Transcriptome and co-expression network revealed molecular mechanism underlying selenium response of foxtail millet (Setaria italica).

Introduction: Selenium-enriched foxtail millet (Setaria italica) represents a functional cereal with significant health benefits for humans. This study endeavors to examine the impact of foliar application of sodium selenite (Na2SeO4) on foxtail millet, specifically focusing on selenium (Se) accumulation and transportation within various plant tissues. Methods: To unravel the molecular mechanisms governing selenium accumulation and transportation in foxtail millet, we conducted a comprehensive analysis of selenium content and transcriptome responses in foxtail millet spikelets across different days (3, 5, 7, and 12) under Na2SeO4 treatment (200 µmol/L). Results: Foxtail millet subjected to selenium fertilizer exhibited significantly elevated selenium levels in each tissue compared to the untreated control. Selenate was observed to be transported and accumulated sequentially in the leaf, stem, and spikes. Transcriptome analysis unveiled a substantial upregulation in the transcription levels of genes associated with selenium metabolism and transport, including sulfate, phosphate, and nitrate transporters, ABC transporters, antioxidants, phytohormone signaling, and transcription factors. These genes demonstrated intricate interactions, both synergistic and antagonistic, forming a complex network that regulated selenate transport mechanisms. Gene co-expression network analysis highlighted three transcription factors in the tan module and three transporters in the turquoise module that significantly correlated with selenium accumulation and transportation. Expression of sulfate transporters (SiSULTR1.2b and SiSULTR3.1a), phosphate transporter (PHT1.3), nitrate transporter 1 (NRT1.1B), glutathione S-transferase genes (GSTs), and ABC transporter (ABCC13) increased with SeO4 2- accumulation. Transcription factors MYB, WRKY, and bHLH were also identified as players in selenium accumulation. Conclusion: This study provides preliminary insights into the mechanisms of selenium accumulation and transportation in foxtail millet. The findings hold theoretical significance for the cultivation of selenium-enriched foxtail millet.

Genome-wide identification and expression profiling of the ABI5 gene family in foxtail millet (Setaria italica).

BACKGROUND: ABA Insensitive 5 (ABI5) is a basic leucine zipper transcription factor that crucially influences plant growth, development, and stress response. However, there is minimal research on the ABI5 family in foxtail millet. RESULTS: In this study, 16 ABI5 genes were identified in foxtail millet, and their sequence composition, gene structures, cis-acting elements, chromosome positions, and gene replication events were analyzed. To more thoroughly evaluate the developmental mechanisms of the SiABI5 family during evolution, we selected three dicotyledons (S. lycopersicum, A. thaliana, F. tataricum) and three (Z. mays, O. sativa, S. bicolor) specific representative monocotyledons associated with foxtail millet for comparative homology mapping. The results showed that foxtail millet ABI5 genes had the best homology with maize. A promoter sequence analysis showed that the SiABI5s contain numerous cis-acting elements related to hormone and stress responses, indicating that the regulation of SiABI5 expression was complex. The expression responses of 16 genes in different tissues, seed germination, and ear development were analyzed. A total of six representative genes were targeted from five subfamilies to characterize their gene expression responses to four different abiotic stresses. Overexpression of SiABI5.12 confers tolerance to osmotic stress in transgenic Arabidopsis thaliana, which demonstrated the function of SiABI5 responded to abiotic stress. CONCLUSIONS: In summary, our research results comprehensively characterized the SiABI5 family and can provide a valuable reference for demonstrating the role of SiABI5s in regulating abiotic stress responses in foxtail millet.

Distinctive physiological and molecular responses of foxtail millet and maize to nicosulfuron.

