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The Journal of Biological Chemistry Nov 2023It is known that metabolic defects in the retinal pigment epithelium (RPE) can cause degeneration of its neighboring photoreceptors in the retina, leading to retinal...
It is known that metabolic defects in the retinal pigment epithelium (RPE) can cause degeneration of its neighboring photoreceptors in the retina, leading to retinal degenerative diseases such as age-related macular degeneration. However, how RPE metabolism supports the health of the neural retina remains unclear. The retina requires exogenous nitrogen sources for protein synthesis, neurotransmission, and energy metabolism. Using N tracing coupled with mass spectrometry, we found human RPE can utilize the nitrogen in proline to produce and export 13 amino acids, including glutamate, aspartate, glutamine, alanine, and serine. Similarly, we found this proline nitrogen utilization in the mouse RPE/choroid but not in the neural retina of explant cultures. Coculture of human RPE with the retina showed that the retina can take up the amino acids, especially glutamate, aspartate, and glutamine, generated from proline nitrogen in the RPE. Intravenous delivery of N proline in vivo demonstrated N-derived amino acids appear earlier in the RPE before the retina. We also found proline dehydrogenase, the key enzyme in proline catabolism is highly enriched in the RPE but not the retina. The deletion of proline dehydrogenase blocks proline nitrogen utilization in RPE and the import of proline nitrogen-derived amino acids in the retina. Our findings highlight the importance of RPE metabolism in supporting nitrogen sources for the retina, providing insight into understanding the mechanisms of the retinal metabolic ecosystem and RPE-initiated retinal degenerative diseases.
Topics: Animals; Humans; Mice; Amino Acids; Aspartic Acid; Glutamates; Glutamine; Nitrogen; Proline; Proline Oxidase; Retina; Retinal Pigment Epithelium
PubMed: 37741457
DOI: 10.1016/j.jbc.2023.105275 -
Journal of Chromatography. B,... Aug 2023The extracellular cellular matrix (ECM) maintains tissue structure and regulates signaling functions by continuous degradation and remodeling. Inflammation or other...
The extracellular cellular matrix (ECM) maintains tissue structure and regulates signaling functions by continuous degradation and remodeling. Inflammation or other disease conditions activate proteases including matrix metalloproteinases (MMPs) that degrade ECM proteins and in particular generate fragments of collagen and elastin, some of which are biologically active ECM peptides or matrikines. Stepwise degradation of collagen by MMP 8, 9 and prolyl endopeptidase release the matrikine proline-glycine-proline (PGP) and its product acetyl-PGP (AcPGP). These peptides are considered as potential biomarkers and therapeutic targets for many disease conditions such as chronic lung disease, heart disease, and cancer. However, there is no published, validated method for the measurement of PGP and AcPGP in plasma and therefore, we developed a sensitive, selective and reliable, isotope dilution LC-multiple reaction monitoring MS method for their determination in human plasma. The chromatographic separation of PGP and AcPGP was achieved in 3 min using Jupiter column with a gradient consisting of acidified acetonitrile and water at a flow rate of 0.5 ml/min. The limit of detection (LOD) for PGP and AcPGP was 0.01 ng/ml and the limit of quantification (LOQ) was 0.05 ng/ml and 0.1 ng/ml, respectively. Precision and accuracy values for all analytes were within 20 % except for the lowest QC of 0.01 ng/ml. The mean extraction recoveries of these analytes were > 90 % using a Phenomenex Phree cartridge and the matrix effect was < 15 % for all the QCs for PGP and AcPGP except the lowest QC. The stability of PGP and AcPGP was > 90 % in several tested conditions including autosampler use, storage at -80 °C, and after 6 times freeze-thaw cycles. Using this method, we successfully extracted and determined PGP levels in human plasma from healthy and COPD subjects. Therefore, this method is suitable for quantification of these peptides in the clinical setting.
