NMNAT promotes glioma growth through regulating post-translational modifications of P53 to inhibit apoptosis

Gliomas are highly malignant brain tumors with poor prognosis and short survival. NAD+ has been shown to impact multiple processes that are dysregulated in cancer; however, anti-cancer therapies targeting NAD+ synthesis have had limited success due to insufficient mechanistic understanding. Here, we adapted a Drosophila glial neoplasia model and discovered the genetic requirement for NAD+ synthase nicotinamide mononucleotide adenylyltransferase (NMNAT) in glioma progression in vivo and in human glioma cells. Overexpressing enzymatically active NMNAT significantly promotes glial neoplasia growth and reduces animal viability. Mechanistic analysis suggests that NMNAT interferes with DNA damage-p53-caspase-3 apoptosis signaling pathway by enhancing NAD+-dependent posttranslational modifications (PTMs) poly(ADP-ribosyl)ation (PARylation) and deacetylation of p53. Since PARylation and deacetylation reduce p53 pro-apoptotic activity, modulating p53 PTMs could be a key mechanism by which NMNAT promotes glioma growth. Our findings reveal a novel tumorigenic mechanism involving protein complex formation of p53 with NAD+ synthetic enzyme NMNAT and NAD+-dependent PTM enzymes that regulates glioma growth.

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NMNAT promotes glioma growth through regulating post-translational modifications of p53 to inhibit apoptosis

10.1101/2021.05.12.443736 ◽

2021 ◽

Author(s):

Rong Grace Zhai ◽

Jiaqi Liu ◽

Yi Zhu ◽

Xianzun Tao ◽

Kai Ruan ◽

...

Keyword(s):

Posttranslational Modifications ◽

Cancer Therapies ◽

Post Translational Modifications ◽

Nicotinamide Mononucleotide ◽

Nad Synthesis ◽

Glioma Growth ◽

Tumorigenic Mechanism ◽

Apoptotic Activity ◽

Glioma Progression

Gliomas are highly malignant brain tumors with poor prognosis and short survival. NAD+ has been shown to impact multiple processes that are dysregulated in cancer; however, anti-cancer therapies targeting NAD+ synthesis have been unsuccessful due to insufficient mechanistic understanding. Here we adapted a Drosophila glial neoplasia model and discovered the genetic requirement for NAD+ synthase nicotinamide mononucleotide adenylyltransferase (NMNAT) in glioma progression in vivo and in human glioma cells. Overexpressing enzymatically active NMNAT significantly promotes glial neoplasia growth and reduces animal viability. Mechanistic analysis suggests that NMNAT interferes with DNA damage-p53-caspase-3 apoptosis signaling pathway by enhancing NAD+-dependent posttranslational modifications (PTMs) poly(ADP-ribosyl)ation (PARylation) and deacetylation of p53. Interestingly, NMNAT forms a complex with p53 and PTM enzyme PARP1 to facilitate PARylation. As PARylation and deacetylation reduce p53 pro-apoptotic activity, our results demonstrate that NMNAT promotes glioma progression through regulating p53 post-translational modifications. Our findings reveal a novel tumorigenic mechanism involving protein complex formation of p53 with NAD+ synthetic enzyme NMNAT and NAD+-dependent PTM enzymes that regulates glioma growth.

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Phytosomal Curcumin Elicits Anti-tumor Properties Through Suppression of Angiogenesis, Cell Proliferation and Induction of Oxidative Stress in Colorectal Cancer

Current Pharmaceutical Design ◽

10.2174/1381612825666190110145151 ◽

2019 ◽

Vol 24 (39) ◽

pp. 4626-4638 ◽

Cited By ~ 13

Author(s):

Reyhaneh Moradi-Marjaneh ◽

Seyed M. Hassanian ◽

Farzad Rahmani ◽

Seyed H. Aghaee-Bakhtiari ◽

Amir Avan ◽

...

