Proline metabolism as a mechanism for the energy dissipation in VaP5CSF129A transgenic tobacco plants under water deficit.

In plants, proline accumulation in cells is a common response to alleviate the stress caused by water deficits. It has been shown that foliar proline spraying, as well as its overaccumulation in transgenic plants can increase drought tolerance, as proline metabolism plays important roles in cell redox balance and on energy dissipation pathways. The aim of this work was to evaluate the role of exogenous proline application or its endogenous overproduction as a potential mechanism for energy dissipation. For this, wild-type and VaP5CSF129A transgenic tobacco plants were sprayed with proline (10 mM) and submitted to water deficit. Changes in plant physiology and biochemistry were evaluated. Transcriptional changes in the relative expression of genes involved in proline synthesis and catabolism, NAD (P)-dependent malate dehydrogenase (NAD(P)-MDH), alternative oxidase (AOX), and VaP5CSF129A transgene were measured. Exogenous proline reduced the negative effects of water deficit on photosynthetic activity in both genotypes; with the transgenic plants even less affected. Water deficit caused an increase in the relative expression of proline biosynthesis genes. On the other hand, the expression of catabolism genes decreased, primarily in transgenic plants. Exogenous proline reduced activity of the NADP-MDH enzyme and decreased expression of the AOX and NADP-MDH genes, mainly in transgenic plants under water stress. Finally, our results suggest that proline metabolism could act as a complementary/compensatory mechanism for the energy dissipation pathways in plants under water deficit.

Glutamine, proline, and isoleucine support maturation and fertilisation of bovine oocytes.

Successful in-vitro production of bovine embryos relies on meiotic maturation of oocytes in vitro (IVM) before they can be fertilised. High levels of IVM are currently achieved using a complex medium that contains all 20 common amino acids, namely TCM199, but can also be achieved using a simple inorganic salt solution containing non-essential amino acids, proline, and glutamine. Further simplification of the amino acid content of medium used for IVM could lead to a more defined medium that provides reproducible IVM. The aim of this study was, therefore, to determine the minimal amino acid requirements for bovine oocyte nuclear maturation, as measured by progression to metaphase II (MII) of meiosis. Supplementation of a simple medium composed of inorganic salts (M1 medium) with multiple amino-acid combinations showed that M1 containing glutamine, proline, and isoleucine resulted in nuclear maturation comparable to that of TCM199 (57.4 ± 3.4% vs 67% ± 1.7%, respectively) but was reduced when cystine (Cys2) to that seen with M1 alone (38.0 ± 2.2%). Viability of oocytes matured in this simplified medium was equal to those matured in TCM199 since the same proportion of zygotes with 2 pronuclei were observed following fertilisation in medium containing no amino acids (33.9 ± 6.5% vs 33.3 ± 3.6%, respectively). Addition of glutamine, proline and isoleucine to fertilisation medium also increased the proportion of zygotes but did not increase blastocyst development rates. Thus, a defined medium containing only glutamine, proline and isoleucine is sufficient for oocyte maturation and successful fertilisation.

Trans-Zeatin induce regulation the biosynthesis of 2-acetyl-1-pyrroline in fragrant rice (Oryza sativa L.) seedlings.

