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Plant networks of oscillating genes coordinate internal processes with external cues, contributing to increased fitness. We hypothesized that the response to submergence stress may dynamically change during different times of the day. In this work, we determined the transcriptome (RNA sequencing) of the model monocotyledonous plant, Brachypodium distachyon, during a day of submergence stress, low light, and normal growth. Two ecotypes of differential tolerance, Bd21 (sensitive) and Bd21-3 (tolerant), were included. We submerged 15-day-old plants under a long-day diurnal cycle (16 h light/8 h dark) and collected samples after 8 h of submergence at ZT0 (dawn), ZT8 (midday), ZT16 (dusk), ZT20 (midnight), and ZT24 (dawn). Rhythmic processes were enriched both with up- and down-regulated genes, and clustering highlighted that the morning and daytime oscillator components (PRRs) show peak expression in the night, and a decrease in the amplitude of the clock genes (GI, LHY, RVE) was observed. Outputs included photosynthesis-related genes losing their known rhythmic expression. Up-regulated genes included oscillating suppressors of growth, hormone-related genes with new late zeniths (e.g., JAZ1, ZEP), and mitochondrial and carbohydrate signaling genes with shifted zeniths. The results highlighted genes up-regulated in the tolerant ecotype such as METALLOTHIONEIN3 and ATPase INHIBITOR FACTOR. Finally, we show by luciferase assays that Arabidopsis thaliana clock genes are also altered by submergence changing their amplitude and phase. This study can guide the research of chronocultural strategies and diurnal-associated tolerance mechanisms.

Drought is a natural disaster that negatively impacts agricultural crops. Understanding the mechanisms of water deficit response is key to improving drought-tolerant crops. This study focused on comprehending the drought-tolerant mechanisms of a mutant chili, CaWDT-2, by investigating its physiological phenotypes and gene expression under PEG-induced drought condition compared to Tavee 60 as the original cultivar. Chili physiological status was monitored by shoot height, root length, leaf number, leaf area, shoot mass and root mass at 10 d after drought initiation. Relative water content (RWC), malondialdehyde (MDA), proline, soluble sugar and sucrose were measured during drought and after recovery. Transcriptome analysis using sequencing of RNA (RNA-seq) was employed to examine gene expression profiling under drought for 24 h. Results revealed diverse changes in physiological mechanism in response to drought between CaWDT-2 and Tavee 60 cultivars. Comparative transcriptomic analysis is helpful in understanding the differences in drought tolerance mechanisms between 2 crop varieties. Major differences that emerged from the physiological and transcriptomic analyses included drought-stressed injury level, proline accumulation, root cell wall adaptation, ABA biosynthesis, stress detoxification, heat shock proteins and transcription factors. Our data will contribute to further research in developing droughttolerant crops. An improved understanding of drought tolerance mechanisms will enhance agricultural resilience and mitigate the impact of drought on crop yield. [ABSTRACT FROM AUTHOR]

Translational regulation contributes to plasticity in metabolism and growth that enables plants to survive in a dynamic environment. Here, we used the precise mapping of ribosome footprints (RFs) on mRNAs to investigate translational regulation under control and sublethal hypoxia stress conditions in seedlings of Arabidopsis thaliana. Ribosomes were obtained by differential centrifugation or immunopurification and were digested with RNase I to generate footprint fragments that were deep-sequenced. Comparison of RF number and position on genic regions with fragmented total and polysomal mRNA illuminated numerous aspects of posttranscriptional and translational control under both growth conditions. When seedlings were oxygen-deprived, the frequency of ribosomes at the start codon was reduced, consistent with a global decline in initiation of translation. Hypoxia-up-regulated gene transcripts increased in polysome complexes during the stress, but the number of ribosomes per transcript relative to normoxic conditions was not enhanced. On the other hand, many mRNAs with limited change in steady-state abundance had significantly fewer ribosomes but with an overall similar distribution under hypoxia, consistent with restriction of initiation rather than elongation of translation. RF profiling also exposed the inhibitory effect of upstream ORFs on the translation of downstream protein-coding regions under normoxia, which was further modulated by hypoxia. The data document translation of alternatively spliced mRNAs and expose ribosome association with some noncoding RNAs. Altogether, we present an experimental approach that illuminates prevalent and nuanced regulation of protein synthesis under optimal and energy-limiting conditions.