Introduction: Nicosulfuron is the leading acetolactate synthase inhibitor herbicide product, and widely used to control gramineous weeds. Here, we investigated the metabolic process of nicosulfuron into foxtail millet and maize, in order to clarify the mechanism of the difference in sensitivity of foxtail millet and maize to nicosulfuron from the perspective of physiological metabolism and provide a theoretical basis for the breeding of nicosulfuron-resistant foxtail millet varieties. Methods: We treated foxtail millet (Zhangzagu 10, Jingu 21) and maize (Nongda 108, Ditian 8) with various doses of nicosulfuron in both pot and field experiments. The malonaldehyde (MDA) content, target enzymes, detoxification enzymes, and antioxidant enzymes, as well as related gene expression levels in the leaf tissues of foxtail millet and maize were measured, and the yield was determined after maturity. Results: The results showed that the recommended dose of nicosulfuron caused Zhangzagu 10 and Jingu 21 to fail to harvest; the yield of the sensitive maize variety (Ditian 8) decreased by 37.09%, whereas that of the resistant maize variety (Nongda 108) did not decrease. Nicosulfuron stress increased the CYP450 enzyme activity, MDA content, and antioxidant enzyme activity of foxtail millet and maize, reduced the acetolactate synthase (ALS) activity and ALS gene expression of foxtail millet and Ditian 8, and reduced the glutathione S-transferase (GST) activity and GST gene expression of foxtail millet. In conclusion, target enzymes, detoxification enzymes, and antioxidant enzymes were involved in the detoxification metabolism of nicosulfuron in plants. ALS and GST are the main factors responsible for the metabolic differences among foxtail millet, sensitive maize varieties, and resistant maize varieties. Discussion: These findings offer valuable insights for exploring the target resistance (TSR) and non-target resistance (NTSR) mechanisms in foxtail millet under herbicide stress and provides theoretical basis for future research of develop foxtail millet germplasm with diverse herbicide resistance traits.

Optimized potassium application rate increases foxtail millet grain yield by improving photosynthetic carbohydrate metabolism.

Photosynthetic carbohydrate metabolism is an important biochemical process related to crop production and quality traits. Potassium (K) critically contributes to the process of photosynthetic carbon assimilation and carbohydrate metabolism. We explored the effects of potassium fertilization on physiological mechanisms including carbohydrate metabolism in foxtail millet and its yield. Field experiments were performed using two foxtail millet (Setaria italica L.) cultivars: 1) Jingu 21 (low-K sensitive); and 2) Zhangza 10 (low-K tolerant). Effect of five different potassium fertilizer (K2O) rates (0, 60, 120, 180, and 240 kg·hm-2) were tested in two consecutive years, 2020 and 2021. We found that potassium application significantly increases the K content, dry matter accumulation and yield. Jingu 21 and Zhangza 10 had maximum yields at 180 kg·hm-2 K application, which were 29.91% and 31.51% larger than without K application, respectively. Excessive K application (K240) did not further improve their yields. The suitable K fertilizer application of Jingu 21 and Zhangza 10 are 195.25-204.27 and 173.95-175.87 kg·K2O·hm-2, respectively. The net photosynthetic rate (Pn), ribulose-1,5-bisphosphate carboxylase-oxygenase (Rubisco), phosphoenolpyruvate carboxylase (PEPC), and fructose-1,6-bisphosphatase (FBPase) were positively correlated with the potassium content. Potassium application improved the availability of carbon sources for carbohydrate synthesis. Compared with the K0 treatment, variations in the activities of sucrose phosphate synthase (SPS) and sucrose synthase (SuSy) in potassium-treated Jingu 21 (K60, K120, K180, and K240) were 17.94%-89.93% and 22.48%-182.10%, respectively, which were greater than those of Zhangza 10 (11.34%-71.12% and 16.18%-109.13%, respectively) and indicate that Jingu 21 is more sensitive to potassium application. The sucrose contents in the Jingu 21 and Zhangza 10 grains were 0.97%-1.15% and 1.04%-1.23%, respectively. The starch contents were 28.99%-37.75% and 24.81%-34.62%, respectively. The sucrose: ratio of Jingu 21 was smaller than that of Zhangza 10, indicating that Jingu 21 utilized nutrients better than Zhangza 10. Stepwise regression and path analysis showed that leaf and grain SuSy activity, by coordinating the source-sink relationship, have the greatest direct effect on Jingu 21 yield, whereas leaf SuSy activity, by promoting the generation of photosynthates at the source leaf, plays a leading role in Zhangza 10 yield increase. In conclusion, optimized K application can increase foxtail millet grain yield by improving photosynthesis and promoting carbohydrate accumulation and distribution.