Keyword(s):

Oxidative Stress ◽

Colorectal Cancer ◽

Therapeutic Potential ◽

Vegf Signaling ◽

Anticancer Effects ◽

Inhibition Of Angiogenesis ◽

Underlying Mechanisms ◽

Apoptotic Activity

Background: Colorectal cancer (CRC) is one of the most common causes of cancer-associated mortality in the world. Anti-tumor effect of curcumin has been shown in different cancers; however, the therapeutic potential of novel phytosomal curcumin, as well as the underlying molecular mechanism in CRC, has not yet been explored. Methods: The anti-proliferative, anti-migratory and apoptotic activity of phytosomal curcumin in CT26 cells was assessed by MTT assay, wound healing assay and Flow cytometry, respectively. Phytosomal curcumin was also tested for its in-vivo activity in a xenograft mouse model of CRC. In addition, oxidant/antioxidant activity was examined by DCFH-DA assay in vitro, measurement of malondialdehyde (MDA), Thiol and superoxidedismutase (SOD) and catalase (CAT) activity and also evaluation of expression levels of Nrf2 and GCLM by qRT-PCR in tumor tissues. In addition, the effect of phytosomal curcumin on angiogenesis was assessed by the measurement of VEGF-A and VEGFR-1 and VEGF signaling regulatory microRNAs (miRNAs) in tumor tissue. Results: Phytosomal curcumin exerts anti-proliferative, anti-migratory and apoptotic activity in-vitro. It also decreases tumor growth and augmented 5-fluorouracil (5-FU) anti-tumor effect in-vivo. In addition, our data showed that induction of oxidative stress and inhibition of angiogenesis through modulation of VEGF signaling regulatory miRNAs might be underlying mechanisms by which phytosomal curcumin exerted its antitumor effect. Conclusion: Our data confirmed this notion that phytosomal curcumin administrates anticancer effects and can be used as a complementary treatment in clinical settings.

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Aryl Hydrocarbon Receptor: Its Regulation and Roles in Transformation and Tumorigenesis

Current Drug Targets ◽

10.2174/1389450120666181109092225 ◽

2019 ◽

Vol 20 (6) ◽

pp. 625-634 ◽

Cited By ~ 1

Author(s):

Xun Che ◽

Wei Dai

Keyword(s):

Target Gene ◽

Tumor Development ◽

Environmental Response ◽

Carcinogenic Effect ◽

Post Translational Modifications ◽

Downstream Target ◽

Aryl Hydrocarbon ◽

Downstream Target Gene ◽

Major Mediator

AhR is an environmental response gene that mediates cellular responses to a variety of xenobiotic compounds that frequently function as AhR ligands. Many AhR ligands are classified as carcinogens or pro-carcinogens. Thus, AhR itself acts as a major mediator of the carcinogenic effect of many xenobiotics in vivo. In this concise review, mechanisms by which AhR trans-activates downstream target gene expression, modulates immune responses, and mediates malignant transformation and tumor development are discussed. Moreover, activation of AhR by post-translational modifications and crosstalk with other transcription factors or signaling pathways are also summarized.

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Effect of Posttranslational Modifications on the Structure and Activity of FTO Demethylase

International Journal of Molecular Sciences ◽

10.3390/ijms22094512 ◽

2021 ◽

Vol 22 (9) ◽

pp. 4512

Author(s):

Michał Marcinkowski ◽

Tomaš Pilžys ◽

Damian Garbicz ◽

Jan Piwowarski ◽

Damian Mielecki ◽

...

Keyword(s):

Catalytic Activity ◽

Size Exclusion Chromatography ◽

Posttranslational Modifications ◽

Size Exclusion ◽

Saxs Data ◽

Wide Range ◽

Exclusion Chromatography ◽

Demethylase Activity ◽

Structure And Activity

The FTO protein is involved in a wide range of physiological processes, including adipogenesis and osteogenesis. This two-domain protein belongs to the AlkB family of 2-oxoglutarate (2-OG)- and Fe(II)-dependent dioxygenases, displaying N6-methyladenosine (N6-meA) demethylase activity. The aim of the study was to characterize the relationships between the structure and activity of FTO. The effect of cofactors (Fe2+/Mn2+ and 2-OG), Ca2+ that do not bind at the catalytic site, and protein concentration on FTO properties expressed in either E. coli (ECFTO) or baculovirus (BESFTO) system were determined using biophysical methods (DSF, MST, SAXS) and biochemical techniques (size-exclusion chromatography, enzymatic assay). We found that BESFTO carries three phosphoserines (S184, S256, S260), while there were no such modifications in ECFTO. The S256D mutation mimicking the S256 phosphorylation moderately decreased FTO catalytic activity. In the presence of Ca2+, a slight stabilization of the FTO structure was observed, accompanied by a decrease in catalytic activity. Size exclusion chromatography and MST data confirmed the ability of FTO from both expression systems to form homodimers. The MST-determined dissociation constant of the FTO homodimer was consistent with their in vivo formation in human cells. Finally, a low-resolution structure of the FTO homodimer was built based on SAXS data.

