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1.
Int J Mol Sci ; 25(5)2024 Mar 01.
Article in English | MEDLINE | ID: mdl-38474102

ABSTRACT

Histone deacetylase SIRT1 represses gene expression through the deacetylation of histones and transcription factors and is involved in the protective cell response to stress and aging. However, upon endoplasmic reticulum (ER) stress, SIRT1 impairs the IRE1α branch of the unfolded protein response (UPR) through the inhibition of the transcriptional activity of XBP-1 and SIRT1 deficiency is beneficial under these conditions. We hypothesized that SIRT1 deficiency may unlock the blockade of transcription factors unrelated to the UPR promoting the synthesis of chaperones and improving the stability of immature proteins or triggering the clearance of unfolded proteins. SIRT1+/+ and SIRT1-/- fibroblasts were exposed to the ER stress inducer tunicamycin and cell survival and expression of heat shock proteins were analyzed 24 h after the treatment. We observed that SIRT1 loss significantly reduced cell sensitivity to ER stress and showed that SIRT1-/- but not SIRT1+/+ cells constitutively expressed high levels of phospho-STAT3 and heat shock proteins. Hsp70 silencing in SIRT1-/- cells abolished the resistance to ER stress. Furthermore, accumulation of ubiquitinated proteins was lower in SIRT1-/- than in SIRT1+/+ cells. Our data showed that SIRT1 deficiency enabled chaperones upregulation and boosted the proteasome activity, two processes that are beneficial for coping with ER stress.


Subject(s)
Heat-Shock Proteins , Sirtuin 1 , Heat-Shock Proteins/metabolism , Up-Regulation , Sirtuin 1/metabolism , Endoribonucleases/metabolism , Protein Serine-Threonine Kinases/metabolism , Endoplasmic Reticulum Stress , Unfolded Protein Response , Molecular Chaperones/metabolism , Transcription Factors/metabolism
2.
Front Cell Dev Biol ; 11: 1293122, 2023.
Article in English | MEDLINE | ID: mdl-38020886

ABSTRACT

Pericentric heterochromatin (PCH) plays an essential role in the maintenance of genome integrity and alterations in PCH have been linked to cancer and aging. HP1 α, ß, and γ, are hallmarks of constitutive heterochromatin that are thought to promote PCH structure through binding to heterochromatin-specific histone modifications and interaction with a wide range of factors. Among the less understood components of PCH is the histone H2A variant H2A.Z, whose role in the organization and maintenance of PCH is poorly defined. Here we show that there is a complex interplay between H2A.Z and HP1 isoforms in PCH. While the loss of HP1α results in the accumulation of H2A.Z.1 in PCH, which is associated with a significant decrease in its mobile fraction, H2A.Z.1 binds preferentially to HP1ß in these regions. Of note, H2A.Z.1 downregulation results in increased heterochromatinization and instability of PCH, reflected by accumulation of the major epigenetic hallmarks of heterochromatin in these regions and increased frequency of chromosome aberrations related to centromeric/pericentromeric defects. Our studies support a role for H2A.Z in genome stability and unveil a key role of H2A.Z in the regulation of heterochromatin-specific epigenetic modifications through a complex interplay with the HP1 isoforms.

3.
Nucleic Acids Res ; 51(13): 6754-6769, 2023 07 21.
Article in English | MEDLINE | ID: mdl-37309898

ABSTRACT

The Sirtuin family of NAD+-dependent enzymes plays an important role in maintaining genome stability upon stress. Several mammalian Sirtuins have been linked directly or indirectly to the regulation of DNA damage during replication through Homologous recombination (HR). The role of one of them, SIRT1, is intriguing as it seems to have a general regulatory role in the DNA damage response (DDR) that has not yet been addressed. SIRT1-deficient cells show impaired DDR reflected in a decrease in repair capacity, increased genome instability and decreased levels of γH2AX. Here we unveil a close functional antagonism between SIRT1 and the PP4 phosphatase multiprotein complex in the regulation of the DDR. Upon DNA damage, SIRT1 interacts specifically with the catalytical subunit PP4c and promotes its inhibition by deacetylating the WH1 domain of the regulatory subunits PP4R3α/ß. This in turn regulates γH2AX and RPA2 phosphorylation, two key events in the signaling of DNA damage and repair by HR. We propose a mechanism whereby during stress, SIRT1 signaling ensures a global control of DNA damage signaling through PP4.


