With persistent elevation in global temperature, water scarcity becomes a major threat to plant growth and development, yield security, agricultural sustainability, and food production. Proline, as a key osmolyte and antioxidant, plays a critical role in regulating drought tolerance in plants, especially its key biosynthetic enzyme, delta-1-pyrroline-5-carboxylate synthase (P5CS), which always positively responds to drought stress. As an important woody oil crop, the expansion of Paeonia ostii cultivation needs to address the issue of plant drought tolerance. Here, we isolated a PoP5CS gene from P. ostii, with an open reading frame of 1842 bp encoding 613 amino acids. PoP5CS expression progressively increased in response to increasing drought stress, and it was localized in the cytoplasm. Silencing of PoP5CS in P. ostii reduced drought tolerance, accompanied by decreased proline content, elevated reactive oxygen species (ROS) accumulation, and increased relative electrical conductivity (REC) and malondialdehyde (MDA) levels. Conversely, overexpression of PoP5CS in Nicotiana tabacum plants enhanced drought resistance, manifested by increased proline levels, reduced ROS accumulation, and lower REC and MDA contents. This study isolates PoP5CS from P. ostii and validates its role in regulating drought tolerance, providing valuable genetic resources and theoretical insights for the development of drought-resistant P. ostii cultivars.

The aim of the study was to investigate changes in proline metabolism in seedlings of tree species during drought stress. One month old Paulownia tomentosa seedlings were exposed to moisture conditions at various levels (irrigation at 100, 75, 50 and 25% of field capacity), and then the material (leaves and roots) was collected three times at 10-day intervals. The activity of enzymes involved in proline metabolism was closely related to drought severity; however, proline content was not directly impacted. The activity of pyrroline-5-carboxylate synthetase (P5CS), which catalyzes proline biosynthesis, increased in response to hydrogen peroxide accumulation, which was correlated with soil moisture. In contrast, the activity of proline dehydrogenase (ProDH), which catalyzes proline catabolism, decreased. Compared to proline, the activity of these enzymes may be a more reliable biochemical marker of stress-induced oxidative changes. The content of proline is dependent on numerous additional factors, i.e., its degradation is an important alternative energy source. Moreover, we noted tissue-specific differences in this species, in which roots appeared to be proline biosynthesis sites and leaves appeared to be proline catabolism sites. Further research is needed to examine a broader view of proline metabolism as a cycle regulated by multiple mechanisms and differences between species.

Introduction: Carnosol exhibited ameliorating effects on muscle atrophy of mice developed cancer cachexia in our previous research. Method: Here, the ameliorating effects of carnosol on the C2C12 myotube atrophy result from simulated cancer cachexia injury, the conditioned medium of the C26 tumor cells or the LLC tumor cells, were observed. To clarify the mechanisms of carnosol, the possible direct target proteins of carnosol were searched using DARTS (drug affinity responsive target stability) assay and then confirmed using CETSA (cellular thermal shift assay). Furthermore, proteomic analysis was used to search its possible indirect target proteins by comparing the protein expression profiles of C2C12 myotubes under treatment of C26 medium, with or without the presence of carnosol. The signal network between the direct and indirect target proteins of carnosol was then constructed. Results: Our results showed that, Delta-1-pyrroline-5-carboxylate synthase (P5CS) might be the direct target protein of carnosol in myotubes. The influence of carnosol on amino acid metabolism downstream of P5CS was confirmed. Carnosol could upregulate the expression of proteins related to glutathione metabolism, anti-oxidant system, and heat shock response. Knockdown of P5CS could also ameliorate myotube atrophy and further enhance the ameliorating effects of carnosol. Discussion: These results suggested that carnosol might ameliorate cancer cachexia-associated myotube atrophy by targeting P5CS and its downstream pathways.

Physiological, metabolic, and genetic changes produced by two plant growth promoting rhizobacteria (PGPR) Pseudomonas sp. (internal code of the laboratory: N 5.12 and N 21.24) inoculated in tomato plants subjected to moderate water stress (10% polyethylene glycol-6000; PEG) were studied. Photosynthesis efficiency, photosynthetic pigments, compatible osmolytes, reactive oxygen species (ROS) scavenging enzymes activities, oxidative stress level and expression of genes related to abscisic acid synthesis (ABA; 9-cis-epoxycarotenoid dioxygenase NCDE1 gene), proline synthesis (Pyrroline-5-carboxylate synthase P5CS gene), and plasma membrane ATPase (PM ATPase gene) were measured. Photosynthetic efficiency was compromised by PEG, but bacterial-inoculated plants reversed the effects: while N5.12 increased carbon fixation (37.5%) maintaining transpiration, N21.24 increased both (14.2% and 31%), negatively affecting stomatal closure, despite the enhanced expression of NCDE1 and plasma membrane ATPase genes, evidencing the activation of different adaptive mechanisms. Among all parameters evaluated, photosynthetic pigments and antioxidant enzymes guaiacol peroxidase (GPX) and ascorbate peroxidase (APX) responded differently to both strains. N 5.12 increased photosynthetic pigments (70% chlorophyll a, 69% chlorophyll b, and 65% carotenoids), proline (33%), glycine betaine (4.3%), and phenolic compounds (21.5%) to a greater extent, thereby decreasing oxidative stress (12.5% in Malondialdehyde, MDA). Both bacteria have highly beneficial effects on tomato plants subjected to moderate water stress, improving their physiological state. The use of these bacteria in agricultural production systems could reduce the amount of water for agricultural irrigation without having a negative impact on food production.

