Epigenetic loss of m1A RNA demethylase ALKBH3 in Hodgkin lymphoma targets collagen, conferring poor clinical outcome.
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ABSTRACT:
SUBMITTER: Esteve-Puig R
PROVIDER: S-EPMC7918181 | biostudies-literature | 2021 Feb
REPOSITORIES: biostudies-literature
Publications
Epigenetic loss of m1A RNA demethylase ALKBH3 in Hodgkin lymphoma targets collagen, conferring poor clinical outcome.
Esteve-Puig Rosaura R Climent Fina F Piñeyro David D Domingo-Domènech Eva E Davalos Veronica V Encuentra Maite M Rea Anna A Espejo-Herrera Nadia N Soler Marta M Lopez Miguel M Ortiz-Barahona Vanessa V Tapia Gustavo G Navarro José-Tomás JT Cid Joan J Farré Lourdes L Villanueva Alberto A Casanova Isolda I Mangues Ramon R Santamarina-Ojeda Pablo P Fernández Agustín F AF Fraga Mario F MF Piris Miguel Angel MA Kol Nitzan N Avrahami Chen C Moshitch-Moshkovitz Sharon S Rechavi Gideon G Sureda Anna A Esteller Manel M
Blood 20210201 7
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Project description:BackgroundEpigenetic inactivation of tumor suppressor genes (TSG) by promoter CpG island hypermethylation is a hallmark of cancer. To assay its extent in human lymphoma, methylation of 24 TSG was analyzed in lymphoma-derived cell lines as well as in patient samples.MethodsWe screened for TSG methylation using methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) in 40 lymphoma-derived cell lines representing anaplastic large cell lymphoma, Burkitt lymphoma (BL), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), Hodgkin lymphoma and mantle cell lymphoma (MCL) as well as in 50 primary lymphoma samples. The methylation status of differentially methylated CD44 was verified by methylation-specific PCR and bisulfite sequencing. Gene expression of CD44 and its reactivation by DNA demethylation was determined by quantitative real-time PCR and on the protein level by flow cytometry. Induction of apoptosis by anti-CD44 antibody was analyzed by annexin-V/PI staining and flow cytometry.ResultsOn average 8 ± 2.8 of 24 TSG were methylated per lymphoma cell line and 2.4 ± 2 of 24 TSG in primary lymphomas, whereas 0/24 TSG were methylated in tonsils and blood mononuclear cells from healthy donors. Notably, we identified that CD44 was hypermethylated and transcriptionally silenced in all BL and most FL and DLBCL cell lines, but was usually unmethylated and expressed in MCL cell lines. Concordant results were obtained from primary lymphoma material: CD44 was not methylated in MCL patients (0/11) whereas CD44 was frequently hypermethylated in BL patients (18/29). In cell lines with CD44 hypermethylation, expression was re-inducible at mRNA and protein levels by treatment with the DNA demethylating agent 5-Aza-2'-deoxycytidine, confirming epigenetic regulation of CD44. CD44 ligation assays with a monoclonal anti-CD44 antibody showed that CD44 can mediate apoptosis in CD44+ lymphoma cells. CD44 hypermethylated, CD44- lymphoma cell lines were consistently resistant towards anti-CD44 induced apoptosis.ConclusionOur data show that CD44 is epigenetically regulated in lymphoma and undergoes de novo methylation in distinct lymphoma subtypes like BL. Thus CD44 may be a promising new epigenetic marker for diagnosis and a potential therapeutic target for the treatment of specific lymphoma subtypes.
| S-EPMC2955612 | biostudies-literature
Project description:BackgroundThe oxidative DNA demethylase ALKBH3 targets single-stranded DNA (ssDNA) in order to perform DNA alkylation damage repair. ALKBH3 becomes upregulated during tumorigenesis and is necessary for proliferation. However, the underlying molecular mechanism remains to be understood.MethodsTo further elucidate the function of ALKBH3 in cancer, we performed ChIP-seq to investigate the genomic binding pattern of endogenous ALKBH3 in PC3 prostate cancer cells coupled with microarray experiments to examine the expression effects of ALKBH3 depletion.ResultsWe demonstrate that ALKBH3 binds to transcription associated locations, such as places of promoter-proximal paused RNA polymerase II and enhancers. Strikingly, ALKBH3 strongly binds to the transcription initiation sites of a small number of highly active gene promoters. These promoters are characterized by high levels of transcriptional regulators, including transcription factors, the Mediator complex, cohesin, histone modifiers, and active histone marks. Gene expression analysis showed that ALKBH3 does not directly influence the transcription of its target genes, but its depletion induces an upregulation of ALKBH3 non-bound inflammatory genes.ConclusionsThe genomic binding pattern of ALKBH3 revealed a putative novel hyperactive promoter type. Further, we propose that ALKBH3 is an intrinsic DNA repair protein that suppresses transcription associated DNA damage at highly expressed genes and thereby plays a role to maintain genomic integrity in ALKBH3-overexpressing cancer cells. These results raise the possibility that ALKBH3 may be a potential target for inhibiting cancer progression.
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Project description:Hodgkin lymphoma (HL), a B cell-derived cancer, is one of the most common lymphomas. In HL, the tumor cells--Hodgkin and Reed-Sternberg (HRS) cells--are usually very rare in the tissue. Although HRS cells are derived from mature B cells, they have largely lost their B cell phenotype and show a very unusual co-expression of markers of various hematopoietic cell types. HRS cells show deregulated activation of multiple signaling pathways and transcription factors. The activation of these pathways and factors is partly mediated through interactions of HRS cells with various other types of cells in the microenvironment, but also through genetic lesions. The transforming events involved in the pathogenesis of HL are only partly understood, but mutations affecting the NF-?B and JAK/STAT pathways are frequent. The dependency of HRS cells on microenvironmental interactions and deregulated signaling pathways may offer novel strategies for targeted therapies.
