Transcriptional repressor RST1 controls salt tolerance and grain yield in rice by regulating gene expression of asparagine synthetase.

Salt stress impairs nutrient metabolism in plant cells, leading to growth and yield penalties. However, the mechanism by which plants alter their nutrient metabolism processes in response to salt stress remains elusive. In this study, we identified and characterized the rice (Oryza sativa) rice salt tolerant 1 (rst1) mutant, which displayed improved salt tolerance and grain yield. Map-based cloning revealed that the gene RST1 encoded an auxin response factor (OsARF18). Molecular analyses showed that RST1 directly repressed the expression of the gene encoding asparagine synthetase 1 (OsAS1). Loss of RST1 function increased the expression of OsAS1 and improved nitrogen (N) utilization by promoting asparagine production and avoiding excess ammonium (NH4+) accumulation. RST1 was undergoing directional selection during domestication. The superior haplotype RST1Hap III decreased its transcriptional repression activity and contributed to salt tolerance and grain weight. Together, our findings unravel a synergistic regulator of growth and salt tolerance associated with N metabolism and provide a new strategy for the development of tolerant cultivars.

Research Note: Creatine monohydrate alleviates protein breakdown induced by corticosterone via inhibiting ubiquitin proteasome pathway in chicken myotubes.

Stress is a common problem diminishing the muscle development of broilers. Creatine (Cr), an energy buffer in skeletal muscle, plays a fundamental role in muscle physiology. This study aimed to evaluate the effect of Cr monohydrate (CMH) on protein breakdown in chicken myotubes challenged by corticosterone (CORT) in vitro. The morphology of myotube was measured and the activation of ubiquitin proteasome (UP) pathway was determined. The result showed that CORT treatment decreased myotube diameter (P < 0.05), increased 3-methyl-histidine (3M-His) content in medium, enhanced the mRNA expression levels of muscle ring finger1(MuRF1) and Atrogin1 (P < 0.001), and Atrogin1 protein level (P < 0.05) compared with control. By contrast, CMH increased myotube diameter (P < 0.05) and myosin heavy chain (MHC) expression (P < 0.001), whereas decreased 3M-His and the mRNA and protein levels of Atrogin1 (P < 0.05), compared to control. In the present of CMH, the decreased myotube diameter and increased 3M-His, mRNA levels of MuRF1 and Atrogin1, and Atrogin1 protein level by CORT were partially relieved (P < 0.05). Hence, the result suggests that CMH alleviates CORT-induced protein breakdown by suppressing Atrogin1 expression in chicken myotubes. The result highlights the potential application of CMH in regulating muscle protein catabolism in chickens under stress.

The optimal dietary arginine level of laying hens fed with low-protein diets.

BACKGROUND: Arginine (Arg) is an essential amino acid (EAA) in poultry, an important substrate for protein synthesis and a precursor of several molecules. Supplementation of EAAs with low protein (LP) diet increases the utilization efficiency of dietary crude protein (CP). However, if the EAA requirement is changed in hens fed a LP diet remains to be elucidated. The aim of the present study was to evaluate the optimal level of dietary Arg in the LP diet of hens. A total of 1350 Hy-Line Brown laying hens were randomly allocated to six dietary treatments: a basal diet (16% CP, positive control), or an isoenergetic LP diet (14% CP, 0.80% Arg) supplemented 0, 0.05%, 0.10%, 0.15%, and 0.20% L-Arg, corresponding to 0.80%, 0.85%, 0.90%, 0.95% and 1.00% dietary Arg, respectively. RESULTS: The feed efficiency was decreased (P < 0.05) by 0.80% and 1.00% Arg-LP diets, compared to control. Within LP diets, dietary Arg level had significant quadratic effects (P < 0.05) on laying rate, egg mass, and feed efficiency. Compared to control, the plasma CAT activity or T-AOC content were decreased by 0.80% (P < 0.001). However, the hens offered 0.85% and 0.90% Arg-LP diets had higher CAT activity (P < 0.001) than 0.80% Arg-LP diet. In contrast, 1.00% Arg-LP group had the highest MDA and the lowest T-AOC content in plasma, liver, duodenal and jejunal mucosa (P < 0.05). Compared to control, the villus height was decreased by 0.80%, 0.95% and 1.00% Arg-LP diets, while the villus height to crypt depth (V/C) ratio was reduced by 0.95% and 1.00% Arg-LP diets in duodenum. CONCLUSION: The result demonstrates that LP diet (14% CP) deficient in Arg (0.80% Arg) result in augmented oxidative damage and impaired development of intestinal mucosa. According to the quadratic broken-line regression model, the optimal dietary arginine levels for Hy-Line Brown laying hens fed with low protein diet (14% CP) aged 33 to 40 weeks are 0.85%, 0.86%, and 0.86% to obtained the maximum laying rate, egg mass, and feed efficiency, respectively.