Salinity is one of the most concerning ecological restrictions influencing plant growth, which poses a devastating threat to global agriculture. Surplus quantities of ROS generated under stress conditions have negative effects on plants\' growth and survival by damaging cellular components, including nucleic acids, lipids, proteins and carbohydrates. However, low levels of ROS are also necessary because of their role as signalling molecules in various development-related pathways. Plants possess sophisticated antioxidant systems for scavenging as well as regulating ROS levels to protect cells from damage. Proline is one such crucial non-enzymatic osmolyte of antioxidant machinery that functions in the reduction of stress. There has been extensive research on improving the tolerance, effectiveness, and protection of plants against stress, and to date, various substances have been used to mitigate the adverse effects of salt. In the present study Zinc (Zn) was applied to elucidate its effect on proline metabolism and stress-responsive mechanisms in proso millet. The results of our study indicate the negative impact on growth and development with increasing treatments of NaCl. However, the low doses of exogenous Zn proved beneficial in mitigating the effects of NaCl by improving morphological and biochemical features. In salt-treated plants, the low doses of Zn (1 mg/L, 2 mg/L) rescued the negative impact of salt (150mM) as evidenced by increase in shoot length (SL) by 7.26% and 25.5%, root length (RL) by 21.84% and 39.07% and membrane stability index (MSI) by 132.57% and 151.58% respectively.The proline content improved at all concentrations with maximum increase of 66.65% at 2 mg/L Zn. Similarly, the low doses of Zn also rescued the salt induced stress at 200mM NaCl. The enzymes related to proline biosynthesis were also improved at lower doses of Zn. In salt treated plants (150mM), Zn (1 mg/L, 2 mg/L) increased the activity of P5CS by 19.344% and 21%. The P5CR and OAT activities were also improved with maximum increase of 21.66% and 21.84% at 2 mg/L Zn respectively. Similarly, the low doses of Zn also increased the activities of P5CS, P5CR and OAT at 200mM NaCl. Whereas P5CDH enzyme activity showed a decrease of 82.5% at 2mg/L Zn+150mM NaCl and 56.7% at 2mg/L Zn+200 mM NaCl. These results strongly imply the modulatory role of Zn in maintaining of proline pool during NaCl stress.

Δ1-Pyrroline-5-carboxylate synthetase (P5CS) acts as the rate-limiting enzyme in the biosynthesis of proline in plants. Although P5CS plays an essential role in plant responses to environmental stresses, its biological functions remain largely unclear in pear (Pyrus betulifolia). In the present study, 11 putative pear P5CSs (PbP5CSs) were identified by comprehensive bioinformatics analysis and classified into five subfamilies. Segmental and tandem duplications contributed to the expansion and evolution of the PbP5CS gene family. Various cis-acting elements associated with plant development, hormone responses, and/or stress responses were identified in the promoters of PbP5CS genes. To investigate the regulatory roles of PbP5CS genes in response to abiotic and biotic stresses, gene expression patterns in publicly available data were explored. The tissue-specific expressional dynamics of PbP5CS genes indicate potentially important roles in pear growth and development. Their spatiotemporal expression patterns suggest key functions in multiple environmental stress responses. Transcriptome and real-time quantitative PCR analyses revealed that most PbP5CS genes exhibited distinct expression patterns in response to drought, waterlogging, salinity-alkalinity, heat, cold, and infection by Alternaria alternate and Gymnosporangium haraeanum. The results provide insight into the versatile functions of the PbP5CS gene family in stress responses. The findings may assist further exploration of the physiological functions of PbP5CS genes for the development and enhancement of stress tolerance in pear and other fruits.

