| 1. |
- Bisteau, Xavier, et al.
(author)
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The Greatwall kinase safeguards the genome integrity by affecting the kinome activity in mitosis
- 2020
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In: Oncogene. - : Springer Science and Business Media LLC. - 0950-9232 .- 1476-5594.
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Journal article (peer-reviewed)abstract
- Progression through mitosis is balanced by the timely regulation of phosphorylation and dephosphorylation events ensuring the correct segregation of chromosomes before cytokinesis. This balance is regulated by the opposing actions of CDK1 and PP2A, as well as the Greatwall kinase/MASTL. MASTL is commonly overexpressed in cancer, which makes it a potential therapeutic anticancer target. Loss of Mastl induces multiple chromosomal errors that lead to the accumulation of micronuclei and multilobulated cells in mitosis. Our analyses revealed that loss of Mastl leads to chromosome breaks and abnormalities impairing correct segregation. Phospho-proteomic data for Mastl knockout cells revealed alterations in proteins implicated in multiple processes during mitosis including double-strand DNA damage repair. In silico prediction of the kinases with affected activity unveiled NEK2 to be regulated in the absence of Mastl. We uncovered that, RAD51AP1, involved in regulation of homologous recombination, is phosphorylated by NEK2 and CDK1 but also efficiently dephosphorylated by PP2A/B55. Our results suggest that MastlKO disturbs the equilibrium of the mitotic phosphoproteome that leads to the disruption of DNA damage repair and triggers an accumulation of chromosome breaks even in noncancerous cells.
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| 2. |
- Caldez, Matias J, et al.
(author)
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Cell cycle regulation in NAFLD: when imbalanced metabolism limits cell division : when imbalanced metabolism limits cell division
- 2020
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In: Hepatology international. - : Springer Science and Business Media LLC. - 1936-0533 .- 1936-0541. ; 14:4, s. 463-474
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Research review (peer-reviewed)abstract
- Cell division is essential for organismal growth and tissue homeostasis. It is exceptionally significant in tissues chronically exposed to intrinsic and external damage, like the liver. After decades of studying the regulation of cell cycle by extracellular signals, there are still gaps in our knowledge on how these two interact with metabolic pathways in vivo. Studying the cross-talk of these pathways has direct clinical implications as defects in cell division, signaling pathways, and metabolic homeostasis are frequently observed in liver diseases. In this review, we will focus on recent reports which describe various functions of cell cycle regulators in hepatic homeostasis. We will describe the interplay between the cell cycle and metabolism during liver regeneration after acute and chronic damage. We will focus our attention on non-alcoholic fatty liver disease, especially non-alcoholic steatohepatitis. The global incidence of non-alcoholic fatty liver disease is increasing exponentially. Therefore, understanding the interplay between cell cycle regulators and metabolism may lead to the discovery of novel therapeutic targets amenable to intervention.
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| 3. |
- Colon-Mesa, Ignacio, et al.
(author)
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p27Kip1 Deficiency Impairs Brown Adipose Tissue Function Favouring Fat Accumulation in Mice
- 2023
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In: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 24:3
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Journal article (peer-reviewed)abstract
- The aim of this work was to investigate the effect of the whole-body deletion of p27 on the activity of brown adipose tissue and the susceptibility to develop obesity and glucose homeostasis disturbances in mice, especially when subjected to a high fat diet. p27 knockout (p27−/−) and wild type (WT) mice were fed a normal chow diet or a high fat diet (HFD) for 10-weeks. Body weight and composition were assessed. Insulin and glucose tolerance tests and indirect calorimetry assays were performed. Histological analysis of interscapular BAT (iBAT) was carried out, and expression of key genes/proteins involved in BAT function were characterized by qPCR and Western blot. iBAT activity was estimated by 18F-fluorodeoxyglucose (18FDG) uptake with microPET. p27−/− mice were more prone to develop obesity and insulin resistance, exhibiting increased size of all fat depots. p27−/− mice displayed a higher respiratory exchange ratio. iBAT presented larger adipocytes in p27−/− HFD mice, accompanied by downregulation of both Glut1 and uncoupling protein 1 (UCP1) in parallel with defective insulin signalling. Moreover, p27−/− HFD mice exhibited impaired response to cold exposure, characterized by a reduced iBAT 18FDG uptake and difficulty to maintain body temperature when exposed to cold compared to WT HFD mice, suggesting reduced thermogenic capacity. These data suggest that p27 could play a role in BAT activation and in the susceptibility to develop obesity and insulin resistance.
