Overexpression of a pearl millet WRKY transcription factor gene, PgWRKY74, in Arabidopsis retards shoot growth under dehydration and salinity-stressed conditions

Pearl millet (Cenchrus americanus) is a cereal crop that can tolerate high temperatures, drought, and low-fertility conditions where other crops lose productivity. However, genes regulating this ability are largely unknown. Transcription factors (TFs) regulate transcription of their target genes, regulate downstream biological processes, and thus are candidates for regulators of such tolerance of pearl millet. PgWRKY74 encodes a group IIc WRKY TF in pearl millet and is downregulated by drought. PgWRKY74 may have a role in drought tolerance. The objective of this study was to gain insights into the physiological and biochemical functions of PgWRKY74. Yeast one-hybrid and gel shift assays were performed to examine transcriptional activation potential and deoxyribonucleic acid (DNA)-binding ability, respectively. Transgenic Arabidopsis thaliana plants overexpressing PgWRKY74-green fluorescent protein (GFP) fusion gene were generated and tested for growth and stress-responsive gene expression under mannitol and NaCl-stressed conditions. A construct with PgWRKY74 enabled yeast reporter cells to survive on test media in the yeast one-hybrid assays. The electrophoretic mobility of DNA with putative WRKY TF-binding motifs was lower in the presence of a recombinant PgWRKY74 protein than its absence. The PgWRKY74-GFP-overexpressing Arabidopsis plants exhibited smaller rosette areas than did wild-type plants under mannitol-stressed and NaCl-stressed conditions, and exhibited weaker expression of RD29B, which is induced by the stress-related phytohormone abscisic acid (ABA), under the mannitol-stressed condition. PgWRKY74 have transcriptional activation potential and DNA-binding ability, and can negatively regulate plant responses to mannitol and NaCl stresses, possibly by decreasing ABA levels or ABA sensitivity

Phenotyping and a genome-wide association study of elite lines of pearl millet

Pearl millet (Pennisetum glaucum (L.) R. BR.) is a cereal crop mainly grown in India and sub-Saharan Africa. In pearl millet, genes and genomic regions associated with traits are largely unknown. Pearl millet parental lines bred at the International Crops Research Institute for the Semi-Arid Tropics (ICRISAT) are useful for the production and breeding of pearl millet. However, the phenotypic diversity of these lines has not been fully evaluated. In this study, 16 traits of 107 of those parental lines were assessed with field trials in Japan, and a genome-wide association study (GWAS) was performed using these phenotypic data to identify the genomic regions and genes associated with those traits. The GWAS revealed genomic regions associated with culm height and pigmentation of the shoot basal part (PS). The genomic region associated with PS contained a homolog of PHENYLALANINE AMMONIA LYASE 2 (PAL2), a gene involved in anthocyanin accumulation in Arabidopsis thaliana. The PAL2 homolog can be a candidate for a gene involved in regulating PS in pearl millet. These results provide a better understanding of the phenotypic diversity of pearl millet and its genetic background

Small RNA sequencing reveals the role of pearl millet miRNAs and their targets in salinity stress responses

Pearl millet [Pennisetum glaucum (L.) R. Br.] is an important crop in arid and semi-arid areas of India and Africa. It is well known for its tolerance to abiotic stresses, but it lags behind other cereals in terms of research and development. MicroRNAs (miRNAs) are a versatile group of small regulatory RNAs of 20�22 bases that play important roles in plant growth, development, and stress responses. However, the regulatory mechanisms underlying miRNA-mediated responses to salinity stress in pearl millet are still unclear. In this study, we performed small RNA sequencing to identify conserved and novel miRNAs from the salinity tolerant pearl millet genotype. In total, 130 million sequence reads were generated, and 81 conserved and 14 novel miRNAs were identified as salinity stress responsive microRNAs. We also performed target prediction for these miRNAs, and a total of 448 pearl millet mRNAs were identified as the targets. Among these target mRNAs, 122 (~25%) encode transcription factors. A pathway analysis showed that differentially expressed miRNAs and their target genes can regulate the auxin response pathway. Quantitative real-time PCR analysis of miRNAs and their targets showed consistent expression patterns. These results suggest that miRNAs play a role in salinity stress tolerance in pearl millet

Genome-wide investigation of SQUAMOSA promoter binding protein-like transcription factor family in pearl millet (Pennisetum glaucum (L) R. Br.)