BACKGROUND: In plants, cytokinin is activated into trans-zeatin to fight abiotic stresses. However, the mechanism of the effect of trans-zeatin on 2-acetyl-1-pyrroline (2-AP) biosynthesis in fragrant rice has yet to be studied. The present study was conducted to explore the effects of exogenous trans-zeatin on enzymes activities, genes expression, and precursors involved in 2-AP biosynthesis and 2-AP contents as well as the seedling quality of a fragrant rice cultivar viz., Meixiangzhan2. Four concentrations of trans-zeatin solutions at 20, 40, and 80 µmol L- 1 (ZT1, ZT2, and ZT3) were sprayed onto rice seedlings. RESULTS: Compared to the control, trans-zeatin treatments showed significantly higher 2-AP contents of fragrant rice seedlings. Increased plant height and stem width were observed due to trans-zeatin treatments. The trans-zeatin application increased 1-pyrroline, methylglyoxal, proline, and P5C contents, enhanced P5CS and OAT activities, and reduced glutamic acid contents. In addition, expressions of ProDH, P5CS2, and DAO4 were comparatively higher under trans-zeatin treatments than CK in fragrant rice seedlings. CONCLUSIONS: Overall, up-regulation of P5C, 1-pyrroline, and proline and down-regulation of glutamic acid under appropriate trans-zeatin concentrations (20 and 40 µmol L- 1) resulted in enhanced 2-AP biosynthesis in fragrant rice seedlings and 20-40 µmol L- 1 was considered as the suggested concentrations of trans-zeatin application in fragrant rice seedling.

Global Responses of Autopolyploid Sugarcane Badila (Saccharum officinarum L.) to Drought Stress Based on Comparative Transcriptome and Metabolome Profiling.

Sugarcane (Saccharum spp. hybrid) is frequently affected by seasonal drought, which causes substantial declines in quality and yield. To understand the drought resistance mechanisms of S. officinarum, the main species of modern sugarcane, at a molecular level, we carried out a comparative analysis of transcriptome and metabolome profiling of the sugarcane variety Badila under drought stress (DS). Compared with control group (CG) plants, plants exposed to DS had 13,744 (6663 up-regulated and 7081 down-regulated) differentially expressed genes (DEGs). GO and KEGG analysis showed that the DEGs were enriched in photosynthesis-related pathways and most DEGs had down-regulated expression. Moreover, the chlorophyll content, photosynthesis (Photo), stomatal conductance (Cond), intercellular carbon dioxide concentration (Ci) and transpiration rate (Trmmol) were sharply decreased under DS. These results indicate that DS has a significant negative influence on photosynthesis in sugarcane. Metabolome analysis identified 166 (37 down-regulated and 129 up-regulated) significantly regulated metabolites (SRMs). Over 50% of SRMs were alkaloids, amino acids and their derivatives, and lipids. The five most significantly enriched KEGG pathways among SRMs were Aminoacyl-tRNA biosynthesis, 2-Oxocarboxylic acid metabolism, Biosynthesis of amino acids, Phenylalanine metabolism, and Arginine and proline metabolism (p < 0.05). Comparing CG with DS for transcriptome and metabolome profiling (T_CG/DS and M_CG/DS, respectively), we found three of the same KEGG-enriched pathways, namely Biosynthesis of amino acids, Phenylalanine metabolism and Arginine and proline metabolism. The potential importance of Phenylalanine metabolism and Arginine and proline metabolism was further analyzed for response to DS in sugarcane. Seven SRMs (five up-regulated and two down-regulated) and 60 DEGs (17 up-regulated and 43 down-regulated) were enriched in Phenylalanine metabolism under DS, of which novel.31261, Sspon.04G0008060-1A, Sspon.04G0008060-2B and Sspon.04G0008060-3C were significantly correlated with 7 SRMs. In Arginine and proline metabolism, eight SRMs (seven up-regulated and one down-regulated) and 63 DEGs (32 up-regulated and 31 down-regulated) were enriched, of which Sspon.01G0026110-1A (OAT) and Sspon.03G0002750-3D (P5CS) were strongly associated with proline (r > 0.99). These findings present the dynamic changes and possible molecular mechanisms of Phenylalanine metabolism as well as Arginine and proline metabolism under DS and provide a foundation for future research and sugarcane improvement.

Application of glutamic acid improved As tolerance in aromatic rice at early growth stage.