The regulating pathway of creatine on muscular protein metabolism depends on the energy state.

Creatine (Cr) is beneficial for increasing muscle mass and preventing muscle atrophy via involving in energy metabolism through the Cr and phosphocreatine (PCr) system. This study aimed to evaluate the supplemental effect of Cr on protein metabolism under normal and starvation conditions. The primary myoblasts were obtained from the breast muscle of chicks. The mammalian target of rapamycin (mTOR)/P70S6 kinase (P70S6K), ubiquitin-proteasome (UP) pathways, and mitochondrial function of myotubes were evaluated at normal or starvation state and with or without glucose supplementation. Under normal condition, Cr supplementation enhanced protein synthesis rate as well as upregulated the total and phosphorylated P70S6K expressions. Cr had little influence on protein catabolism and mitochondrial function. In a starvation state, however, Cr alleviated myotube atrophy and enhanced protein accretion by inhibiting Atrogin1 and myostatin (MSTN) expression. Furthermore, Cr treatment upregulated the transcriptional coactivators peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) expression and decreased reactive oxygen species (ROS) accumulation under starvation condition. In the presence of glucose, however, the favorable effect of Cr on protein content and myotube diameter did not occur under starvation condition. The present result indicates that at a normal state, Cr stimulated protein synthesis via the mTOR/P70S6K pathway. In a starvation state, Cr mainly takes a favorable effect on protein accumulation via suppression of the UP pathway and mediated mitochondrial function mainly by serving as an energy supplier. The result highlights the potential clinical application for the modulation of muscle mass under different nutritional conditions.

Research Note: Effects of dietary L-arginine on the production performance and gene expression of reproductive hormones in laying hens fed low crude protein diets.

The present study was designed to investigate whether L-arginine (Arg) supplementation would influence the production performance and reproductive traits of laying hens fed low crude protein (LCP) diets. Hy-Line brown laying hens were randomly assigned to dietary treatments of control, LCP, and LCP supplemented with 0.05, 0.10, 0.15, or 0.20% Arg for 7 wk. The results showed no significant variations on the production performance, and relative organ indexes of hens. However, significant transcriptional and structural changes were evident along the HPG axis of hens. Arg supplementation linearly upregulated (P < 0.05) the gonadotropin-releasing hormone 1 (GnRH1), and gonadotropin inhibitory hormone in the hypothalamus. The pituitary growth hormone, GnRH receptor, and follicle-stimulating hormone (FSHß) were also increased (P < 0.05). In the ovary, GnRH1, and estrogen receptor ß were linearly increased by Arg, and the ovarian morphology revealed that LCP induced structural alterations which were minimally recovered by Arg supplementation. Serum insulin-like growth factor 1 (IGF-1) and nitric oxide (NO) were increased (P < 0.05) at higher levels of Arg supplementation. Therefore, supplementing high Arg (0.20%) to LCP hens influenced the ovarian morphology and modulated the gene expression of reproductive hormones in the hypothalamic-pituitary-gonadal axis of laying hens via actions that may be related to NO and IGF-1 activity.

Encapsulated crystalline lysine and DL-methionine have higher efficiency than the crystalline form in broilers.