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E3 ligase Nedd4l promotes antiviral innate immunity by catalyzing K29-linked cysteine ubiquitination of TRAF3

Nature Communications ◽

10.1038/s41467-021-21456-1 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Peng Gao ◽

Xianwei Ma ◽

Ming Yuan ◽

Yulan Yi ◽

Guoke Liu ◽

...

Keyword(s):

Posttranslational Modifications ◽

Type I Interferon ◽

Zinc Fingers ◽

E3 Ligase ◽

Type I ◽

E3 Ligases ◽

Protein Posttranslational Modifications ◽

Antiviral Innate Immunity

AbstractUbiquitination is one of the most prevalent protein posttranslational modifications. Here, we show that E3 ligase Nedd4l positively regulates antiviral immunity by catalyzing K29-linked cysteine ubiquitination of TRAF3. Deficiency of Nedd4l significantly impairs type I interferon and proinflammatory cytokine production induced by virus infection both in vitro and in vivo. Nedd4l deficiency inhibits virus-induced ubiquitination of TRAF3, the binding between TRAF3 and TBK1, and subsequent phosphorylation of TBK1 and IRF3. Nedd4l directly interacts with TRAF3 and catalyzes K29-linked ubiquitination of Cys56 and Cys124, two cysteines that constitute zinc fingers, resulting in enhanced association between TRAF3 and E3 ligases, cIAP1/2 and HECTD3, and also increased K48/K63-linked ubiquitination of TRAF3. Mutation of Cys56 and Cys124 diminishes Nedd4l-catalyzed K29-linked ubiquitination, but enhances association between TRAF3 and the E3 ligases, supporting Nedd4l promotes type I interferon production in response to virus by catalyzing ubiquitination of the cysteines in TRAF3.

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TMOD-24. GENERATION AND CHARACTERIZATION OF A NOVEL TRANSGENIC IDO REPORTER MOUSE FOR IDO POSTTRANSLATIONAL MODIFICATION ANALYSIS IN SITU

Neuro-Oncology ◽

10.1093/neuonc/noaa215.974 ◽

2020 ◽

Vol 22 (Supplement_2) ◽

pp. ii233-ii233

Author(s):

April Bell ◽

Lijie Zhai ◽

Erik Ladomersky ◽

Kristen Lauing ◽

Lakshmi Bollu ◽

...

Keyword(s):

Mouse Model ◽

Tumor Cell ◽

Central Nervous System Tumor ◽

Post Translational Modifications ◽

Reporter Mouse ◽

C Terminus ◽

Human Gbm

Abstract Glioblastoma (GBM) is the most common and aggressive primary central nervous system tumor in adults with a median survival of 14.6 months. GBM is a potently immunosuppressive cancer due in-part to the prolific expression of immunosuppressive indoleamine 2,3 dioxygenase 1 (IDO). Tumor cell IDO facilitates the intratumoral accumulation of regulatory T cells (Tregs; CD4+CD25+FoxP3+). Although immunosuppressive IDO activity is canonically characterized by the conversion of tryptophan into kynurenine, we have utilized transgenic and syngeneic mouse models and mutant glioma lines to demonstrate that tumor cell IDO increases Treg accumulation independent of tryptophan metabolism. Here, we address the gap in our understanding of IDO signaling activity in vivo. Subcutaneously-engrafted human GBM expressing human IDO-GFP cDNA was isolated from immunodeficient humanized NSG-SGM3 mice. The tumor was immunoprecipitated for the GFP tag using GFP-TRAP followed by mass spectrometry which revealed a novel methylation site on a lysine residue at amino acid 373 in the IDO C-terminus region. Western blot analysis of IDO protein also revealed the presence of tyrosine phosphorylation. Additionally, we recently created a new transgenic IDO reporter mouse model whereby endogenous IDO is fused to GFP via a T2A linker (IDO→GFP). This model allows for the isolation of IDO+ cells in real-time and without causing cell death, thereby creating the opportunity for downstream molecular analysis of in situ-isolated GFP+ cells. Collectively, our work suggests that IDO non-enzyme activity may involve the post-translational modifications we recently identified. As IDO activity may differ between in vitro and in vivo modeling systems, we will use the new IDO→GFP reporter mouse model for an improved mechanistic understanding of how immunosuppressive IDO facilitates Treg accumulation in vivo.