To alleviate the arsenic (As) toxicity in aromatic rice, a hydroponic experiment of two As concentrations (0 and 100 µM sodium arsenite: A0, A1), three glutamic acid (Glu) concentrations (0, 100, and 500 µM l-glutamic acid: G0, G1, and G2) with Xiangyaxiangzhan and Meixiangzhan 2 was conducted. Results showed that the root As content were increased under A1G2 but reduced under A1G1 for Xiangyaxiangzhan as compared with A1G0. A decrement of As was transported from root to shoot caused by up-regulated OsABCC1 relative expression in Meixiangzhan 2. Likewise, As stress enhanced the H2O2 and malondialdehyde content, resulting in the impaired cell wall observed by transmission electron microscopy. However, compared with A1G0, the superoxide dismutase activity, ascorbic acid, glutathione, proline, and soluble sugar content were increased under A1G1. Additionally, arsenate reductase, monodehydroascorbate reductase activity, Glu, proline, and soluble sugar content were found positively associated with the As accumulation. Further, the metabolome analysis indicated that the pathway of amino acid and arginine biosynthesis were notably enriched after Glu application. Generally, 100 µM Glu application was the better treatment to enhance As tolerance in aromatic rice through up-regulating amino acid biosynthesis with increasing antioxidants and osmolytes to scavenge excessive reactive oxygen species.

Do Amino Acid Antiporters Have Asymmetric Substrate Specificity?

Amino acid antiporters mediate the 1:1 exchange of groups of amino acids. Whether substrate specificity can be different for the inward and outward facing conformation has not been investigated systematically, although examples of asymmetric transport have been reported. Here we used LC-MS to detect the movement of 12C- and 13C-labelled amino acid mixtures across the plasma membrane of Xenopus laevis oocytes expressing a variety of amino acid antiporters. Differences of substrate specificity between transporter paralogs were readily observed using this method. Our results suggest that antiporters are largely symmetric, equalizing the pools of their substrate amino acids. Exceptions are the antiporters y+LAT1 and y+LAT2 where neutral amino acids are co-transported with Na+ ions, favouring their import. For the antiporters ASCT1 and ASCT2 glycine acted as a selective influx substrate, while proline was a selective influx substrate of ASCT1. These data show that antiporters can display non-canonical modes of transport.

The proline-rich domain of fission yeast WASp (Wsp1p) interacts with actin filaments and inhibits actin polymerization.

Members of the Wiskott-Aldrich Syndrome protein (WASp) family activate Arp2/3 complex (actin-related proteins 2 and 3 complex) to form actin filament branches. The proline-rich domain (PRD) of WASp contributes to branching nucleation, and the PRD of budding yeast Las17 binds actin filaments [Urbanek AN et al. (2013) Curr Biol 23, 196-203]. Biochemical assays showed the recombinant PRD of fission yeast Schizosaccharomyces pombe Wsp1p binds actin filaments with micromolar affinity. Recombinant PRDs of both Wsp1p and Las17p slowed the elongation of actin filaments by Mg-ATP-actin monomers by half and slowed the spontaneous polymerization of Mg-ATP-actin monomers modestly. The affinity of PRDs of WASp-family proteins for actin filaments is high enough to contribute to the reported stimulation of actin filament branching by Arp2/3 complex.

Crystal Structure of the SH3 Domain of ASAP1 in Complex with the Proline Rich Motif (PRM) of MICAL1 Reveals a Unique SH3/PRM Interaction Mode.