Subject(s)
DNA Damage , Sirtuin 1 , Animals , Humans , Mammals/metabolism , Phosphoric Monoester Hydrolases , Phosphorylation , Signal Transduction , Sirtuin 1/metabolism
4.
Aging (Albany NY) ; 13(9): 12308-12333, 2021 04 26.
Article in English | MEDLINE | ID: mdl-33901008

ABSTRACT

UV radiation is one of the main contributors to skin photoaging by promoting the accumulation of cellular senescence, which in turn induces a proinflammatory and tissue-degrading state that favors skin aging. The members of the sirtuin family of NAD+-dependent enzymes play an anti-senescence role and their activation suggests a promising approach for preventing UV-induced senescence in the treatment of skin aging. A two-step screening designed to identify compounds able to protect cells from UV-induced senescence through sirtuin activation identified shikimic acid (SA), a metabolic intermediate in many organisms, as a bona-fide candidate. The protective effects of SA against senescence were dependent on specific activation of SIRT1 as the effect was abrogated by the SIRT1 inhibitor EX-527. Upon UV irradiation SA induced S-phase accumulation and a decrease in p16INK4A expression but did not protect against DNA damage or increased polyploidies. In contrast, SA reverted misfolded protein accumulation upon senescence, an effect that was abrogated by EX-527. Consistently, SA induced an increase in the levels of the chaperone BiP, resulting in a downregulation of unfolded protein response (UPR) signaling and UPR-dependent autophagy, avoiding their abnormal hyperactivation during senescence. SA did not directly activate SIRT1 in vitro, suggesting that SIRT1 is a downstream effector of SA signaling specifically in the response to cellular senescence. Our study not only uncovers a shikimic acid/SIRT1 signaling pathway that prevents cellular senescence, but also reinforces the role of sirtuins as key regulators of cell proteostasis.


Subject(s)
NAD/drug effects , Shikimic Acid/pharmacology , Sirtuin 1/drug effects , Skin Aging/drug effects , Cell Proliferation/drug effects , Cellular Senescence/physiology , Humans , NAD/metabolism , Oxidative Stress/drug effects , Protective Agents/pharmacology , Signal Transduction/drug effects , Sirtuin 1/metabolism , Skin/drug effects , Skin/metabolism , Ultraviolet Rays/adverse effects
5.
Nat Commun ; 9(1): 101, 2018 01 09.
Article in English | MEDLINE | ID: mdl-29317652

ABSTRACT

Sirtuins are NAD+-dependent deacetylases that facilitate cellular stress response. They include SirT6, which protects genome stability and regulates metabolic homeostasis through gene silencing, and whose loss induces an accelerated aging phenotype directly linked to hyperactivation of the NF-κB pathway. Here we show that SirT6 binds to the H3K9me3-specific histone methyltransferase Suv39h1 and induces monoubiquitination of conserved cysteines in the PRE-SET domain of Suv39h1. Following activation of NF-κB signaling Suv39h1 is released from the IκBα locus, subsequently repressing the NF-κB pathway. We propose that SirT6 attenuates the NF-κB pathway through IκBα upregulation via cysteine monoubiquitination and chromatin eviction of Suv39h1. We suggest a mechanism based on SirT6-mediated enhancement of a negative feedback loop that restricts the NF-κB pathway.


Subject(s)
Cysteine/metabolism , Methyltransferases/metabolism , NF-kappa B/metabolism , PR-SET Domains , Repressor Proteins/metabolism , Sirtuins/metabolism , Animals , Cell Line, Tumor , Cells, Cultured , Chromatin/metabolism , Cysteine/genetics , HCT116 Cells , HEK293 Cells , HeLa Cells , Humans , Methyltransferases/genetics , Mice , NF-KappaB Inhibitor alpha/metabolism , NIH 3T3 Cells , Protein Binding , Repressor Proteins/genetics , Signal Transduction , Sirtuins/genetics , Ubiquitination , Up-Regulation
6.
Epigenetics ; 12(2): 166-175, 2017 02.
Article in English | MEDLINE | ID: mdl-28059589

ABSTRACT

The presence of H3K9me3 and heterochromatin protein 1 (HP1) are hallmarks of heterochromatin conserved in eukaryotes. The spreading and maintenance of H3K9me3 is effected by the functional interplay between the H3K9me3-specific histone methyltransferase Suv39h1 and HP1. This interplay is complex in mammals because the three HP1 isoforms, HP1α, ß, and γ, are thought to play a redundant role in Suv39h1-dependent deposition of H3K9me3 in pericentric heterochromatin (PCH). Here, we demonstrate that despite this redundancy, HP1α and, to a lesser extent, HP1γ have a closer functional link to Suv39h1, compared to HP1ß. HP1α and γ preferentially interact in vivo with Suv39h1, regulate its dynamics in heterochromatin, and increase Suv39h1 protein stability through an inhibition of MDM2-dependent Suv39h1-K87 polyubiquitination. The reverse is also observed, where Suv39h1 increases HP1α stability compared HP1ß and γ. The interplay between Suv39h1 and HP1 isoforms appears to be relevant under genotoxic stress. Specifically, loss of HP1α and γ isoforms inhibits the upregulation of Suv39h1 and H3K9me3 that is observed under stress conditions. Reciprocally, Suv39h1 deficiency abrogates stress-dependent upregulation of HP1α and γ, and enhances HP1ß levels. Our work defines a specific role for HP1 isoforms in regulating Suv39h1 function under stress via a feedback mechanism that likely regulates heterochromatin formation.