Abiotic stress, including drought, salinity, and temperature extremes, regulates gene expression at the transcriptional and posttranscriptional levels. Expression profiling of total messenger RNAs (mRNAs) from rice (Oryza sativa) leaves grown under stress conditions revealed that the transcript levels of photosynthetic genes are reduced more rapidly than others, a phenomenon referred to as stress-induced mRNA decay (SMD). By comparing RNA polymerase II engagement with the steady-state mRNA level, we show here that SMD is a posttranscriptional event. The SMD of photosynthetic genes was further verified by measuring the half-lives of the small subunit of Rubisco (RbcS1) and Chlorophyll a/b-Binding Protein1 (Cab1) mRNAs during stress conditions in the presence of the transcription inhibitor cordycepin. To discern any correlation between SMD and the process of translation, changes in total and polysome-associated mRNA levels after stress were measured. Total and polysome-associated mRNA levels of two photosynthetic (RbcS1 and Cab1) and two stress-inducible (Dehydration Stress-Inducible Protein1 and Salt-Induced Protein) genes were found to be markedly similar. This demonstrated the importance of polysome association for transcript stability under stress conditions. Microarray experiments performed on total and polysomal mRNAs indicate that approximately half of all mRNAs that undergo SMD remain polysome associated during stress treatments. To delineate the functional determinant(s) of mRNAs responsible for SMD, the RbcS1 and Cab1 transcripts were dissected into several components. The expressions of different combinations of the mRNA components were analyzed under stress conditions, revealing that both 3' and 5' untranslated regions are necessary for SMD. Our results, therefore, suggest that the posttranscriptional control of photosynthetic mRNA decay under stress conditions requires both 3' and 5' untranslated regions and correlates with differential polysome association. [ABSTRACT FROM AUTHOR]

Enhancing crop tolerance to waterlogging is critical for improving food and biofuel security. In waterlogged soils, roots are exposed to a low oxygen environment. The group VII ethylene response factors (ERFVIIs) were recently identified as key regulators of plant low oxygen response. Oxygen-dependent N-end rule pathways can regulate the stability of ERFVIIs. This study aims to characterize the function of the Jatropha curcas ERFVIIs and the impact of N-terminal modification that stabilized the protein toward low oxygen response. This study revealed that all three JcERFVII proteins are substrates of the N-end rule pathway. Overexpression of JcERFVII2 conferred tolerance to low oxygen stress in Arabidopsis. In contrast, the constitutive overexpression of stabilized JcERFVII2 reduced low oxygen tolerance. RNA-seq was performed to elucidate the functional roles of JcERFVII2 and the impact of its N-terminal modification. Overexpression of both wildtype and stabilized JcERFVII2 constitutively upregulated the plant core hypoxia-responsive genes. Besides, overexpression of the stabilized JcERFVII2 further upregulated various genes controlling fermentative metabolic processes, oxidative stress, and pathogen responses under aerobic conditions. In summary, JcERFVII2 is an N-end rule regulated waterlogging-responsive transcription factor that modulates the expression of multiple stress-responsive genes; therefore, it is a potential candidate for molecular breeding of multiple stress-tolerant crops. [ABSTRACT FROM AUTHOR]

Vigna vexillata (zombi pea) is an underutilized legume crop considered to be a potential gene source in breeding for abiotic stress tolerance. This study focuses on the molecular characterization of mechanisms controlling waterlogging tolerance using two zombi pea varieties with contrasting waterlogging tolerance. Morphological examination revealed that in contrast to the sensitive variety, the tolerant variety was able to grow, maintain chlorophyll, form lateral roots, and develop aerenchyma in hypocotyl and taproots under waterlogging. To find the mechanism controlling waterlogging tolerance in zombi pea, comparative transcriptome analysis was performed using roots subjected to short-term waterlogging. Functional analysis indicated that glycolysis and fermentative genes were strongly upregulated in the sensitive variety, but not in the tolerant one. In contrast, the genes involved in auxin-regulated lateral root initiation and formation were expressed only in the tolerant variety. In addition, cell wall modification, aquaporin, and peroxidase genes were highly induced in the tolerant variety under waterlogging. Our findings suggest that energy management and root plasticity play important roles in mitigating the impact of waterlogging in zombi pea. The basic knowledge obtained from this study can be used in the molecular breeding of waterlogging-tolerant legume crops in the future. [ABSTRACT FROM AUTHOR]