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Satureja subspicata and S. horvatii Extracts Induce Overexpression of the BCl-2 Family of Anti-apoptotic Genes and Reduce Micronuclei Frequency in Mice

Natural Product Communications ◽

10.1177/1934578x1801300617 ◽

2018 ◽

Vol 13 (6) ◽

pp. 1934578X1801300

Author(s):

Jasmina Čakar ◽

Naida Kadrić Lojo ◽

Anja Haverić ◽

Maida Hadžić ◽

Lejla Lasić ◽

...

Keyword(s):

Human Lymphocyte ◽

Micronucleus Assay ◽

Balkan Peninsula ◽

Lymphocyte Culture ◽

Cytotoxic Activities ◽

Cytotoxic Effects ◽

Apoptotic Genes ◽

Apoptotic Activity

Satureja subspicata and S. horvatii are endemic species of the Balkan Peninsula and often used in traditional medicine in Bosnia and Herzegovina to treat different health conditions. We aimed to analyze the unevaluated apoptotic, genotoxic and cytotoxic effects of two Satureja species, as well as their content of phenolics that are mainly responsible for the plant's biological activity. Apoptotic and geno/cytotoxic activities of S. subspicata and S. horvatii were investigated in vitro in human lymphocyte culture and in vivo in mice. The content of the main phenolics in plant extracts was determined by ultra-high pressure liquid chromatography-MS-MS (UHPLC–MS/MS). Genotoxic and cytotoxic activities of Satureja extracts were evaluated in vitro by applying a cytokinesis-block micronucleus cytome assay in human lymphocyte culture and in vivo applying a mice reticulocytes micronucleus assay. SALSA RT-MLPA R011-C1 apoptosis assay was used for measuring the relative expression of 44 genes associated with the regulation of the apoptotic pathways in human lymphocyte cultures treated with different concentrations of two Satureja extracts. The first analysis of phenolic compounds in S. horvatii and S. subspicata determined by an UHPLC-MS/MS method revealed high levels of rosmarinic and caffeic acids. Minor genotoxic potential was determined in relation to the tested concentrations while no cytostatic and cytotoxic effects were revealed in vitro. However, when applied in concentrations of 200 mg/kg per os, aqueous extracts of two Satureja species significantly decreased frequency of reticulocytes micronuclei in treated mice against controls. Extracts of S. subspicata and S. horvatii in concentrations of 0.2 mg/mL, regardless of solvent used, downregulated pro-apoptotic and upregulated anti-apoptotic genes, showing anti-apoptotic activity. Our results indicate that the registered anti-genotoxic and anti-apoptotic activity is most likely related to the high level of phenolic acids (particularly rosmarinic and caffeic) in the tested extracts.

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Spanning the gap: unraveling RSC dynamics in vivo

Current Genetics ◽

10.1007/s00294-020-01144-1 ◽

2021 ◽

Author(s):

Heinz Neumann ◽

Bryan J. Wilkins

Keyword(s):

Cell Cycle ◽

Posttranslational Modifications ◽

Transcriptional Activation ◽

Binding Mode ◽

Binding Modes ◽

Binding Domains ◽

Binding Interface ◽

The Many ◽

And Function

AbstractMultiple reports over the past 2 years have provided the first complete structural analyses for the essential yeast chromatin remodeler, RSC, providing elaborate molecular details for its engagement with the nucleosome. However, there still remain gaps in resolution, particularly within the many RSC subunits that harbor histone binding domains.Solving contacts at these interfaces is crucial because they are regulated by posttranslational modifications that control remodeler binding modes and function. Modifications are dynamic in nature often corresponding to transcriptional activation states and cell cycle stage, highlighting not only a need for enriched spatial resolution but also temporal understanding of remodeler engagement with the nucleosome. Our recent work sheds light on some of those gaps by exploring the binding interface between the RSC catalytic motor protein, Sth1, and the nucleosome, in the living nucleus. Using genetically encoded photo-activatable amino acids incorporated into histones of living yeast we are able to monitor the nucleosomal binding of RSC, emphasizing the regulatory roles of histone modifications in a spatiotemporal manner. We observe that RSC prefers to bind H2B SUMOylated nucleosomes in vivo and interacts with neighboring nucleosomes via H3K14ac. Additionally, we establish that RSC is constitutively bound to the nucleosome and is not ejected during mitotic chromatin compaction but alters its binding mode as it progresses through the cell cycle. Our data offer a renewed perspective on RSC mechanics under true physiological conditions.