SH3 domains are common protein binding modules. The target sequence of SH3 domains is usually a proline-rich motif (PRM) containing a minimal "PxxP" sequence. The mechanism of how different SH3 domains specifically choose their targets from vast PxxP-containing sequences is still not very clear, as many reported SH3/PRM interactions are weak and promiscuous. Here, we identified the binding of the SH3 domain of ASAP1 to the PRM of MICAL1 with a sub-µM binding affinity, and determined the crystal structure of ASAP1-SH3 and MICAL1-PRM complex. Our structural and biochemical analyses revealed that the target-binding pocket of ASAP1-SH3 contains two negatively charged patches to recognize the "xPx + Px+" sequence in MICAL1-PRM and consequently strengthen the interaction, differing from the typical SH3/PRM interaction. This unique PRM-binding pocket is also found in the SH3 domains of GTPase Regulator associated with focal adhesion kinase (GRAF) and Src kinase associated phosphoprotein 1 (SKAP1), which we named SH3AGS. In addition, we searched the Swiss-Prot database and found ~130 proteins with the SH3AGS-binding PRM in silico. Finally, gene ontology analysis suggests that the strong interaction between the SH3AGS-containing proteins and their targets may play roles in actin cytoskeleton regulation and vesicle trafficking.

Yield, quality and physiological variation of strawberry in response to irrigation regimes and exogenous proline with a cost benefit analysis.

Strawberry (Fragaria × ananassa Duch.) production is a major aspect of the agricultural economy in Turkey's Mediterranean region, offering high rates of employment and farm revenue. The effects of treatment of the exogenous amino acid on yield, quality, and physiological diversity for strawberry production was analyzed considering the economic aspects. To achieve this, 4 different irrigation regimes (IR (irrigation) 125, IR100, IR75, IR50) with proline treatment was tested. The total berry yield, photosynthesis and leaf water potential (LWP) significantly decreased as irrigation application rates declined. The IR125 treatment with proline (PIR125) produced the highest overall berry production. The use of proline significantly increased berry production by 23%. Exogenous proline generated 3.5 kg/1 m3 greater yields than control in terms of irrigation water use efficiency (IWUE). Under the IR50 conditions, the treatment of proline yielded a 32% higher than the control. Moreover, proline significantly increased fruit soluble solids content (SSC) by 6.4%. The production system achieves the highest cost-benefit ratio (CBR) under PIR125 whereas the lowest cost-benefit ratio under IR50. When each scenario was considered at individually, it was evident that the more water used the more efficient high tunnel strawberry production. The fact that proline generated an increase in CBR in all four irrigation regimes proves this amino acid's economic effectiveness. As a result, PIR125 is recommended for the highest efficiency and CBR in Mediterranean environment. However, it has been discovered that using proline to alleviate this problem in areas where water is limited could be quite helpful.

Molecular mechanism of Tsukamurella tyrosinosolvens strain P9 in response to root exudates of peanut.

Tsukamurella tyrosinosolvens strain P9 is a rare actinomycete with plant growth-promoting properties and can improve the growth of peanut. We analyzed the differentially expressed genes (DEGs) of P9 under the influence of peanut root exudates from RNA-sequencing data and analyzed the effects of root exudates and their organic acid and amino acid components on the growth and growth-promoting effects of this strain to explore the molecular mechanism of the P9 response. The results showed that peanut root exudates promoted the growth and growth-promoting activity of P9. Transcriptome analysis revealed 126 DEGs in P9, comprising 81 up-regulated and 45 down-regulated genes. The DEGs were significantly enriched in 17 KEGG metabolic pathways, including arginine biosynthesis, butyric acid metabolism, fatty acid degradation, and tryptophan metabolism. Peanut root exudates induced up-regulation of nutrient transport, carbohydrate metabolism and energy production, siderophore and IAA biosynthesis, adhesion, and biofilm formation, and down-regulation of arginine biosynthesis and the urea cycle in P9. Organic acids and amino acids are the major components of peanut root exudates. Glycine, proline, and alanine promoted the growth and IAA secretion of P9. Proline, alanine (40 mM), and oxalic acid significantly enhanced siderophore biosynthesis, whereas citric acid, oxalic acid, and malic acid significantly promoted biofilm formation of P9. This study clarifies the response of T. tyrosinosolvens P9 to peanut root exudates at the molecular level, examining the molecular basis of the relationship between P9 and peanut, and provides a theoretical foundation for improved exertion of the growth-promoting properties of P9.

NKCC transport mediates the insulinotropic effects of taurine and other small neutral amino acids.