Topics: Humans; Chromatography, Liquid; Proline; Glycine; Tandem Mass Spectrometry; Peptides; Collagen
PubMed: 37453387
DOI: 10.1016/j.jchromb.2023.123815 -
Frontiers of Medicine Dec 2023The coronavirus disease 2019 (COVID-19) pandemic has stimulated tremendous efforts to develop therapeutic agents that target severe acute respiratory syndrome... (Review)
Review
The coronavirus disease 2019 (COVID-19) pandemic has stimulated tremendous efforts to develop therapeutic agents that target severe acute respiratory syndrome coronavirus 2 to control viral infection. So far, a few small-molecule antiviral drugs, including nirmatrelvir-ritonavir (Paxlovid), remdesivir, and molnupiravir have been marketed for the treatment of COVID-19. Nirmatrelvir-ritonavir has been recommended by the World Health Organization as an early treatment for outpatients with mild-to-moderate COVID-19. However, the existing treatment options have limitations, and effective treatment strategies that are cost-effective and convenient for tackling COVID-19 are still needed. To date, four domestically developed oral anti-COVID-19 drugs have been granted conditional market approval in China. These drugs include azvudine, simnotrelvir-ritonavir (Xiannuoxin), leritrelvir, and mindeudesivir (VV116). Preclinical and clinical studies have explored the efficacy and tolerability of mindeudesivir and supported its early use in mild-to-moderate COVID-19 cases at high risk for progression. In this review, we discuss the most recent findings regarding the pharmacological mechanism and therapeutic effects focusing on mindeudesivir and other small-molecule antiviral agents for COVID-19. These findings will expand our understanding and highlight the potential widespread application of China's homegrown anti-COVID-19 drugs.
Topics: Humans; Ritonavir; COVID-19; Antiviral Agents; China; Nitriles; Lactams; Proline; Adenosine; Leucine
PubMed: 38165534
DOI: 10.1007/s11684-023-1037-3 -
PLoS Pathogens Nov 2023Candida albicans, the primary etiology of human mycoses, is well-adapted to catabolize proline to obtain energy to initiate morphological switching (yeast to hyphal) and...
Candida albicans, the primary etiology of human mycoses, is well-adapted to catabolize proline to obtain energy to initiate morphological switching (yeast to hyphal) and for growth. We report that put1-/- and put2-/- strains, carrying defective Proline UTilization genes, display remarkable proline sensitivity with put2-/- mutants being hypersensitive due to the accumulation of the toxic intermediate pyrroline-5-carboxylate (P5C), which inhibits mitochondrial respiration. The put1-/- and put2-/- mutations attenuate virulence in Drosophila and murine candidemia models and decrease survival in human neutrophils and whole blood. Using intravital 2-photon microscopy and label-free non-linear imaging, we visualized the initial stages of C. albicans cells infecting a kidney in real-time, directly deep in the tissue of a living mouse, and observed morphological switching of wildtype but not of put2-/- cells. Multiple members of the Candida species complex, including C. auris, are capable of using proline as a sole energy source. Our results indicate that a tailored proline metabolic network tuned to the mammalian host environment is a key feature of opportunistic fungal pathogens.
Topics: Animals; Mice; Humans; Candida albicans; Virulence; Saccharomyces cerevisiae; Proline; Candida; Mammals
PubMed: 37917600
DOI: 10.1371/journal.ppat.1011677 -
Science Advances May 2024Metabolic reprogramming is critical in the onset of pressure overload-induced cardiac remodeling. Our study reveals that proline dehydrogenase (PRODH), the key enzyme in...
Metabolic reprogramming is critical in the onset of pressure overload-induced cardiac remodeling. Our study reveals that proline dehydrogenase (PRODH), the key enzyme in proline metabolism, reprograms cardiomyocyte metabolism to protect against cardiac remodeling. We induced cardiac remodeling using transverse aortic constriction (TAC) in both cardiac-specific PRODH knockout and overexpression mice. Our results indicate that PRODH expression is suppressed after TAC. Cardiac-specific PRODH knockout mice exhibited worsened cardiac dysfunction, while mice with PRODH overexpression demonstrated a protective effect. In addition, we simulated cardiomyocyte hypertrophy in vitro using neonatal rat ventricular myocytes treated with phenylephrine. Through RNA sequencing, metabolomics, and metabolic flux analysis, we elucidated that PRODH overexpression in cardiomyocytes redirects proline catabolism to replenish tricarboxylic acid cycle intermediates, enhance energy production, and restore glutathione redox balance. Our findings suggest PRODH as a modulator of cardiac bioenergetics and redox homeostasis during cardiac remodeling induced by pressure overload. This highlights the potential of PRODH as a therapeutic target for cardiac remodeling.
Topics: Animals; Ventricular Remodeling; Proline; Myocytes, Cardiac; Mice; Mice, Knockout; Rats; Proline Oxidase; Energy Metabolism; Myocardium; Cardiomegaly; Disease Models, Animal; Oxidation-Reduction; Male; Metabolic Reprogramming
PubMed: 38718121
DOI: 10.1126/sciadv.adl3549 -
Cells Jul 2023Despite significant progress in clinical management, colorectal cancer (CRC) remains the third most common cause of cancer-related deaths. A positive association between...
BACKGROUND
Despite significant progress in clinical management, colorectal cancer (CRC) remains the third most common cause of cancer-related deaths. A positive association between PYCR2 (pyrroline-5-carboxylate reductase-2), a terminal enzyme of proline metabolism, and CRC aggressiveness was recently reported. However, how PYCR2 promotes colon carcinogenesis remains ill understood.