Abiotic stress negatively affects the yield of many crops. The SQUAMOSA promoter binding protein-like proteins (SBPs) represent a family of plant-specific transcription factors which play essential roles in plant growth, development, and stress responsiveness. In this study, 18 putative SBPs (PgSBPs) were identified in the genome of pearl millet on the basis of the SBP domain. One or two zinc finger-like structure(s) and a nuclear localization signal (NLS) were found in the SBP domains of all the PgSBPs. Fourteen PgSBPs were distributed on 7 chromosomes unevenly, while the other 4 were located on the scaffolds (i.e., non-chromosomal genomic sequences). Moreover, all the PgSBPs were clustered into seven groups (I-VII) based on the phylogenetic analysis. The intron/ exon structures and the motif composition were similar between PgSBPs within the same groups. PgSBPs in groups V and VII were predicted as the targets of two microRNAs, PgmiR156q and PgmiR529b. Some PgSBPs have abscisic acid (ABA)-responsive elements and stress-responsive elements in their promoters. Expression levels of these PgSBPs were upregulated by abiotic stresses and downregulated by ABA. Together, this study shows a comprehensive overview of PgSBPs and provides vital information for elucidating the biological functions of PgSBPs

Analyses of Sequence Features and Expression Patterns of DOF Transcription Factor Genes in Pearl Millet (Pennisetum glaucum (L.) R. Br.)

Pearl millet (Pennisetum glaucum (L.) R. Br.) is a cereal crop that is tolerant to drought, high-temperature and poor-nutrient stresses throughout its life cycle. The DOF (DNA binding with One Finger) family genes encode plant-specific zinc finger transcription factors. These transcription factors have a variety of functions and can be involved in regulating the stress tolerance of pearl millet. In this study, we identified 12 DOF family genes (PgDOFs) in pearl millet. Ten of them were distributed on four chromosomes and the other two were on scaffolds (i.e., non-chromosomal sequences). Protein sequence analysis showed that PgDOFs have the DOF domain in their N-terminal regions and that other conserved motifs are also present in them. PgDOFs and DOF proteins from five other plant species were divided into seven groups, and the 12 PgDOFs could be classified into six of those seven groups. In gene expression analysis, most PgDOFs were upregulated by cold, heat, dehydration and salinity stress. These data can be useful for further characterization of the PgDOFs

Comparative de novo transcriptomic profiling of the salinity stress responsiveness in contrasting pearl millet lines

Pearl millet (Pennisetum glaucum (L) R. Br.) is a staple crop for more than 90 million poor farmers. It is known for its tolerance against drought, salinity, and high temperature. To understand the molecular mechanisms underlying its salinity tolerance, physiological analyses and a comparative transcriptome analysis between salinity tolerant (ICMB 01222) and salinity susceptible (ICMB 081) lines were conducted under control and salinity conditions. The physiological studies revealed that the tolerant line ICMB 01222 had a higher growth rate and accumulated higher amount of sugar in leaves under salinity stress. Sequencing using the Illumina HiSeq 2500 system generated a total of 977 million reads, and these reads were assembled de novo into contigs corresponding to gene products. A total of 11,627 differentially expressed genes (DEGs) were identified in both lines. These DEGs are involved in various metabolic pathways such as plant hormone signal transduction, mitogen-activated protein kinase signaling pathways, and so on. Genes involved in ubiquitin-mediated proteolysis and phenylpropanoid biosynthesis pathways were upregulated in the tolerant line. In contrast, unigenes involved in glycolysis/gluconeogenesis and genes for ribosomes were downregulated in the susceptible line. Genes encoding SBPs (SQUAMOSA promoter binding proteins), which are plant-specific transcription factors, were differentially expressed only in the tolerant line. Functional unigenes and pathways that are identified can provide useful clues for improving salinity stress tolerance in pearl millet

Data of RNA sequencing of pearl millet panicles treated with a high temperature

Pearl millet (Pennisetum glaucum) is a cereal crop that can grow and set seeds even under drought, high temperatures and nutrient-poor conditions. Panicles of two pearl millet cultivars that differ in seed-setting rates were exposed to two different high-temperature treatments at three different developmental stages with three replicates, and RNA was prepared from these panicles. The resulting RNA samples were subjected to sequencing with the Illumina NovaSeq 6000 sequencer. The obtained data were 150-base-paired-end reads and were approximately 5 Gb/sample in total. These read data were deposited as those for a project in the NCBI (National Center for Biotechnology Information) BioProject database

Data from collection and analysis of RNA sequencing data from pearl millet

Pearl millet (Pennisetum glaucum, also known as Cenchrus americanus) is a cereal crop that has a C4 photosynthesis system and that can grow and develop seeds even under stressed conditions including drought-stressed, high temperature-stressed and nutrient-poor conditions. In previous studies, transcriptomes of pearl millet were studied by RNA sequencing (RNA-Seq) to understand mechanisms regulating its development and tolerance to such stressed conditions. Here, RNA-Seq reads from 565 pearl millet samples from 25 projects in the NCBI (National Center for Biotechnology Information) BioProject database were collected and mapped to the pearl millet reference genome to obtain read counts and transcripts per million (TPM) for each pearl millet gene. The count and TPM data for all the 565 samples as well as the attributes of those samples and projects were deposited in the figshare repository (https://doi.org/10.6084/m9.figshare.24902100)