AIMS: Despite its high concentration in pancreatic islets of Langerhans and broad range of antihyperglycemic effects, the route facilitating the import of dietary taurine into pancreatic ß-cell and mechanisms underlying its insulinotropic activity are unclear. We therefore studied the impact of taurine on beta-cell function, alongside that of other small neutral amino acids, L-alanine and L-proline. MAIN METHODS: Pharmacological profiling of insulin secretion was conducted using clonal BRIN BD11 ß-cells, the impact of taurine on the metabolic fate of glucose carbons was assessed using NMR and the findings were verified by real-time imaging of Ca2+ dynamics in the cytosol of primary mouse and human islet beta-cells. KEY FINDINGS: In our hands, taurine, alanine and proline induced secretory responses that were dependent on the plasma membrane depolarisation, import of Ca2+, homeostasis of K+ and Na+ as well as on cell glycolytic and oxidative metabolism. Taurine shifted the balance between the oxidation and anaplerosis towards the latter, in BRIN BD11 beta-cells. Furthermore, the amino acid signalling was significantly attenuated by inhibition of Na+-K+-Cl- symporter (NKCC). SIGNIFICANCE: These data suggest that taurine, like L-alanine and L-proline, acutely induces glucose-dependent insulin-secretory responses by modulating electrogenic Na+ transport, with potential role of intracellular K+ and Cl- in the signal transduction. The acute action delineated would be consistent with antidiabetic potential of dietary taurine supplementation.

FK506-Binding Protein 2 Participates in Proinsulin Folding.

Apart from chaperoning, disulfide bond formation, and downstream processing, the molecular sequence of proinsulin folding is not completely understood. Proinsulin requires proline isomerization for correct folding. Since FK506-binding protein 2 (FKBP2) is an ER-resident proline isomerase, we hypothesized that FKBP2 contributes to proinsulin folding. We found that FKBP2 co-immunoprecipitated with proinsulin and its chaperone GRP94 and that inhibition of FKBP2 expression increased proinsulin turnover with reduced intracellular proinsulin and insulin levels. This phenotype was accompanied by an increased proinsulin secretion and the formation of proinsulin high-molecular-weight complexes, a sign of proinsulin misfolding. FKBP2 knockout in pancreatic ß-cells increased apoptosis without detectable up-regulation of ER stress response genes. Interestingly, FKBP2 mRNA was overexpressed in ß-cells from pancreatic islets of T2D patients. Based on molecular modeling and an in vitro enzymatic assay, we suggest that proline at position 28 of the proinsulin B-chain (P28) is the substrate of FKBP2's isomerization activity. We propose that this isomerization step catalyzed by FKBP2 is an essential sequence required for correct proinsulin folding.

Regulation of chromium translocation to shoot and physiological, metabolomic, and ionomic adjustments confer chromium stress tolerance in the halophyte Suaeda maritima.

Chromium (Cr) is a highly toxic element adversely affecting the environment, cultivable lands, and human populations. The present study investigated the effects of Cr (VI) (100-400 µM) on plant morphology and growth, photosynthetic pigments, organic osmolytes, ionomics, and metabolomic dynamics of the halophyte Suaeda maritima to decipher the Cr tolerance mechanisms. Cr exposure reduced the growth and biomass in S. maritima. The photosynthetic pigments content significantly declined at higher Cr concentrations (400 µM). However, at lower Cr concentrations (100-300 µM), the photosynthetic pigments remained unaffected or increased. The results suggest that a high concentration of Cr exposure might have adverse effects on PS II in S. maritima. The enhanced uptake of Na+ in S. maritima imposed to Cr stress indicates that Na+ might have a pivotal role in osmotic adjustment, thereby maintaining water status under Cr stress. The proline content was significantly upregulated in Cr-treated plants suggesting its role in maintaining osmotic balance and scavenging ROS. The metabolomic analysis of control and 400 µM Cr treated plants led to the identification of 62 metabolites. The fold chain analysis indicated the upregulation of several metabolites, including phytohormones (SA and GA3), polyphenols (cinnamic acid, sinapic acid, coumaric acid, vanillic acid, and syringic acid), and amino acids (alanine, leucine, proline, methionine, and cysteine) under Cr stress. The upregulation of these metabolites suggests the enhanced metal chelation and sequestration in vacuoles, reducing oxidative stress by scavenging ROS and promoting photosynthesis by maintaining the chloroplast membrane structure and photosynthetic pigments. Furthermore, in S. maritima, Cr tolerance index (Ti) was more than 60% in all the treatments, and Cr bio-concentration factor (BCF) and translocation factor (Tf) values were all greater than 1.0, which clearly indicates the Cr-hyperaccumulator characteristics of this halophyte.

