsCHI1-7 promoters contained jasmonic acid-, salicylic acid-, gibberellins-, abscisic acid-, auxin-, ethylene-, and stress-responsive elements associated with defense against pathogens. The expression of AsCHI2, AsCHI3, and AsCHI7 genes was constitutive in Fusarium-resistant and -susceptible garlic cultivars and was mostly induced at the early stage of F. proliferatum infection. In roots, AsCHI2 and AsCHI3 mRNA levels were increased in the susceptible and decreased in the resistant cultivar, whereas in cloves, AsCHI7 and AsCHI5 expression was decreased in the susceptible but increased in the resistant plants, suggesting that these genes are involved in the garlic response to Fusarium proliferatum attack. Our results provide insights into the role of chitinases in garlic and may be useful for breeding programs to increase the resistance of Allium crops to Fusarium infections.

Characterization of the complete chloroplast genome of leek Allium porrum L. (Amaryllidaceae).

The complete chloroplast genome sequence of Allium porrum was determined by Illumina single-end sequencing. The complete plastid genome was 152,732 bp in length containing a large single copy (LSC) of 81,744 bp and a small single copy (SSC) of 17,910 bp, which were separated by a pair of 26,524 bp inverted repeats (IRs). A total of 133 genes were annotated, including 80 protein-coding genes, 38 tRNA genes, 8 rRNA genes, and 7 pseudogenes. The overall GC contents of the plastid genome were 36.7%. Unlike A. sativum and A. obliquum in the leek, plastome infA gene is absent, and rpl23 gene is a pseudogene due to a 4 bp deletion and the formation of a premature stop codon.

The YABBY Genes of Leaf and Leaf-Like Organ Polarity in Leafless Plant Monotropa hypopitys.

Monotropa hypopitys is a mycoheterotrophic, nonphotosynthetic plant acquiring nutrients from the roots of autotrophic trees through mycorrhizal symbiosis, and, similar to other extant plants, forming asymmetrical lateral organs during development. The members of the YABBY family of transcription factors are important players in the establishment of leaf and leaf-like organ polarity in plants. This is the first report on the identification of YABBY genes in a mycoheterotrophic plant devoid of aboveground vegetative organs. Seven M. hypopitys YABBY members were identified and classified into four clades. By structural analysis of putative encoded proteins, we confirmed the presence of YABBY-defining conserved domains and identified novel clade-specific motifs. Transcriptomic and qRT-PCR analyses of different tissues revealed MhyYABBY transcriptional patterns, which were similar to those of orthologous YABBY genes from other angiosperms. These data should contribute to the understanding of the role of the YABBY genes in the regulation of developmental and physiological processes in achlorophyllous leafless plants.

Characterization of the complete plastid genome of lop-sided onion Allium obliquum L. (Amaryllidaceae).

The complete chloroplast genome sequence of Allium obliquum was determined by Illumina single-end sequencing. The complete plastid genome was 152,387 bp in length, containing a large single copy (LSC) of 81,588 bp and a small single copy (SSC) of 18,059 bp, which were separated by a pair of 26,370 bp inverted repeats (IRs). A total of 134 genes were annotated, including 83 protein coding genes, 38 tRNA genes, eight rRNA genes, and five pseudogenes. The overall GC contents of the plastid genome were 36.8%. Unlike A. cepa (onion) and A. sativum (garlic), A. obliquum encodes a functional intact infA gene.

De novo transcriptome assembly of the mycoheterotrophic plant Monotropa hypopitys.