Crystalline amino acids (AAs) exhibit high nutritional values when supplemented AA-deficient diets. However, the AAs in crystalline form in the diet are absorbed quickly than protein-bound AAs, which may take an effect on AA utilization efficiency. In this study, 2 experiments were conducted to investigate the effect of encapsulated lysine-HCl (Lys) and DL-methionine (DL-Met) on the growth performance of broiler chickens. In experiment 1, a total of 432 one-day-old male Arbor Acres broilers were subjected to 3 dietary treatments (27 pens; 16 birds per pen) for 42 d. The control group was basal diets supplemented with crystalline Lys and DL-Met, and treatment groups had basal diets supplemented with encapsulated Lys and DL-Met at the levels of 80% and 60% of control diets (80CLM, 60CLM), respectively. The growth performance, intestinal development, and transcription of AA transporters were determined. In experiment 2, 24 broiler chickens were subjected to the same treatments as in experiment 1. The plasma concentrations of free AAs were measured 0, 2, 4, and 6 h after feeding. The results showed that 80CLM treatment had no significant influence on production performance, carcass characteristics, and plasma free AAs content during the experiment compared with the control group (P > 0.05). In addition, the 80CLM group moderately enhanced gut morphology development and increased AAs' absorption capacity. However, broilers fed the 60CLM diet had lower production performance and breast muscle weight than the control group (P < 0.05), but increased villi height and B0AT mRNA expression level (P < 0.05). At h 4 after feeding, the 60CLM broilers exhibited higher concentration of Ala, Cys, and total dispensable AAs than the control group (P < 0.05). In conclusion, the result suggests that the supplemental levels of crystalline Lys and DL-Met can be effectively saved approximately for 20% by using the encapsulated form in broilers, with improvements to AAs utilization efficiency, while posing no detrimental effects on production performance. Encapsulated Lys and DL-Met would have greater potential for application when replacing crystalline AAs in broiler chickens.

Use of encapsulated L-lysine-HCl and DL-methionine improves postprandial amino acid balance in laying hens.

The supplementation of dietary limiting amino acids (AA) with crystalline AA makes the use of low-protein diets an option in poultry production. The differing absorption rates of crystalline and protein-bound AA may lead to temporally imbalanced AA in the postabsorptive period. In this study, two experiments were conducted to evaluate the effect of encapsulated L-lysine-HCl (L-Lys-HCl) and DL-methionine (DL-Met) on the laying performance of hens. In exp. 1, a total of 135 forty-seven-wk-old Hy-Line Brown hens were subjected to three dietary treatments for 8 wk: basal diet supplemented with 0.14% L-Lys-HCl and 0.17% DL-Met to satisfy the NRC (1994) total Lys and Met recommendation (control) and basal diet supplemented with encapsulated L-Lys-HCl and DL-Met at the levels of 60% (60CLM, 0.084% L-Lys-HCl and 0.102% DL-Met) or 80% of control (80CLM, 0.112% L-Lys-HCl and 0.136% DL-Met), respectively. In exp. 2, 24 fifty-five-wk-old Hy-Line Brown hens were individually reared in cages and subjected to the same treatments as in exp. 1. The plasma concentrations of free AA and nitrogen metabolites were measured 2, 4, and 6 h after fed. The results showed that dietary AA treatment had no significant influence on body weight (BW), feed intake, laying rate, egg weight, egg mass, or feed efficiency. The expression levels of AA transporters CAT-1, y+LAT1, b0,+AT, B0AT, rBAT, EAAT3, and PepT1 in the duodenum, jejunum, and ileum were not influenced (P > 0.05) by dietary treatment. There was an interaction of dietary AA treatment and time (P < 0.05) and the 80CLM hens exhibited higher concentrations of Lys (P < 0.05) than the controls at 2-h time point. In contrast, plasma Met concentration was not influenced (P > 0.05), while Cys was reduced in the 60CLM hens at every time point. The 80CLM hens had higher taurine concentrations than those receiving the control diet at every postprandial time point. In conclusion, these findings demonstrate that by using encapsulated form, the supplemental levels of synthetic L-Lys-HCl and DL-Met can be effectively reduced by approximately 20% with no negative effect on laying performance. The result suggests that encapsulated Lys and Met may ameliorate the postabsorptive AA balance and contribute to the reduced dietary AA supplemental levels.