<b>Background and Objective:</b> <i>Flowering locus T</i> (<i>FT</i>) genes are involved in the flower induction mechanism in plants as florigen signals. The objective of this study was to study the relationship between the expression of <i>Flowering locus T</i> genes (<i>MeFTs</i>) in cassava and flowering traits under the different flowering induction conditions. <b>Materials and Methods:</b> The experimental design for flowering induction was RCBD for 4 replications. There were 5 treatment factors which were control, red light set from 5 pm to 7 am, 0.5 mM 6-benzyladenine (BA) with 2 mM silver thio-sulfate (STS), paclobutrazol for 6 g/plant and potassium chlorate (KCIO<sub>3</sub>) for 250 g/plant. The number of plants with flower bunches and the average number of bunches per plant in two cassava varieties were collected each month from 5-9 months after planting (MAP). The leaf samples were collected from HB80 and R9 varieties at 5-7 MAP for RNA extraction to study <i>MeFT1</i> and <i>MeFT2</i> expression. <b>Results:</b> The results show that <i>MeFT1</i> expression level positively correlated with flowering traits in the same month, while <i>MeFT2</i> expression level positively correlated with flowering traits in the following months. <b>Conclusion:</b> Therefore, expression of <i>MeFT2</i> can be used for the prediction of cassava flowering in the following month which will assist the breeder for the crossing management.

Background: Asian Palmyra palm, the source of palm-sugar, is dioecious with a long juvenile period requiring at least 12 years to reach its maturity. To date, there is no reliable molecular marker for identifying sexes before the first bloom, limiting crop designs and utilization. We aimed to identify sex-linked markers for this palm using PCR-based DNA fingerprinting, suppression subtractive hybridization (SSH) and transcriptome sequencing., Methods: DNA fingerprints were generated between males and females based on RAPD, AFLP, SCoT, modified SCoT, ILP, and SSR techniques. Large-scale cloning and screening of SSH libraries and de novo transcriptome sequencing of male and female cDNA from inflorescences were performed to identify sex-specific genes for developing sex-linked markers., Results: Through extensive screening and re-testing of the DNA fingerprints (up to 1,204 primer pairs) and transcripts from SSH (>10,000 clones) and transcriptome data, however, no sex-linked marker was identified. Although de novo transcriptome sequencing of male and female inflorescences provided ∼32 million reads and 187,083 assembled transcripts, PCR analysis of selected sex-highly represented transcripts did not yield any sex-linked marker. This result may suggest the complexity and small sex-determining region of the Asian Palmyra palm. To this end, we provide the first global transcripts of male and female inflorescences of Asian Palmyra palm. Interestingly, sequence annotation revealed a large proportion of transcripts related to sucrose metabolism, which corresponds to the sucrose-rich sap produced in the inflorescences, and these transcripts will be useful for further understanding of sucrose production in sugar crop plants. Provided lists of sex-specific and differential-expressed transcripts would be beneficial to the further study of sexual development and sex-linked markers in palms and related species., Competing Interests: The authors declare there are no competing interests.

Translational regulation is important for plant growth, metabolism, and acclimation to environmental challenges. Ribosome profiling involves the nuclease digestion of mRNAs associated with ribosomes and mapping of the generated ribosome-protected footprints to transcripts. This is useful for investigation of translational regulation. Here we present a detailed method to generate, purify, and high-throughput-sequence ribosome footprints from Arabidopsis thaliana using two different isolation methods, namely, conventional differential centrifugation and the translating ribosome affinity purification (TRAP) technology. These methodologies provide researchers with an opportunity to quantitatively assess with high-resolution the translational activity of individual mRNAs by determination of the position and number of ribosomes in the corresponding mRNA. The results can provide insights into the translation of upstream open reading frames, alternatively spliced transcripts, short open reading frames, and other aspects of translation.

Translating Ribosome Affinity Purification (TRAP) is a technology to isolate the population of mRNAs associated with at least one 80S ribosome, referred as the translatome. TRAP is based on the expression of an epitope-tagged version of a ribosomal protein and the affinity purification of ribosomes and associated mRNAs using antibodies conjugated to agarose beads. Quantitative assessment of the translatome is achieved by direct RNA sequencing (RNA-SEQ), which provides accurate quantitation of ribosome-associated mRNAs and reveals alternatively spliced isoforms. Here we present a detailed procedure for TRAP, as well as a guide for preparation of RNA-SEQ libraries (TRAP-SEQ) and a primary data analysis. This methodology enables the study of translational dynamic by assessing rapid changes in translatomes, at organ or cell-type level, during development or in response to endogenous or exogenous stimuli.