Evolutionary Analysis of StSnRK2 Family Genes and Their Overexpression in Transgenic Tobacco Improve Drought Tolerance.

Sucrose non-ferment 1-related protein kinase 2 (SnRK2) is a highly conserved protein kinase in plants that plays an important role in regulating plant response to drought stress. Although it has been reported in some plants, the evolutionary relationship of potato SnRK2s and their function in drought resistance have not been systematically analyzed. In this study, molecular characteristic analysis showed that 8 StSnRK2s were distributed on six chromosomes, coding proteins were divided into three subgroups, and StSnRK2s clustered in the same subgroup had similar conserved motifs and domains. In addition, StSnRK2 has a wide range of replication events in some species, making it closer to dicots in the process of evolution. In addition, the average nonsynonymous substitution rate/synonymous substitution rate (Ka/Ks) value of SnRK2s in monocots was higher than that of dicots. The codon usage index showed that SnRK2s prefer to use cytosine 3 (C3s), guanine 3 (G3s) and GC content (GC3s) in monocots, whereas thymine 3 (T3s) and adenine 3 (A3s) are preferred in dicots. Furthermore, stress response analysis showed that the expression of StSnRK2s under different degrees of drought stress significantly correlated with one or more stress-related physiological indices, such as proline and malondialdehyde (MDA) content, superoxide dismutase (SOD) and catalase (CAT) activity, ion leakage (IL) etc. The drought resistance of StSnRK2 transgenic plants was determined to occur in the order of StSnRK2.1/2.8 > StSnRK2.2/2.5 > StSnRK2.4/2.6 > StSnRK2.3 > StSnRK2.7, was attributed to not only lower IL but also higher proline, soluble sugar contents and stress-related genes in transgenic plants compared to wild type (WT). In conclusion, this study provides useful insights into the evolution and function of StSnRK2s and lays a foundation for further study on the molecular mechanism of StSnRK2s regulating potato drought resistance.

Triticum aestivum: antioxidant gene profiling and morpho-physiological studies under salt stress.