Monotropa hypopitys (pinesap) is a non-photosynthetic obligately mycoheterotrophic plant of the family Ericaceae. It obtains the carbon and other nutrients from the roots of surrounding autotrophic trees through the associated mycorrhizal fungi. In order to understand the evolutionary changes in the plant genome associated with transition to a heterotrophic lifestyle, we performed de novo transcriptomic analysis of M. hypopitys using next-generation sequencing. We obtained the RNA-Seq data from flowers, flower bracts and roots with haustoria using Illumina HiSeq2500 platform. The raw data obtained in this study can be available in NCBI SRA database with accession number of SRP069226. A total of 10.3 GB raw sequence data were obtained, corresponding to 103,357,809 raw reads. A total of 103,025,683 reads were filtered after removing low-quality reads and trimming the adapter sequences. The Trinity program was used to de novo assemble 98,349 unigens with an N50 of 1342 bp. Using the TransDecoder program, we predicted 43,505 putative proteins. 38,416 unigenes were annotated in the Swiss-Prot protein sequence database using BLASTX. The obtained transcriptomic data will be useful for further studies of the evolution of plant genomes upon transition to a non-photosynthetic lifestyle and the loss of photosynthesis-related functions.

The complete plastid genome sequence of garlic Allium sativum L.

The complete plastid genome sequence of garlic Allium sativum was determined using Illumina sequencing. The plastid DNA is 153,172 bp in length and includes a large single copy region (LSC) of 82,035 bp and a small single copy region (SSC) of 18,015 bp, which are separated by a pair of 26,561 bp inverted repeat regions (IRs). In total, 134 genes are identified, containing 82 protein-coding genes, 38 tRNA genes, eight rRNA genes and six pseudogenes. Most of genes occur as a single copy, while 19 genes are duplicated in IRs. Among 15 intron-containing genes, clpP and ycf3 contain two introns and the rest have one intron.

The loss of photosynthetic pathways in the plastid and nuclear genomes of the non-photosynthetic mycoheterotrophic eudicot Monotropa hypopitys.

BACKGROUND: Chloroplasts of most plants are responsible for photosynthesis and contain a conserved set of about 110 genes that encode components of housekeeping gene expression machinery and photosynthesis-related functions. Heterotrophic plants obtaining nutrients from other organisms and their plastid genomes represent model systems in which to study the effects of relaxed selective pressure on photosynthetic function. The most evident is a reduction in the size and gene content of the plastome, which correlates with the loss of genes encoding photosynthetic machinery which become unnecessary. Transition to a non-photosynthetic lifestyle is expected also to relax the selective pressure on photosynthetic machinery in the nuclear genome, however, the corresponding changes are less known. RESULTS: Here we report the complete sequence of the plastid genome of Monotropa hypopitys, an achlorophyllous obligately mycoheterotrophic plant belonging to the family Ericaceae. The plastome of M. hypopitys is greatly reduced in size (35,336 bp) and lacks the typical quadripartite structure with two single-copy regions and an inverted repeat. Only 45 genes remained presumably intact- those encoding ribosomal proteins, ribosomal and transfer RNA and housekeeping genes infA, matK, accD and clpP. The clpP and accD genes probably remain functional, although their sequences are highly diverged. The sets of genes for ribosomal protein and transfer RNA are incomplete relative to chloroplasts of a photosynthetic plant. Comparison of the plastid genomes of two subspecies-level isolates of M. hypopitys revealed major structural rearrangements associated with repeat-driven recombination and the presence of isolate-specific tRNA genes. Analysis of the M. hypopitys transcriptome by RNA-Seq showed the absence of expression of nuclear-encoded components of photosystem I and II reaction center proteins, components of cytochrome b6f complex, ATP synthase, ribulose bisphosphate carboxylase components, as well as chlorophyll from protoporphyrin IX biosynthesis pathway. CONCLUSIONS: With the complete loss of genes related to photosynthesis, NADH dehydrogenase, plastid-encoded RNA polymerase and ATP synthase, the M. hypopitys plastid genome is among the most functionally reduced ones characteristic of obligate non-photosynthetic parasitic species. Analysis of the M. hypopitys transcriptome revealed coordinated evolution of the nuclear and plastome genomes and the loss of photosynthesis-related functions in both genomes.