METHODS
A comprehensive analysis was performed using publicly available cancer databases and CRC patient cohorts. Proteomics and biochemical evaluations were performed along with genetic manipulations and in vivo tumor growth assays to gain a mechanistic understanding.
RESULTS
PYCR2 expression was significantly upregulated in CRC and associated with poor patient survival, specifically among PYCR isoforms (PYCR1, 2, and 3). The genetic inhibition of PYCR2 inhibited the tumorigenic abilities of CRC cells and in vivo tumor growth. Coinciding with these observations was a significant decrease in cellular proline content. PYCR2 overexpression promoted the tumorigenic abilities of CRC cells. Proteomics (LC-MS/MS) analysis further demonstrated that PYCR2 loss of expression in CRC cells inhibits survival and cell cycle pathways. A subsequent biochemical analysis supported the causal role of PYCR2 in regulating CRC cell survival and the cell cycle, potentially by regulating the expression of MASTL, a cell-cycle-regulating protein upregulated in CRC. Further studies revealed that PYCR2 regulates Wnt/β-catenin-signaling in manners dependent on the expression of MASTL and the cancer stem cell niche.
CONCLUSIONS
PYCR2 promotes MASTL/Wnt/β-catenin signaling that, in turn, promotes cancer stem cell populations and, thus, colon carcinogenesis. Taken together, our data highlight the significance of PYCR2 as a novel therapeutic target for effectively treating aggressive colon cancer.
Topics: Humans; Protein Serine-Threonine Kinases; beta Catenin; Chromatography, Liquid; Tandem Mass Spectrometry; Carcinogenesis; Wnt Signaling Pathway; Colonic Neoplasms; Microtubules; Proline; Oxidoreductases; Serine; Microtubule-Associated Proteins
PubMed: 37508547
DOI: 10.3390/cells12141883 -
Cells Nov 2023The culture of embryos in the non-essential amino acid L-proline (Pro) or its analogues pipecolic acid (PA) and L-4-thiazolidine carboxylic acid (L4T) improves embryo...
The culture of embryos in the non-essential amino acid L-proline (Pro) or its analogues pipecolic acid (PA) and L-4-thiazolidine carboxylic acid (L4T) improves embryo development, increasing the percentage that develop to the blastocyst stage and hatch. Staining of 2-cell and 4-cell embryos with tetramethylrhodamine methyl ester and 2',7'-dichlorofluorescein diacetate showed that the culture of embryos in the presence of Pro, or either of these analogues, reduced mitochondrial activity and reactive oxygen species (ROS), respectively, indicating potential mechanisms by which embryo development is improved. Inhibition of the Pro metabolism enzyme, proline oxidase, by tetrahydro-2-furoic-acid prevented these reductions and concomitantly prevented the improved development. The ways in which Pro, PA and L4T reduce mitochondrial activity and ROS appear to differ, despite their structural similarity. Specifically, the results are consistent with Pro reducing ROS by reducing mitochondrial activity while PA and L4T may be acting as ROS scavengers. All three may work to reduce ROS by contributing to the GSH pool. Overall, our results indicate that reduction in mitochondrial activity and oxidative stress are potential mechanisms by which Pro and its analogues act to improve pre-implantation embryo development.
Topics: Animals; Mice; Reactive Oxygen Species; Proline; Oxidative Stress; Blastocyst; Embryonic Development
PubMed: 37998375
DOI: 10.3390/cells12222640 -
Andrology Oct 2023Asthenozoospermia is one of the essential causes of male infertility, and its incidence is significantly higher in obese men. Due to its complex etiology and unknown...
BACKGROUND
Asthenozoospermia is one of the essential causes of male infertility, and its incidence is significantly higher in obese men. Due to its complex etiology and unknown pathomechanism, the diagnosis and treatment of obesity-induced asthenozoospermia is a prevalent problem in reproductive medicine.
OBJECTIVE
This study aims to explore major differential metabolites and metabolic pathways in seminal plasma and pathological mechanisms for obesity-induced asthenozoospermia.
MATERIALS AND METHODS
We performed non-target metabolomic studies on the seminal plasma of healthy men with normal semen parameters (HN group, n = 20), obese men with normal semen parameters (ON group, n = 20), and men with obesity-induced asthenozoospermia (OA group, n = 20) based on gas chromatography-mass spectrometry. Metabolic profilings and related pathway analyses were conducted to discriminate differential metabolites and metabolic pathways.