BACKGROUND: Soil salinity drastically reduced wheat growth and production in Pakistan. It is a need of an hour to identify the best suitable salt tolerance or resistant wheat varieties which shows good growth under salinity affected areas. In presented study, two wheat varieties Johar (salt tolerant) and Sarsabaz (salt sensitive) were examined under NaCl stress conditions. METHODS: Antioxidant enzyme activities were investigated in 10-days old wheat seedlings under 200 mM NaCl stress in hydroponic conditions. To investigate the various growth parameters, antioxidant enzyme activities such as superoxide dismutase (SOD: EC 1.15.1.1), catalase (CAT: EC 1.11.1.6) and ascorbate peroxidase (APX: EC 1.11.1.11) were monitored and studied. Besides this various growth parameters such as length of the roots, shoots, as well as Physiological parameters likes lipid peroxidation by malondialdehyde (MDA), hydrogen peroxide (H2O2), and proline contents and antioxidant enzyme activities were estimated. The effect of salinity was also observed on gene transcription level and eventually expression level. RESULTS: Shoot and root length were decreased in Sarsabaz variety while it showed opposite trend in johar at 200 mM salt concentration. The concentration of proline showed a noticeable rise in salt dependency. Higher concentrations of Proline in Johar were observed as compared to Sarsabaz. SOD showed the increase in activity for antioxidant enzymes. Significant increase of SOD levels were observed in shoot tissues as compared to root tissues. The results indicated that the shoots were more susceptible to salt stress. Activity of APX showed similar affects in both varieties. The production of CAT enzyme in the shoot and root tissues of both varieties showed substantial growth under increased salt stress. Furthermore, NaCl stress has increased the expression of certain genes coding for antioxidant enzymes such as catalase, superoxide dismutase, and peroxidase. Maximum expression of all the antioxidant enzyme coding genes were observed in Johar (tolerant) at 48 h exposure to salt. In contrast the expression of the all mentioned genes in Sarsabaz variety were found maximum at early hours (24 h) and gradually decreased at 48 h. CONCLUSION: The study showed that the selected salt tolerant wheat variety Johar is significantly resistant to 200 mM NaCl salt level as compared to Sarsabaz.

Zebrafish and medaka T1R (taste receptor type 1) proteins mediate highly sensitive recognition of l-proline.

In vertebrates, nutritional tastants, such as amino acids and sugars, are recognized by G-protein-coupled receptors of the taste receptor type 1 (T1R) family. Previous studies have shown that fish T1Rs are functionally distinct from mammalian T1Rs in certain regards. Here, we report the existence of oral receptors with high sensitivity to amino acids in zebrafish and medaka fish. We describe the construction of multiple cell lines stably expressing functional T1Rs (from medaka fish or zebrafish) with a chimeric G-protein (G16gust44) using the Flp-In system. Through functional assays with these cell lines, medaka fish and zebrafish were confirmed to possess particular T1Rs highly sensitive to l-proline, possibly reflecting the physiological importance of l-proline in teleosts, in line with previous studies.

The impact of glucose on mitochondria and life span is determined by the integrity of proline catabolism in Caenorhabditis elegans.

Mutations in genes involved in mitochondrial proline catabolism lead to the rare genetic disorder hyperprolinemia in humans. We have previously reported that mutations of proline catabolic genes in Caenorhabditis elegans impair mitochondrial homeostasis and shorten life span, and that these effects surprisingly occur in a diet type-dependent manner. Therefore, we speculated that a specific dietary component may mitigate the adverse effects of defective proline catabolism. Here, we discovered that high dietary glucose, which is generally detrimental to health, actually improves mitochondrial homeostasis and life span in C. elegans with faulty proline catabolism. Mechanistically, defective proline catabolism results in a shift of glucose catabolism toward the pentose phosphate pathway, which is crucial for cellular redox balance. This shift helps to maintain mitochondrial reactive oxygen species homeostasis and to extend life span, as suppression of the pentose phosphate pathway enzyme GSPD-1 prevents the favorable effects of high glucose. In addition, we demonstrate that this crosstalk between proline and glucose catabolism is mediated by the transcription factor DAF-16. Altogether, these findings suggest that a glucose-rich diet may be advantageous in certain situations and might represent a potentially viable treatment strategy for disorders involving impaired proline catabolism.

TaERF87 and TaAKS1 synergistically regulate TaP5CS1/TaP5CR1-mediated proline biosynthesis to enhance drought tolerance in wheat.