Humic acid improves the physiological and photosynthetic characteristics of millet seedlings under drought stress.

We aimed to determine whether humic acid (HA) can alleviate the injury of millet caused by drought and its potential mechanism. Millet seeds (Jingu 21 and Zhangza 10) were soaked in different concentrations of HA (0, 50, 10, 200, and 300 mg L-1) for 12 h. The physiological and photosynthetic characteristics of millet seedlings, including growth parameters, osmotic regulators, antioxidase activity, photosynthesis, chlorophyll fluorescence, and P700 parameters, were determined before and after drought stress. HA significantly promoted the growth of millet seedlings under drought stress. Pretreatment with 100 mg L-1 or 200 mg L-1 HA significantly increased free proline, soluble protein, and activity of the antioxidant enzyme system (superoxide dismutase, peroxidase, and catalase) in both Zhangza 10 and Jingu 21. The accumulation of reactive oxygen species ([Formula: see text] and H2O2) was reduced in HA treatments compared with that of the control (P < .05). Moreover, HA (100 mg L-1) significantly increased net photosynthetic rate, stomatal conductance, effective quantum yield of photosystem II, relative photosynthetic electron transfer rate of photosystem II, and photochemical quenching. HA also reduced intercellular CO2 concentration and non-photochemical quenching. Furthermore, 200 mg L-1 HA significantly increased the maximum P700, effective quantum yield of photosystem I, and relative photosynthetic electron transfer rate of photosystem I in Zhangza 10 and decreased non-photochemical energy dissipation in Jingu 21 and Zhangza 10 under drought stress. HA promoted the growth of millet seedlings under drought stress by promoting the osmotic adjustment ability and antioxidant capacity of seedlings and increased photosynthesis.

Comprehensive evaluation of fluroxypyr herbicide on physiological parameters of spring hybrid millet.

Foxtail millet (Setaria italic L.) is an important food and fodder crop that is cultivated worldwide. Quantifying the effects of herbicides on foxtail millet is critical for safe herbicide application. In this study, we analyzed the effects of different fluroxypyr dosages on the growth parameters and physiological parametric of foxtail millet, that is, peroxidation characteristics, photosynthetic characteristics, and endogenous hormone production, by using multivariate statistical analysis. Indicators were screened via Fisher discriminant analysis, and the growth parameters, peroxidation characteristics, photosynthesis characteristics and endogenous hormones of foxtail millet at different fluroxypyr dosages were comprehensively evaluated by principal component analysis. On the basis of the results of principal component analysis, the cumulative contribution rate of the first two principal component factors was 93.72%. The first principal component, which explained 59.23% of total variance, was selected to represent the photosynthetic characteristics and endogenous hormones of foxtail millet. The second principal component, which explained 34.49% of total variance, represented the growth parameters of foxtail millet. According to the principal component analysis, the indexes were simplified into comprehensive index Z, and the mathematical model of comprehensive index Z was set as F = 0.592Z 1 + 0.345Z 2. The results showed that the comprehensive evaluation score of fluroxypyr at moderate concentrations was higher than at high concentrations. Consequently, one L (active ingredient, ai) ha-1 fluroxypyr exerted minimal effects on growth parameters, oxidase activity, photosynthetic activity, and endogenous hormones, and had highest value of comprehensive evaluation, which had efficient and safe benefits in foxtail millet field.

Comparative Genomics Analysis in Grass Species Reveals Two Distinct Evolutionary Strategies Adopted by R Genes.