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| 4. |
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| 5. |
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| 6. |
- Kaldis, Philipp, et al.
(author)
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Molecular basis of the reaction mechanism of the methyltransferase HENMT1
- 2024
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In: PLoS ONE. - 1932-6203. ; 19:1
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Journal article (peer-reviewed)abstract
- PIWI-interacting RNAs (piRNAs) are important for ensuring the integrity of the germline. 3'-terminal 2'-O-methylation is essential for piRNA maturation and to protect them from degradation. HENMT1 (HEN Methyltransferase 1) carries out the 2'-O-methylation, which is of key importance for piRNA stability and functionality. However, neither the structure nor the catalytic mechanism of mammalian HENMT1 have been studied. We have constructed a catalytic-competent HENMT1 complex using computational approaches, in which Mg2+ is primarily coordinated by four evolutionary conserved residues, and is further auxiliary coordinated by the 3'-O and 2'-O on the 3'-terminal nucleotide of the piRNA. Our study suggests that metal has limited effects on substrate and cofactor binding but is essential for catalysis. The reaction consists of deprotonation of the 2'-OH to 2'-O and a methyl transfer from SAM to the 2'-O. The methyl transfer is spontaneous and fast. Our in-depth analysis suggests that the 2'-OH may be deprotonated before entering the active site or it may be partially deprotonated at the active site by His800 and Asp859, which are in a special alignment that facilitates the proton transfer out of the active site. Furthermore, we have developed a detailed potential reaction scenario indicating that HENMT1 is Mg2+ utilizing but is not a Mg2+ dependent enzyme.
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| 7. |
- Keles, Umur, et al.
(author)
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Liver-derived metabolites as signaling molecules in fatty liver disease
- 2023
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In: Cellular and Molecular Life Sciences. - : Springer Science and Business Media LLC. - 1420-9071 .- 1420-682X. ; 80:1, s. 1-14
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Research review (peer-reviewed)abstract
- Excessive fat accumulation in the liver has become a major health threat worldwide. Unresolved fat deposition in the liver can go undetected until it develops into fatty liver disease, followed by steatohepatitis, fibrosis, cirrhosis, and eventually hepatocellular carcinoma. Lipid deposition in the liver is governed by complex communication, primarily between metabolic organs. This can be mediated by hormones, organokines, and also, as has been more recently discovered, metabolites. Although how metabolites from peripheral organs affect the liver is well documented, the effect of metabolic players released from the liver during the development of fatty liver disease or associated comorbidities needs further attention. Here we focus on interorgan crosstalk based on metabolites released from the liver and how these molecules act as signaling molecules in peripheral tissues. Due to the liver's specific role, we are covering lipid and bile mechanism-derived metabolites. We also discuss the high sucrose intake associated with uric acid release from the liver. Excessive fat deposition in the liver during fatty liver disease development reflects disrupted metabolic processes. As a response, the liver secretes a variety of signaling molecules as well as metabolites which act as a footprint of the metabolic disruption. In the coming years, the reciprocal exchange of metabolites between the liver and other metabolic organs will gain further importance and will help to better understand the development of fatty liver disease and associated diseases.
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| 8. |
- Na Zhao, Li, et al.