Autosomal recessive cutis laxa type IIIA is a very rare genetic condition, caused by pathogenic variants in ALDH18A1, encoding delta-1-pyrroline-5-carboxylate synthase (P5CS). This enzyme catalyzes the reduction of glutamic acid to delta1-pyrroline-5-carboxylate, playing a key role in the de novo biosynthesis of proline, ornithine, and arginine. Autosomal recessive cutis laxa type IIIA is characterized by abundant and wrinkled skin, skeletal anomalies, cataract or corneal clouding and neuro-developmental disorders of variable degree. We report on a patient with autosomal recessive cutis laxa type IIIA, due to a homozygous missense c.1273C > T; p. (Arg425Cys) pathogenic variant in ALDH18A1. The patient presented a severe phenotype with serious urological involvement, peculiar cerebro-vascular abnormalities and neurodevelopmental compromise. This description contributes to better characterize the phenotypic spectrum associated with ALDH18A1 pathogenic variants, confirming the systemic involvement as a typical feature of autosomal recessive cutis laxa type IIIA.

The bifunctional enzyme Δ1-pyrroline-5-carboxylate synthase (P5CS) is vital to the synthesis of proline and ornithine, playing an essential role in human health and agriculture. Pathogenic mutations in the P5CS gene (ALDH18A1) lead to neurocutaneous syndrome and skin relaxation connective tissue disease in humans, and P5CS deficiency seriously damages the ability to resist adversity in plants. We have recently found that P5CS forms cytoophidia in vivo and filaments in vitro. However, it is difficult to appreciate the function of P5CS filamentation without precise structures. Using cryo-electron microscopy, here we solve the structures of Drosophila full-length P5CS in three states at resolution from 3.1 to 4.3 Å. We observe distinct ligand-binding states and conformational changes for the GK and GPR domains, respectively. Divergent helical filaments are assembled by P5CS tetramers and stabilized by multiple interfaces. Point mutations disturbing those interfaces prevent P5CS filamentation and greatly reduce the enzymatic activity. Our findings reveal that filamentation is crucial for the coordination between the GK and GPR domains, providing a structural basis for the catalytic function of P5CS filaments.

Cadmium (Cd) accumulation in agricultural soils constitutes a serious problem for crop yields and food safety. It is known that proline (Pro) can rapidly accumulate in plant tissues in response to abiotic stress. To analyze the potential protective effect of Pro accumulation against Cd toxicity, we compared the response to Cd stress of wild-type (WT) Medicago truncatula and a transgenic line that we had previously obtained and characterized (p18), which expressed the Δ 1-pyrroline-5-carboxylate synthetase gene from Vigna aconitifolia (VaP5CS), and accumulated high Pro levels. Cadmium significantly reduced germination of WT seeds compared to p18 seeds, and seedling relative root growth, a valid indicator of metal tolerance, was significantly higher for p18 than WT seedlings. We analyzed the relative expression of genes related to Pro metabolism, phytochelatin biosynthesis. antioxidant machinery, and NADPH recycling, which are relevant mechanisms in the response to Cd stress. They presented differential expression in the seedlings of both genotypes both under control conditions and under Cd stress, suggesting that the Cd response mechanisms might be constitutively activated in the transgenic line. Pro accumulation promoted higher survival, enhanced growth performance, and minor nutrient imbalance in transgenic p18 plants compared to WT plants. These facts, together with the recorded gluthatione levels, lipid peroxidation and antioxidant enzyme activities strongly suggested that VaP5CS expression and Pro accumulation conferred enhanced Cd tolerance to M. truncatula p18 plants, which was likely mediated by changes in Pro metabolism, increased phytochelatin biosynthesis and a more efficient antioxidant response. Moreover, p18 roots accumulated significantly higher Cd amounts than WT roots, while Cd translocation to the aerial part was similar to WT plants, thus suggesting that high Pro levels increased not only Cd tolerance, but also Cd phytostabilization by rhizosequestration.

Free proline has multiple functions in plant cells, such as regulating osmotic potential and protecting both proteins and cell membranes. The expression of Δ1-Pyrroline-5-carboxylate synthase (P5CS), a key enzyme in the proline biosynthetic pathway, increases under drought, salt and cold stress conditions, causing plant cells to accumulate large amounts of proline. In this study, we cloned and identified the P5CS gene from Stipa purpurea, which has a full-length of 2196 bp and encodes 731 amino acids. A subcellular localization analysis indicated that SpP5CS localized to the cytoplasm. The ectopic overexpression of SpP5CS in Arabidopsis thaliana resulted in higher proline contents, longer roots, higher survival rates and less membrane damage under drought stress conditions compared with wild-type controls. SpP5CS-overexpressing A. thaliana was more resistant to drought stress than the wild type, whereas the deletion mutant sp5cs was less resistant to drought stress. Thus, SpP5CS may be a potential candidate target gene for increasing plant resistance to drought stress.