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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 -
The EMBO Journal Oct 2023Mitochondria play essential roles in cancer cell adaptation to hypoxia, but the underlying mechanisms remain elusive. Through mitochondrial proteomic profiling, we here...
Mitochondria play essential roles in cancer cell adaptation to hypoxia, but the underlying mechanisms remain elusive. Through mitochondrial proteomic profiling, we here find that the prolyl hydroxylase EglN1 (PHD2) accumulates on mitochondria under hypoxia. EglN1 substrate-binding region in the β2β3 loop is responsible for its mitochondrial translocation and contributes to breast tumor growth. Furthermore, we identify AMP-activated protein kinase alpha (AMPKα) as an EglN1 substrate on mitochondria. The EglN1-AMPKα interaction is essential for their mutual mitochondrial translocation. After EglN1 prolyl-hydroxylates AMPKα under normoxia, they rapidly dissociate following prolyl-hydroxylation, leading to their immediate release from mitochondria. In contrast, hypoxia results in constant EglN1-AMPKα interaction and their accumulation on mitochondria, leading to the formation of a Ca /calmodulin-dependent protein kinase 2 (CaMKK2)-EglN1-AMPKα complex to activate AMPKα phosphorylation, ensuring metabolic homeostasis and breast tumor growth. Our findings identify EglN1 as an oxygen-sensitive metabolic checkpoint signaling hypoxic stress to mitochondria through its β2β3 loop region, suggesting a potential therapeutic target for breast cancer.
Topics: Female; Humans; AMP-Activated Protein Kinases; Breast Neoplasms; Hypoxia; Hypoxia-Inducible Factor-Proline Dioxygenases; Mitochondria; Proteomics
PubMed: 37661833
DOI: 10.15252/embj.2023113743 -
Pathogens (Basel, Switzerland) Sep 2023In , proline dehydrogenase (PruB) and ∆-pyrroline-5-carboxylate (P5C) dehydrogenase (PruA) are monofunctional enzymes that catalyze proline oxidation to glutamate via...
In , proline dehydrogenase (PruB) and ∆-pyrroline-5-carboxylate (P5C) dehydrogenase (PruA) are monofunctional enzymes that catalyze proline oxidation to glutamate via the intermediates P5C and L-glutamate-γ-semialdehyde. Both enzymes are essential for the replication of pathogenic . Highly active enzymes were expressed and purified using a expression system. The purified enzymes were characterized using natural substrates and chemically synthesized analogs. The structural requirements of the quinone electron acceptor were examined. PruB displayed activity with all tested lipoquinone analogs (naphthoquinone or benzoquinone). In PruB assays utilizing analogs of the native naphthoquinone [MK-9 (II-H)] specificity constants were an order of magnitude greater for the menaquinone analogs than the benzoquinone analogs. In addition, mycobacterial PruA was enzymatically characterized for the first time using exogenous chemically synthesized P5C. A value of 120 ± 0.015 µM was determined for P5C, while the value for NAD was shown to be 33 ± 4.3 µM. Furthermore, proline competitively inhibited PruA activity and coupled enzyme assays, suggesting that the recombinant purified monofunctional PruB and PruA enzymes of channel substrate likely increase metabolic flux and protect the bacterium from methylglyoxal toxicity.
PubMed: 37764979
DOI: 10.3390/pathogens12091171 -
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 -
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 -
Frontiers in Oncology 2023Emerging evidence suggests that proline metabolism is important for regulating the survival and death of different types of cancer cells. Proline dehydrogenase (PRODH),... (Review)
Review
Emerging evidence suggests that proline metabolism is important for regulating the survival and death of different types of cancer cells. Proline dehydrogenase (PRODH), an enzyme catalyzing proline catabolism, and the degradation products of proline by PRODH, such as ATP and ROS, are known to play critical roles in cancer progression. Notably, the role of PRODH in cancer is still complicated and unclear, and primarily depends on the cancer type and tumor microenvironment. For instance, PRODH induces apoptosis and senescence through ROS signaling in different types of cancers, while as a protumor factor, PRODH promotes malignant phenotypes of certain tumors under stresses such as hypoxia. In order to assess whether PRODH can serve as a novel target for cancer therapy, we will provide an overview of the biological functions of PRODH and its double-edged role in cancer in this article.
PubMed: 38023181
DOI: 10.3389/fonc.2023.1254439 -
Human Reproduction (Oxford, England) Nov 2023Is the abundance of certain biochemical compounds in human cumulus cells (CCs) related to oocyte quality?
STUDY QUESTION
Is the abundance of certain biochemical compounds in human cumulus cells (CCs) related to oocyte quality?
SUMMARY ANSWER
Malonate, 5-oxyproline, and erythronate were positively associated with pregnancy potential.