Resistance genes play an important role in the defense of plants against the invasion of pathogens. In Setaria italica and closely related grass species, R genes have been identified through genetic mapping and genome-wide homologous/domain searching. However, there has been to date no systematic analysis of the evolutionary features of R genes across all sequenced grass genomes. Here, we determined and comprehensively compared R genes in all 12 assembled grass genomes and an outgroup species (Arabidopsis thaliana) through synteny and selection analyses of multiple genomes. We found that the two groups of nucleotide binding site (NBS) domains containing R genes-R tandem duplications (TD) and R singletons-adopted different strategies and showed different features in their evolution. Based on Ka/Ks analysis between syntenic R loci pairs of TDs or singletons, we conclude that R singletons are under stronger purifying selection to be conserved among different grass species than R TDs, while R genes located at TD arrays have evolved much faster through diversifying selection. Furthermore, using the variome datasets of S. italica populations, we scanned for selection signals on genes and observed that a part of R singleton genes have been under purifying selection in populations of S. italica, which is consistent with the pattern observed in syntenic R singletons among different grass species. Additionally, we checked the synteny relationships of reported R genes in grass species and found that the functionally mapped R genes for novel resistance traits are prone to appear in TDs and are heavily divergent from their syntenic orthologs in other grass species, such the black streak R gene Rxo1 in Z. mays and the blast R gene Pi37 in O. sativa. These findings indicate that the R genes from TDs adopted tandem duplications to evolve faster and accumulate more mutations to facilitate functional innovation to cope with variable threats from a fluctuating environment, while R singletons provide a way for R genes to maintain sequence stability and retain conservation of function.

Spraying Brassinolide improves Sigma Broad tolerance in foxtail millet (Setaria italica L.) through modulation of antioxidant activity and photosynthetic capacity.

To explore the role of Brassinolide (BR) in improving the tolerance of Sigma Broad in foxtail millet (Setaria italica L.), effects of 0.1 mg/L of BR foliar application 24 h before 3.37 g/ha of Sigma Broad treatment at five-leaf stage of foxtail millet on growth parameters, antioxidant enzymes, malondialdehyde (MDA), chlorophyll, net photosynthetic rate (P N), chlorophyll fluorescence and P700 parameters were studied 7 and 15 d after herbicide treatment, respectively. Results showed that Sigma Broad significantly decreased plant height, activities of superoxide dismutase (SOD), chlorophyll content, P N, PS II effective quantum yield (Y (II)), PS II electron transport rate (ETR (II)), photochemical quantum yield of PSI(Y (I)) and PS I electron transport rate ETR (I), but significantly increased MDA. Compared to herbicide treatment, BR dramatically increased plant height, activities of SOD, Y (II), ETR (II), Y (I) and ETR (I). This study showed BR pretreatment could improve the tolerance of Sigma Broad in foxtail millet through improving the activity of antioxidant enzymes, keeping electron transport smooth, and enhancing actual photochemical efficiency of PS II and PSI.

Increasing Selenium and Yellow Pigment Concentrations in Foxtail Millet (Setaria italica L.) Grain with Foliar Application of Selenite.

Although addition of selenium (Se) is known to increase Se in crops, it is unclear whether exogenous Se is linked to nutritional and functional components in foxtail millet (Setaria italica L.). In this study, we examined the potential of increasing Se and yellow pigment (YP) in foxtail millet grain by foliar application of Se. Field experiments were conducted during the growing season of foxtail millet in 2013 and 2014 to assess the effects of foliar spray of sodium selenite (10-210 g Se ha(-1)) on the yield, Se uptake and accumulation, total YP, and microminerals in the grain. Average grain yields with Se application were 5.60 and 4.53 t ha(-1) in the 2 years, showing no significant differences from the unfertilized control. However, grain Se concentration increased linearly with Se application rate, by 8.92 and 6.09 µg kg(-1) in the 2 years with application of 1 g Se ha(-1) (maximum grain recovery rates of Se fertilizer, 52 and 28 %). Likewise, total grain YP concentration markedly increased by 0.038 and 0.031 mg kg(-1) in the 2 years with application of 1 g Se ha(-1). Grain Mn, Cu, Fe, and Zn concentrations were not significantly affected by Se application. This study indicated that foliar application of Se effectively and reliably increased the concentrations of Se and YP in foxtail millet grain without affecting the yield or mineral micronutrient concentrations. Thus, foliar-applied selenite has a significant potential to increase the concentrations of selenium and YP (putative lutein (Shen, J Cereal Sci 61:86-93, 2015; Abdel-Aal, Cereal Chem 79:455-457, 2002; Abdel-Aal, J Agric Food Chem 55:787-794, 2007)) of foxtail millet and, thus, the health benefits of this crop.