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Project description:Albeit N1-Methyladenosine (m1A) RNA modification represents an important regulator of RNA metabolism, the role of m1A modification in carcinogenesis remains enigmatic. Herein, we found that histone lactylation enhances ALKBH3 expression and simultaneously attenuates the formation of tumor-suppressive promyelocytic leukemia protein (PML) condensates by removing the m1A methylation of SP100A, promoting the malignant transformation of cancers. First, ALKBH3 is specifically upregulated in high-risk ocular melanoma due to excessive histone lactylation levels, referring to m1A hypomethylation status. Moreover, the multiomics analysis subsequently identified that SP100A, a core component for PML bodies, serves as a downstream candidate target for ALKBH3. Therapeutically, the silencing of ALKBH3 exhibits efficient therapeutic efficacy in melanoma both in vitro and in vivo, which could be reversed by the depletion of SP100A. Mechanistically, we found that YTHDF1 is responsible for recognition of the m1A methylated SP100A transcript, which increases its RNA stability and translational efficacy. Conclusively, we initially demonstrated that m1A modification is necessary for tumor suppressor gene expression, expanding the current understandings of dynamic m1A function during tumor progression. In addition, our results indicate that lactylation-driven ALKBH3 is essential for the formation of PML nuclear condensates, which bridges our knowledge of m1A modification, metabolic reprogramming, and phase-separation events.
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Project description:Transfer RNA is heavily modified and plays a central role in protein synthesis and cellular functions. Here we demonstrate that ALKBH3 is a 1-methyladenosine (m1A) and 3-methylcytidine (m3C) demethylase of tRNA. ALKBH3 can promote cancer cell proliferation, migration and invasion. In vivo study confirms the regulation effects of ALKBH3 on growth of tumor xenograft. The m1A demethylated tRNA is more sensitive to angiogenin (ANG) cleavage, followed by generating tRNA-derived small RNAs (tDRs) around the anticodon regions. tDRs are conserved among species, which strengthen the ribosome assembly and prevent apoptosis triggered by cytochrome c (Cyt c). Our discovery opens a potential and novel paradigm of tRNA demethylase, which regulates biological functions via generation of tDRs.
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Project description:Human AlkB homolog 3 (ALKBH3), a homolog of the Escherichia coli protein AlkB, demethylates 1-methyladenine and 3-methylcytosine (3-meC) in single-stranded DNA and RNA by oxidative demethylation. Immunohistochemical analyses on clinical cancer specimens and knockdown experiments using RNA interference in vitro and in vivo indicate that ALKBH3 is a promising molecular target for the treatment of prostate, pancreatic, and non-small cell lung cancer. Therefore, an inhibitor for ALKBH3 demethylase is expected to be a first-in-class molecular-targeted drug for cancer treatment. Here, we report the development of a novel, quantitative real-time PCR-based assay for ALKBH3 demethylase activity against 3-meC by highly active recombinant ALKBH3 protein using a silkworm expression system. This assay enables us to screen for inhibitors of ALKBH3 demethylase, which may result in the development of a novel molecular-targeted drug for cancer therapy.
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Project description:Comparative proteome analysis of HL and DLBCL educated human macrophages by SWATH-MS.
2020-01-29 | PXD016128 | Pride
Project description:The integrity of our DNA is challenged daily by a variety of chemicals that cause DNA base alkylation. DNA alkylation repair is an essential cellular defence mechanism to prevent the cytotoxicity or mutagenesis from DNA alkylating chemicals. Human oxidative demethylase ALKBH3 is a central component of alkylation repair, especially from single-stranded DNA. However, the molecular mechanism of ALKBH3-mediated damage recognition and repair is less understood. We report that ALKBH3 has a direct protein-protein interaction with human RAD51 paralogue RAD51C. We also provide evidence that RAD51C-ALKBH3 interaction stimulates ALKBH3-mediated repair of methyl-adduct located within 3'-tailed DNA, which serves as a substrate for the RAD51 recombinase. We further show that the lack of RAD51C-ALKBH3 interaction affects ALKBH3 function in vitro and in vivo. Our data provide a molecular mechanism underlying upstream events of alkyl adduct recognition and repair by ALKBH3.
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Project description:In the last 10 years, major advances have been made in the diagnosis and development of selective therapies for several blood cancers, including B-cell non-Hodgkin lymphoma (B-NHL), a heterogeneous group of malignancies arising from the mature B lymphocyte compartment. However, most of these entities remain incurable and current treatments are associated with variable efficacy, several adverse events, and frequent relapses. Thus, new diagnostic paradigms and novel therapeutic options are required to improve the prognosis of patients with B-NHL. With the recent deciphering of the mutational landscapes of B-cell disorders by high-throughput sequencing, it came out that different epigenetic deregulations might drive and/or promote B lymphomagenesis. Consistently, over the last decade, numerous epigenetic drugs (or epidrugs) have emerged in the clinical management of B-NHL patients. In this review, we will present an overview of the most relevant epidrugs tested and/or used so far for the treatment of different subtypes of B-NHL, from first-generation epigenetic therapies like histone acetyl transferases (HDACs) or DNA-methyl transferases (DNMTs) inhibitors to new agents showing selectivity for proteins that are mutated, translocated, and/or overexpressed in these diseases, including EZH2, BET, and PRMT. We will dissect the mechanisms of action of these epigenetic inhibitors, as well as the molecular processes underlying their lack of efficacy in refractory patients. This review will also provide a summary of the latest strategies being employed in preclinical and clinical settings, and will point out the most promising lines of investigation in the field.
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