WHAT IS KNOWN ALREADY
CCs are removed and discarded prior to ICSI, thereby constituting an interesting biological material on which to perform molecular analysis aimed to predict oocyte developmental competence. Mitochondrial DNA content and transcriptional analyses in CC have been shown to provide a poor predictive value of oocyte competence, but the untargeted analysis of biochemical compounds (metabolomics) has been unexplored.
STUDY DESIGN, SIZE, DURATION
CCs were obtained from three groups of cumulus-oocyte complexes (COCs) of known developmental potential: oocytes not developing to blastocyst following ICSI (Bl-); oocytes developing to blastocyst but failing to establish pregnancy following embryo transfer (P-); and oocytes developing to blastocyst able to establish a pregnancy (P+). Metabolomics analyses were performed on 12 samples per group, each sample comprising the CC recovered from a single COC.
PARTICIPANTS/MATERIALS, SETTING, METHODS
Human CC samples were obtained from IVF treatments. Only unfrozen oocytes and embryos not submitted to preimplantation genetic testing were included in the analysis. Metabolomics analysis was performed by ultra-high performance liquid chromatography-tandem mass spectroscopy.
MAIN RESULTS AND THE ROLE OF CHANCE
The analysis identified 98 compounds, five of which were differentially abundant (P < 0.05) between groups: asparagine, proline, and malonate were less abundant in P- compared to Bl-, malonate and 5-oxoproline were less abundant in P- group compared to P+, and erythronate was less abundant in Bl- group compared to P+. No significant association between the abundance of the compounds identified and donor age or BMI was noted.
LIMITATIONS, REASONS FOR CAUTION
Data dispersion and the lack of coherence between developmental groups preclude the direct use of metabolic markers in clinical practice, where the uterine environment plays a major role in pregnancy outcome. The abundance of other compounds not detected by the analysis may be associated with oocyte competence. As donors were lean (only two with BMI > 30 kg/m2) and young (<34 years old), a possible effect of obesity or advanced age on the CC metabolome could not be determined.
WIDER IMPLICATIONS OF THE FINDINGS
The abundance of malonate, 5-oxyproline, and erythronate in CC was significantly higher in COCs ultimately establishing pregnancy, providing clues on the pathways required for oocyte competence. The untargeted analysis uncovered the presence of compounds that were not expected in CC, such as β-citrylglutamate and the neurotransmitter N-acetyl-aspartyl-glutamate, which may play roles in chromatin remodeling and signaling, respectively.
STUDY FUNDING/COMPETING INTEREST(S)
Research was supported by the Industrial Doctorate Project IND2017/BIO-7748 funded by Madrid Region Government. The authors declare no competing interest.
TRIAL REGISTRATION NUMBER
N/A.
Topics: Female; Humans; Pregnancy; Adult; Cumulus Cells; Hydroxyproline; Oocytes; Oogenesis; Malonates
PubMed: 37697661
DOI: 10.1093/humrep/dead181 -
BMC Geriatrics Oct 2023Sleep duration and amino acid intake are independently associated with cognitive decline. This study aimed to determine the longitudinal association between sleep...
Dietary amino acid intake and sleep duration are additively involved in future cognitive decline in Japanese adults aged 60 years or over: a community-based longitudinal study.
BACKGROUND
Sleep duration and amino acid intake are independently associated with cognitive decline. This study aimed to determine the longitudinal association between sleep duration and cognitive impairment incidence and to examine the involvement of diet, particularly amino acid intake, in these associations in community dwellers.
METHODS
In this longitudinal study in a community-based setting, we analyzed data from 623 adults aged 60-83 years without cognitive impairment at baseline. Sleep duration was assessed using a self-report questionnaire. Amino acid intake was assessed using 3-day dietary records. Cognitive impairment was defined as a Mini-Mental State Examination score ≤ 27. Participants were classified into short-, moderate-, and long-sleep groups according to baseline sleep duration (≤ 6, 7-8, and > 8 h, respectively). Using moderate sleep as a reference, odds ratios (ORs) and 95% confidence intervals (CIs) of short- and long-sleep for cognitive-impairment incidence were estimated using the generalized estimating equation. Participants were classified according to sex-stratified quartiles (Q) of 19 amino acid intake: Q1 and Q2-Q4 were low- and middle to high-intake groups, respectively. Using middle- to high-intake as a reference, ORs and 95% CIs of low intake for cognitive impairment incidence were estimated using the generalized estimating equation in each sleep-duration group. Follow-up period, sex, age, body mass index, depressive symptoms, education, smoking status, employment status, sleep aids use, physical activity, medical history, and Mini-Mental State Examination score at baseline were covariates.