Comparison of leaf transcriptome in response to Rhizoctonia solani infection between resistant and susceptible rice cultivars.

BACKGROUND: Sheath blight (SB), caused by Rhizoctonia solani, is a common rice disease worldwide. Currently, rice cultivars with robust resistance to R. solani are still lacking. To provide theoretic basis for molecular breeding of R. solani-resistant rice cultivars, the changes of transcriptome profiles in response to R. solani infection were compared between a moderate resistant cultivar (Yanhui-888, YH) and a susceptible cultivar (Jingang-30, JG). RESULTS: In the present study, 3085 differentially express genes (DEGs) were detected between the infected leaves and the control in JG, with 2853 DEGs in YH. A total of 4091 unigenes were significantly upregulated in YH than in JG before infection, while 3192 were significantly upregulated after infection. Further analysis revealed that YH and JG showed similar molecular responses to R. solani infection, but the responses were earlier in JG than in YH. Expression levels of trans-cinnamate 4-monooxygenase (C4H), ethylene-insensitive protein 2 (EIN2), transcriptome factor WRKY33 and the KEGG pathway plant-pathogen interaction were significantly affected by R. solani infection. More importantly, these components were all over-represented in YH cultivar than in JG cultivar before and/or after infection. CONCLUSIONS: These genes possibly contribute to the higher resistance of YH to R. solani than JG and were potential target genes to molecularly breed R. solani-resistant rice cultivar.

Impacts of soil salinity on Bt protein concentration in square of transgenic Bt cotton.

Insect-resistance of transgenic Bacillus thuringiensis (Bt) cotton varies among plants organs and with different environmental conditions. The objective of this study was to examine the influence of soil salinity on Bt protein concentration in cotton squares and to elucidate the potential mechanism of Bt efficacy reduction. Two cotton cultivars (NuCOTN 33B and CCRI 07, salt-sensitive and salt-tolerant) were subjected to salinity stress under four natural saline levels in field conditions in 2015 and 2016 and seven regimes of soil salinity ranged from 0.5 to 18.8 dS m-1 in greenhouse conditions in 2017. Results of field studies revealed that Bt protein content was not significantly changed at 7.13 dS m-1 salinity, but exhibited a significant drop at the 10.41 and 14.16 dS m-1 salinity. The greenhouse experiments further showed similar trends that significant declines of the insecticidal protein contents in squares were detected when soil salinity exceeded 9.1 dS m-1. Meanwhile, high salinity resulted in significant reduction in contents of soluble protein and total nitrogen, activities of nitrate reductase (NR), glutamine synthetase (GS) and glutamic-pyruvic transaminase (GPT), but increased amino acid content, activities of protease and peptidase in cotton squares. High salinity also decreased root vigor (RV), root total absorption area (RTA) and root active absorption area (RAA). The extent of decrease of Bt protein content was more pronounced in NuCOTN 33B than CCRI 07, and CCRI07 exhibited stronger enzymes activities involved in square protein synthesis and higher levels of RV, RTA and RAA. Therefore, the results of our present study indicated that insecticidal protein expression in cotton squares were significantly affected by higher salinity (equal to or higher than 9.1 dS m-1), reduced protein synthesis and increased protein degradation in squares and reduced metabolic activities in roots might lead to the decrease of Bt protein content in squares.

Characterization of a new semi-dominant dwarf allele of SLR1 and its potential application in hybrid rice breeding.

The widespread introduction of semi-dwarf1 (sd1), also known as the 'Green Revolution' gene, has dramatically increased rice yield. However, the extensive use of limited sources of dwarf genes may cause 'bottleneck' effects in breeding new rice varieties. Alternative dwarf germplasms are quite urgent for rice breeding. Here, we characterized a new allele of the rice Slr1-d mutant, Slr1-d6, which reduced plant height by 37%, a much milder allele for dwarfism. Slr-d6 was still responsive to gibberellin (GA) to a reduced extent. The mutation site in Slr1-d6 was less conserved in the TVHYNP domain, leading to the specific semi-dominant dwarf phenotype. Expression of SLR1 and five key GA biosynthetic genes was disturbed in Slr1-d6, and the interaction between Slr1-d6 and GID1 was decreased. In the genetic background of cultivar 9311 with sd1 eliminated, Slr1-d6 homozygous plants were ~70 cm tall. Moreover, Slr1-d6 heterozygous plants were equivalent in height to the standard sd1 semi-dwarf 9311, but with a 25% yield increase, showing its potential application in hybrid rice breeding.