Grain Yield and Quality of Foxtail Millet (Setaria italica L.) in Response to Tribenuron-Methyl.

Foxtail millet (Setaria italica L.) is cultivated around the world for human and animal consumption. There is no suitable herbicide available for weed control in foxtail millet fields during the post-emergence stage. In this study, we investigated the effect and safety of the post-emergence herbicide tribenuron-methyl (TBM) on foxtail millet in terms of grain yield and quality using a split-plot field design. Field experiments were conducted using two varieties in 2013 and 2014, i.e., high-yielding hybrid Zhangzagu 10 and high-quality conventional Jingu 21. TBM treatments at 11.25 to 90 g ai ha(-1) reduced root and shoot biomass and grain yield to varying degrees. In each of the two years, grain yield declined by 50.2% in Zhangzagu 10 with a herbicide dosage of 45 g ai ha(-1) and by 45.2% in Jingu 21 with a herbicide dosage of 22.5 g ai ha(-1) (recommended dosage). Yield reduction was due to lower grains per panicle, 1000-grain weight, panicle length, and panicle diameter. Grain yield was positively correlated with grains per panicle and 1000-grain weight, but not with panicles ha(-1). With respect to grain protein content at 22.5 g ai ha(-1,) Zhangzagu 10 was similar to the control, whereas Jingu 21 was markedly lower. An increase in TBM dosage led to a decrease in grain Mn, Cu, Fe, and Zn concentrations. In conclusion, the recommended dosage of TBM was relatively safe for Zhangzagu 10, but not for Jingu 21. Additionally, the hybrid variety Zhangzagu 10 had a greater tolerance to TBM than the conventional variety Jingu 21.

Photosynthetic physiological response of Radix Isatidis (Isatis indigotica Fort.) seedlings to nicosulfuron.

Radix Isatidis (Isatis indigotica Fort.) is one of the most important traditional Chinese medicine plants. However, there is no suitable herbicide used for weed control in Radix Isatidis field during postemergence stage. To explore the safety of sulfonylurea herbicide nicosulfuron on Radix Isatidis (Isatis indigotica Fort.) seedlings and the photosynthetic physiological response of the plant to the herbicide, biological mass, leaf area, photosynthetic pigment content, photosynthetic rate, chlorophyll fluorescence characteristics, and P700 parameters of Radix Isatidis seedlings were analyzed 10 d after nicosulfuron treatment at 5th leaf stage in this greenhouse research. The results showed that biological mass, total chlorophyll, chlorophyll a, and carotenoids content, photosynthetic rate, stomatal conductance, PS II maximum quantum yield, PS II effective quantum yield, PS II electron transport rate, photochemical quenching, maximal P700 change, photochemical quantum yield of PS I, and PS I electron transport rate decreased with increasing herbicide concentrations, whereas initial fluorescence, quantum yield of non-regulated energy dissipation in PS II and quantum yield of non-photochemical energy dissipation due to acceptor side limitation in PS I increased. It suggests that nicosulfuron ≥1 mg L-1 causes the damage of chloroplast, PS II and PS I structure. Electron transport limitations in PS I receptor side, and blocked dark reaction process may be the main cause of the significantly inhibited growth and decreased photosynthetic rate of Radix Isatidis seedlings.