RESULTS
Mean follow-up period was 6.9 ± 2.1 years. Adjusted ORs (95% CIs) for cognitive impairment in short- and long-sleep groups were 0.81 (0.49-1.35, P = 0.423) and 1.41 (1.05-1.87, P = 0.020), respectively. Particularly in long sleepers (i.e., > 8 h), cognitive impairment was significantly associated with low cystine, proline, and serine intake [adjusted ORs (95% CIs) for cognitive impairment were 2.17 (1.15-4.11, P = 0.017), 1.86 (1.07-3.23, P = 0.027), and 2.21 (1.14-4.29, P = 0.019), respectively].
CONCLUSIONS
Community-dwelling adults aged ≥ 60 years who sleep longer are more likely to have cognitive decline, and attention should be paid to the low cystine, proline, and serine intake.
Topics: Humans; Cognitive Dysfunction; Cystine; Diet; East Asian People; Longitudinal Studies; Proline; Serine; Sleep; Sleep Duration; Surveys and Questionnaires; Dietary Proteins; Amino Acids; Eating; Middle Aged; Incidence; Aged; Aged, 80 and over; Independent Living; Diet Records; Dyssomnias
PubMed: 37821805
DOI: 10.1186/s12877-023-04359-2 -
Cancer Research Communications Nov 2023IFNγ alters the immunopeptidome presented on HLA class I (HLA-I), and its activity on cancer cells is known to be important for effective immunotherapy responses. We...
UNLABELLED
IFNγ alters the immunopeptidome presented on HLA class I (HLA-I), and its activity on cancer cells is known to be important for effective immunotherapy responses. We performed proteomic analyses of untreated and IFNγ-treated colorectal cancer patient-derived organoids and combined this with transcriptomic and HLA-I immunopeptidomics data to dissect mechanisms that lead to remodeling of the immunopeptidome through IFNγ. IFNγ-induced changes in the abundance of source proteins, switching from the constitutive to the immunoproteasome, and differential upregulation of different HLA alleles explained some, but not all, observed peptide abundance changes. By selecting for peptides which increased or decreased the most in abundance, but originated from proteins with limited abundance changes, we discovered that the amino acid composition of presented peptides also influences whether a peptide is upregulated or downregulated on HLA-I through IFNγ. The presence of proline within the peptide core was most strongly associated with peptide downregulation. This was validated in an independent dataset. Proline substitution in relevant core positions did not influence the predicted HLA-I binding affinity or stability, indicating that proline effects on peptide processing may be most relevant. Understanding the multiple factors that influence the abundance of peptides presented on HLA-I in the absence or presence of IFNγ is important to identify the best targets for antigen-specific cancer immunotherapies such as vaccines or T-cell receptor engineered therapeutics.
SIGNIFICANCE
IFNγ remodels the HLA-I-presented immunopeptidome. We showed that peptide-specific factors influence whether a peptide is upregulated or downregulated and identified a preferential loss or downregulation of those with proline near the peptide center. This will help selecting immunotherapy target antigens which are consistently presented by cancer cells.
Topics: Humans; Proteomics; Neoplasms; Interferon-gamma; Antigens; Peptides; Proline
PubMed: 37991387
DOI: 10.1158/2767-9764.CRC-23-0121 -
Plant Biotechnology (Tokyo, Japan) Dec 2023Turfgrasses show a wide range of salinity tolerance. In this study, twenty wild turfgrasses were collected from coastal regions in Japan, and their species; evolutionary...
Turfgrasses show a wide range of salinity tolerance. In this study, twenty wild turfgrasses were collected from coastal regions in Japan, and their species; evolutionary lineage; salt tolerance levels; shoot and root K, Na, and proline contents; and amounts of ions secreted from their salt glands were determined. Among them, eighteen turfgrass species were determined based on the internal transcribed spacer 1 sequences. All collected wild turfgrasses were identified as halophytes and were divided into two salt-tolerant levels. They maintained the shoot relative water contents and suppressed excess Na accumulation in their shoots and roots and K content homeostasis compared with rice, resulting in the maintenance of a higher K/Na ratio under salt stress. These characteristics must be part of the salt tolerance mechanisms. Among the four turfgrasses with salt glands, three selectively secreted Na from their salt glands; however, interestingly, one secreted K over Na, although it still maintained a K/Na ratio comparable to that of the other turfgrasses. A significant amount of proline synthesis was observed in most of the turfgrasses in response to salt stress, and the proline content was highly correlated with the salt tolerance, suggesting its key role in the salt tolerance mechanisms. These wild turfgrasses with such diverse ion control mechanisms and proline synthesis profiles are useful materials for investigating the salt tolerant mechanisms and breeding salt tolerant turfgrasses.
PubMed: 38434114
DOI: 10.5511/plantbiotechnology.23.0721a