Semi-Rolled Leaf2 modulates rice leaf rolling by regulating abaxial side cell differentiation.

Moderate leaf rolling maintains the erectness of leaves and minimizes the shadowing between leaves which is helpful to establish ideal plant architecture. Here, we describe asrl2(semi-rolled leaf2) rice mutant, which has incurved leaves due to the presence of defective sclerenchymatous cells on the abaxial side of the leaf and displays narrow leaves and reduced plant height. Map-based cloning revealed that SRL2 encodes a novel plant-specific protein of unknown biochemical function.SRL2 was mainly expressed in the vascular bundles of leaf blades, leaf sheaths, and roots, especially in their sclerenchymatous cells. The transcriptional activities of several leaf development-related YABBY genes were significantly altered in the srl2 mutant. Double mutant analysis suggested that SRL2 and SHALLOT-LIKE1(SLL1)/ROLLED LEAF9(RL9) function in distinct pathways that regulate abaxial-side leaf development. Hence, SRL2 plays an important role in regulating leaf development, particularly during sclerenchymatous cell differentiation.

Manipulation of the rice L-galactose pathway: evaluation of the effects of transgene overexpression on ascorbate accumulation and abiotic stress tolerance.

Ascorbic acid (AsA) is the most abundant water-soluble antioxidant in plants, and it plays a crucial role in plant growth, development and abiotic stress tolerance. In the present study, six key Arabidopsis or rapeseed genes involved in AsA biosynthesis were constitutively overexpressed in an elite Japonica rice cultivar. These genes encoded the GDP-mannose pyrophosphorylase (GMP), GDP-mannose-3',5'-epimerase (GME), GDP-L-galactose phosphorylase (GGP), L-galactose-1-phosphate phosphatase (GPP), L-galactose dehydrogenase (GDH), and L-galactono-1,4-lactone dehydrogenase (GalLDH). The effects of transgene expression on rice leaf AsA accumulation were carefully evaluated. In homozygous transgenic seedlings, AtGGP transgenic lines had the highest AsA contents (2.55-fold greater than the empty vector transgenic control), followed by the AtGME and AtGDH transgenic lines. Moreover, with the exception of the AtGPP lines, the increased AsA content also provoked an increase in the redox state (AsA/DHA ratio). To evaluate salt tolerance, AtGGP and AtGME transgenic seedlings were exposed to salt stress for one week. The relative plant height, root length and fresh weight growth rates were significantly higher for the transgenic lines compared with the control plants. Altogether, our results suggest that GGP may be a key rate-limiting step in rice AsA biosynthesis, and the plants with elevated AsA contents demonstrated enhanced tolerance for salt stress.

The use of microdebrider for the treatment of accessory axillary breast.

Accessory axillary breast tissue can be fairly common occurring in 2-6% of women. Treatment modalities thus far include direct excision as well as liposuction. While direct excision allows for accessible and adequate tissue resection, it results in long unsightly scars and the creation of significant amount of dead space. This may be complicated by seroma and haematoma formation. Liposuction is not without its drawbacks either. It is often very difficult to remove fibro-glandular breast tissue resulting in inadequate excision, thus leaving behind a visible core of breast tissue. This has led some surgeons to use a combination of direct excision and liposuction to manage accessory axillary breast tissue. Hence, we present the use of the microdebrider for sharp and precise excision of accessory axillary breast tissue. This day procedure can be performed through a single 5-mm incision which is well hidden in the axillary skin folds while allowing the operator the amount of control needed to accurately remove fibro-glandular breast tissue and restore an aesthetically pleasing contour to the axilla.