Subject(s)
Chromosomal Proteins, Non-Histone/metabolism , DNA Damage , Feedback, Physiological , Methyltransferases/genetics , Repressor Proteins/genetics , Cell Line , Chromatin Assembly and Disassembly , Chromobox Protein Homolog 5 , Chromosomal Proteins, Non-Histone/genetics , Histones/metabolism , Humans , Methyltransferases/metabolism , Protein Binding , Protein Isoforms/genetics , Protein Isoforms/metabolism , Protein Stability , Repressor Proteins/metabolism , Ubiquitination
7.
Genes Dev ; 27(6): 639-53, 2013 Mar 15.
Article in English | MEDLINE | ID: mdl-23468428

ABSTRACT

The establishment of the epigenetic mark H4K20me1 (monomethylation of H4K20) by PR-Set7 during G2/M directly impacts S-phase progression and genome stability. However, the mechanisms involved in the regulation of this event are not well understood. Here we show that SirT2 regulates H4K20me1 deposition through the deacetylation of H4K16Ac (acetylation of H4K16) and determines the levels of H4K20me2/3 throughout the cell cycle. SirT2 binds and deacetylates PR-Set7 at K90, modulating its chromatin localization. Consistently, SirT2 depletion significantly reduces PR-Set7 chromatin levels, alters the size and number of PR-Set7 foci, and decreases the overall mitotic deposition of H4K20me1. Upon stress, the interaction between SirT2 and PR-Set7 increases along with the H4K20me1 levels, suggesting a novel mitotic checkpoint mechanism. SirT2 loss in mice induces significant defects associated with defective H4K20me1-3 levels. Accordingly, SirT2-deficient animals exhibit genomic instability and chromosomal aberrations and are prone to tumorigenesis. Our studies suggest that the dynamic cross-talk between the environment and the genome during mitosis determines the fate of the subsequent cell cycle.


Subject(s)
Cell Cycle/physiology , Genomic Instability , Sirtuin 2/metabolism , Acetylation , Amino Acid Sequence , Animals , Cell Transformation, Neoplastic/genetics , Chromatin/metabolism , DNA Damage/genetics , Gene Knockout Techniques , HeLa Cells , Histone-Lysine N-Methyltransferase/metabolism , Histones/metabolism , Humans , M Phase Cell Cycle Checkpoints/physiology , Methylation , Mice , Mice, Knockout , Mitosis , Protein Binding , Sirtuin 2/genetics
8.
Mol Cell ; 42(2): 210-23, 2011 Apr 22.
Article in English | MEDLINE | ID: mdl-21504832

ABSTRACT

Sirtuins are NAD-dependent deacetylases that sense oxidative stress conditions and promote a protective cellular response. The Sirtuin SirT1 is involved in facultative heterochromatin formation through an intimate functional relationship with the H3K9me3 methyltransferase Suv39h1, a chromatin organization protein. However, SirT1 also regulates Suv39h1-dependent constitutive heterochromatin (CH) through an unknown mechanism; interestingly, SirT1 does not significantly localize in these regions. Herein, we report that SirT1 controls global levels of Suv39h1 by increasing its half-life through inhibition of Suv39h1 lysine 87 polyubiquitination by the E3-ubiquitin ligase MDM2. This in turn increases Suv39h1 turnover in CH and ensures genome integrity. Stress conditions that lead to SirT1 upregulation, such as calorie restriction, also induce higher levels of Suv39h1 in a SirT1-dependent manner in vivo. These observations reflect a direct link between oxidative stress response and Suv39h1 and support a dynamic view of heterochromatin, in which its structure adapts to cell physiology.


Subject(s)
Chromatin Assembly and Disassembly , Genomic Instability , Heterochromatin/metabolism , Methyltransferases/metabolism , Oxidative Stress , Repressor Proteins/metabolism , Sirtuin 1/metabolism , Amino Acid Sequence , Animals , Caloric Restriction , Enzyme Stability , HEK293 Cells , Half-Life , HeLa Cells , Humans , Lysine , Male , Methyltransferases/genetics , Mice , Molecular Sequence Data , Mutation , NIH 3T3 Cells , Polyubiquitin/metabolism , Proto-Oncogene Proteins c-mdm2/metabolism , RNA Interference , Rats , Rats, Sprague-Dawley , Recombinant Fusion Proteins/metabolism , Repressor Proteins/genetics , Sirtuin 1/genetics , Time Factors , Transfection , Ubiquitination
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