RESULTS
A total of 20 differential metabolites including fructose, succinic acid, aconitic acid, methylmaleic acid, glucopyranose, serine, valine, leucine, phenylalanine, glycine, glutamic acid, alanine, proline and threonine were identified in HN group and ON group; 24 differential metabolites including glucose, fructose, pyruvic acid, citric acid, succinic acid, aconitic acid, glucopyranose, glutamic acid, valine, leucine, glycine, phenylalanine, lysine, citrulline, proline and alanine were produced in OA group and ON group; and 28 differential metabolites including glucose, fructose, citric acid, succinic acid, glucopyranose, valine, glycine, serine, leucine, phenylalanine, alanine, threonine, proline, glutamic acid, citrulline, lysine and tyrosine were produced in OA group and HN group. In addition, abnormal energy metabolism including carbohydrate metabolism (TCA cycle, glycolysis/gluconeogenesis, and pyruvate metabolism) and amino acid metabolism (phenylalanine, tyrosine, and tryptophan biosynthesis, D-glutamine and D-glutamate metabolism; phenylalanine metabolism, etc.) were found in ON group and OA group.
CONCLUSION
Obesity could affect the metabolite composition in seminal plasma and abnormal energy metabolism in seminal plasma mainly including carbohydrate metabolism and amino acid metabolism were closely related to obesity-induced asthenozoospermia.
Topics: Male; Humans; Semen; Asthenozoospermia; Leucine; Lysine; Glutamic Acid; Citrulline; Aconitic Acid; Succinic Acid; Metabolomics; Alanine; Proline; Glycine; Tyrosine; Phenylalanine; Valine; Serine; Threonine; Glucose
PubMed: 36841993
DOI: 10.1111/andr.13412 -
Journal of Bioscience and Bioengineering Dec 2023Proline, which is a predominant amino acid in grape musts, is involved in the taste and flavor of foods and beverages. The yeast Saccharomyces cerevisiae poorly utilizes...
Proline, which is a predominant amino acid in grape musts, is involved in the taste and flavor of foods and beverages. The yeast Saccharomyces cerevisiae poorly utilizes proline in wine-making processes, leading to a nitrogen deficiency during fermentation and proline accumulation in wine. Previous studies have shown that the protein kinase A (PKA) pathway is involved in inhibitory mechanisms of proline utilization. In this study, we screened the PKA pathway-related genes that regulate proline utilization. Using a yeast culture collection of disrupted strains associated with the downstream of the PKA cascade, we revealed that the stress-responsive transcription factor genes MSN2/4 regulate proline utilization. Moreover, we found that Msn2/4 up-regulate the SHY1 gene during the cell growth of the wine fermentation model, which may cause the inhibition of proline utilization. The SHY1-deleted strain of the commercial wine yeast clearly showed proline consumption and average ethanol production under the wine fermentation model. The present data indicate that the PKA-Msn2/4-Shy1 cascade controls the inhibition of proline utilization under wine-making processes. Our study could hold promise for the development of wine yeast strains that can efficiently reduce proline during wine fermentation.
Topics: Fermentation; Saccharomyces cerevisiae; Wine; Proline; Amino Acids; DNA-Binding Proteins; Transcription Factors; Saccharomyces cerevisiae Proteins
PubMed: 37940488
DOI: 10.1016/j.jbiosc.2023.10.005 -
International Journal of Molecular... Sep 2023represents a type of single-transmembrane adaptor protein containing an N-terminal cysteine-rich domain and a proline-rich C-terminal region. Nine subfamily genes have...
represents a type of single-transmembrane adaptor protein containing an N-terminal cysteine-rich domain and a proline-rich C-terminal region. Nine subfamily genes have been proposed in most vertebrates; however, some might be species-specific. The number of genes present in zebrafish remains unclear. This study aimed to investigate the evolutionary relationships among family genes in zebrafish (TU strain) using phylogenetic and syntenic analyses. The function of was preliminarily examined via CRISPR/Cas13d-mediated knockdown. Following identification in zebrafish, 10 family genes, namely , , , , , , , , , and , were classified into three main clades and six subclades. Their encoding proteins contained a cysteine-rich N-terminal domain and a proline-rich C-terminal region containing different motifs. A specific syntenic block containing and was observed to be conserved across all species. Furthermore, all these genes were expressed during embryogenesis. was expressed in the presomitic mesoderm, somites, and so on. was identified as a regulator of the expression of the somite formation marker . Overall, our study provides new insights into the evolution of family genes and the control of over the convergent extension cells of somitic precursors in zebrafish.
Topics: Animals; Zebrafish; Zebrafish Proteins; Phylogeny; Cysteine; Membrane Proteins; Proline; Gene Expression Regulation, Developmental
PubMed: 37762365
DOI: 10.3390/ijms241814062