Drought stress limits wheat production and threatens food security world-wide. While ethylene-responsive factors (ERFs) are known to regulate plant response to drought stress, the regulatory mechanisms responsible for a tolerant phenotype remain unclear. Here, we describe the positive regulatory role of TaERF87 in mediating wheat tolerance to drought stress. TaERF87 overexpression (OE) enhances drought tolerance, while silencing leads to drought sensitivity in wheat. RNA sequencing with biochemical assays revealed that TaERF87 activates the expression of the proline biosynthesis genes TaP5CS1 and TaP5CR1 via direct binding to GCC-box elements. Furthermore, proline accumulates to higher levels in TaERF87- and TaP5CS1-OE lines than that in wild-type plants under well-watered and drought stress conditions concomitantly with enhanced drought tolerance in these transgenic lines. Moreover, the interaction between TaERF87 and the bHLH transcription factor TaAKS1 synergistically enhances TaP5CS1 and TaP5CR1 transcriptional activation. TaAKS1 OE also increases wheat drought tolerance by promoting proline accumulation. Additionally, our findings verified that TaERF87 and TaAKS1 are targets of abscisic acid-responsive element binding factor 2 (TaABF2). Together, our study elucidates the mechanisms underlying a positive response to drought stress mediated by the TaABF2-TaERF87/TaAKS1-TaP5CS1/TaP5CR1 module, and identifies candidate genes for the development of elite drought-tolerant wheat varieties.

Protamine-derived peptide RPR (Arg-Pro-Arg) ameliorates oleic acid-induced lipogenesis via the PepT1 pathway in HepG2 cells.

The protamine-derived peptide arginine-proline-arginine (RPR) can ameliorate lifestyle-related diseases such as obesity and hypercholesterolemia. Thus, we hypothesized that the hypolipidemic activity of RPR could attenuate events leading to non-alcoholic fatty liver disease. Addition of 2 m m oleic acid (OA) to the culture medium induced fatty liver conditions in HepG2 cells. The OA + RPR group showed significantly decreased cellular or medium triglyceride (TG) level compared with the OA group. Stearoyl-CoA desaturase-1 (SCD1) or sterol regulatory element-binding protein 1 (SREBP1) protein level was significantly lower in the OA + RPR group than in the OA group. In the R + P + R amino acid mixture-treated group, the TG level was not significantly different from that in the OA-treated group. The OA + RP- or OA + PR-treated groups showed significantly decreased cellular TG level compared with the OA group. Moreover, the effect of RPR disappeared when the peptide transporter 1 (PepT1) was knocked down with a siRNA. Collectively, our results demonstrated that RPR effectively ameliorated hepatic steatosis in HepG2 cells via the PepT1 pathway.

Pyrroline-5-carboxylate dehydrogenase is an essential enzyme for proline dehydrogenase function during dark-induced senescence in Arabidopsis thaliana.

During leaf senescence, nitrogen is remobilized and carbon backbones are replenished by amino acid catabolism, with many of the key reactions occurring in mitochondria. The intermediate Δ1 -pyrroline-5-carboxylate (P5C) is common to some catabolic pathways, thus linking the metabolism of several amino acids, including proline and arginine. Specifically, mitochondrial proline catabolism involves sequential action of proline dehydrogenase (ProDH) and P5C dehydrogenase (P5CDH) to produce P5C and then glutamate. Arginine catabolism produces urea and ornithine, the latter in the presence of α-ketoglutarate being converted by ornithine δ-aminotransferase (OAT) into P5C and glutamate. Metabolic changes during dark-induced leaf senescence (DIS) were studied in Arabidopsis thaliana leaves of Col-0 and in prodh1prodh2, p5cdh and oat mutants. Progression of DIS was followed by measuring chlorophyll and proline contents for 5 days. Metabolomic profiling of 116 compounds revealed similar profiles of Col-0 and oat metabolism, distinct from prodh1prodh2 and p5cdh metabolism. Metabolic dynamics were accelerated in p5cdh by 1 day. Notably, more P5C and proline accumulated in p5cdh than in prodh1prodh2. ProDH1 enzymatic activity and protein amount were significantly down-regulated in p5cdh mutant at Day 4 of DIS. Mitochondrial P5C levels appeared critical in determining the flow through interconnected amino acid remobilization pathways to sustain senescence.