(author)
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Cascading proton transfers are a hallmark of the catalytic mechanism of SAM-dependent methyltransferases
- 2020
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In: FEBS Letters. - : Wiley. - 1873-3468 .- 0014-5793. ; 594:13, s. 2128-2139
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Journal article (peer-reviewed)abstract
- The S-adenosyl-L-methionine (SAM)-dependent methyltransferases attach a methyl group to the deprotonated methyl lysine (Kme0) using SAM as a donor. An intriguing, yet unanswered, question is how the deprotonation of the methyl lysine takes place which results in a lone pair of electrons at the Nϵ atom of the methyl lysine for the following methyl transfer. PRDM9, one of the few methyltransferases with well-defined enzyme activity in vitro and in vivo, is a good representative of the PR/SET domain methyltransferase family to study the deprotonation and subsequently the methyl transfer. The reaction consists of two progressing steps: (i) the absolutely required substrate methyl lysine deprotonation and (ii) the transfer of the methyl group to the deprontonated methyl lysine. We use empirical valence bond (EVB) simulations to evaluate Y357 at the active site as potential general base for the deprotonation of the methyl lysine. Indeed, our study has found that the pKa of Tyr357 is low enough to make it an ideal candidate for proton abstraction from the methyl lysine. The partially deprontonated Tyr357 is able to change its H-bond pattern thus bridging two proton tunneling states (OH- H 0-Tyr357 and Kme0-Nϵ H O-Tyr357) and providing a cascading proton transfer from Tyr357 to hydroxide, generating deprotonated Tyr357 and then from Kme0 to the deprotonated Tyr357 resulting in deprotonated methyl lysine. This cascading proton transfer shortens the lifespan of the labile intermediates, and affects the conformational changes during the product release important to promote the proton release to the bulk solvent. Our computational efforts have uncovered a new catalytic mechanism to unravel the unanswered question about the deprotonation of the methyl lysine in methyltransferases.
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| 9. |
- Narayanaswamy, Pradeep, et al.
(author)
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MetaboKit : a comprehensive data extraction tool for untargeted metabolomics
- 2020
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In: Molecular omics. - : Royal Society of Chemistry (RSC). - 2515-4184. ; 16:5, s. 436-447
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Journal article (peer-reviewed)abstract
- We have developed MetaboKit, a comprehensive software package for compound identification and relative quantification in mass spectrometry-based untargeted metabolomics analysis. In data dependent acquisition (DDA) analysis, MetaboKit constructs a customized spectral library with compound identities from reference spectral libraries, adducts, dimers, in-source fragments (ISF), MS/MS fragmentation spectra, and more importantly the retention time information unique to the chromatography system used in the experiment. Using the customized library, the software performs targeted peak integration for precursor ions in DDA analysis and for precursor and product ions in data independent acquisition (DIA) analysis. With its stringent identification algorithm requiring matches by both MS and MS/MS data, MetaboKit provides identification results with significantly greater specificity than the competing software packages without loss in sensitivity. The proposed MS/MS-based screening of ISFs also reduces the chance of unverifiable identification of ISFs considerably. MetaboKit's quantification module produced peak area values highly correlated with known concentrations in a DIA analysis of the metabolite standards at both MS1 and MS2 levels. Moreover, the analysis of Cdk1Liv-/- mouse livers showed that MetaboKit can identify a wide range of lipid species and their ISFs, and quantitatively reconstitute the well-characterized fatty liver phenotype in these mice. In DIA data, the MS1-level and MS2-level peak area data produced similar fold change estimates in the differential abundance analysis, and the MS2-level peak area data allowed for quantitative comparisons in compounds whose precursor ion chromatogram was too noisy for peak integration.
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| 10. |
- Ow, Jin Rong, et al.
(author)
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Remodelling of whole-body lipid metabolism and a diabetic-like phenotype caused by loss of CDK1 and hepatocyte division
- 2020
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In: eLife. - 2050-084X. ; 9
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Journal article (peer-reviewed)abstract
- Cell cycle progression and lipid metabolism are well-coordinated processes required for proper cell proliferation. In liver diseases that arise from dysregulated lipid metabolism, proliferation is diminished. To study the outcome of CDK1 loss and blocked hepatocyte proliferation on lipid metabolism and the consequent impact on whole-body physiology, we performed lipidomics, metabolomics, and RNA-seq analyses on a mouse model. We observed reduced triacylglycerides in liver of young mice, caused by oxidative stress that activated FOXO1 to promote expression of ATGL. Additionally, we discovered that hepatocytes displayed malfunctioning b-oxidation, reflected by increased acylcarnitines and reduced b-hydroxybutyrate. This led to elevated plasma free fatty acids, which were transported to the adipose tissue for storage and triggered greater insulin secretion. Upon aging, chronic hyperinsulinemia resulted in insulin resistance and hepatic steatosis through activation of LXR. Here we demonstrate that loss of hepatocyte proliferation is not only an outcome but possibly causative for liver pathology.
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