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Asciminib Mitigates DNA Damage Stress Signaling Induced by Cyclophosphamide in the Ovary

International Journal of Molecular Sciences ◽

10.3390/ijms22031395 ◽

2021 ◽

Vol 22 (3) ◽

pp. 1395

Author(s):

Luca Mattiello ◽

Giulia Pucci ◽

Francesco Marchetti ◽

Marc Diederich ◽

Stefania Gonfloni

Keyword(s):

Ovarian Reserve ◽

Kinase Inhibitor ◽

Concomitant Administration ◽

Cancer Therapies ◽

Cancer Treatments ◽

Abl Tyrosine Kinase ◽

Negative Impacts ◽

Abl Tyrosine Kinase Inhibitor

Cancer treatments can often adversely affect the quality of life of young women. One of the most relevant negative impacts is the loss of fertility. Cyclophosphamide is one of the most detrimental chemotherapeutic drugs for the ovary. Cyclophosphamide may induce the destruction of dormant follicles while promoting follicle activation and growth. Herein, we demonstrate the in vivo protective effect of the allosteric Bcr-Abl tyrosine kinase inhibitor Asciminib on signaling pathways activated by cyclophosphamide in mouse ovaries. We also provide evidence that Asciminib does not interfere with the cytotoxic effect of cyclophosphamide in Michigan Cancer Foundation (MCF)7 breast cancer cells. Our data indicate that concomitant administration of Asciminib mitigates the cyclophosphamide-induced ovarian reserve loss without affecting the anticancer potential of cyclophosphamide. Taken together, these observations are relevant for the development of effective ferto-protective adjuvants to preserve the ovarian reserve from the damaging effects of cancer therapies.

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Targeting Post-Translational Modifications of the p73 Protein: A Promising Therapeutic Strategy for Tumors

Cancers ◽

10.3390/cancers13081916 ◽

2021 ◽

Vol 13 (8) ◽

pp. 1916

Author(s):

Ziad Omran ◽

Mahmood H. Dalhat ◽

Omeima Abdullah ◽

Mohammed Kaleem ◽

Salman Hosawi ◽

...

Keyword(s):

Current Knowledge ◽

Protein A ◽

P53 Gene ◽

Cancer Therapies ◽

Transactivation Domain ◽

P53 Family ◽

Post Translational Modifications ◽

P53 Status ◽

Apoptotic Effects ◽

High Degree

The tumor suppressor p73 is a member of the p53 family and is expressed as different isoforms with opposing properties. The TAp73 isoforms act as tumor suppressors and have pro-apoptotic effects, whereas the ΔNp73 isoforms lack the N-terminus transactivation domain and behave as oncogenes. The TAp73 protein has a high degree of similarity with both p53 function and structure, and it induces the regulation of various genes involved in the cell cycle and apoptosis. Unlike those of the p53 gene, the mutations in the p73 gene are very rare in tumors. Cancer cells have developed several mechanisms to inhibit the activity and/or expression of p73, from the hypermethylation of its promoter to the modulation of the ratio between its pro- and anti-apoptotic isoforms. The p73 protein is also decorated by a panel of post-translational modifications, including phosphorylation, acetylation, ubiquitin proteasomal pathway modifications, and small ubiquitin-related modifier (SUMO)ylation, that regulate its transcriptional activity, subcellular localization, and stability. These modifications orchestrate the multiple anti-proliferative and pro-apoptotic functions of TAp73, thereby offering multiple promising candidates for targeted anti-cancer therapies. In this review, we summarize the current knowledge of the different pathways implicated in the regulation of TAp73 at the post-translational level. This review also highlights the growing importance of targeting the post-translational modifications of TAp73 as a promising antitumor strategy, regardless of p53 status.

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