Functional epigenetic approach identifies frequently methylated genes in Ewing sarcoma

Alholle, Abdullah; Brini, A.T.; Gharanei, Seley; Vaiyapuri, Sumathi; Arrigoni, E.; Dallol, Ashraf; Gentle, Dean; Kishida, Takeshi; Hiruma, Toru; Avigad, Smadar; Grimer, R. J.; Latif, Farida

doi:10.4161/epi.26266

Cited by 39 publications

(30 citation statements)

References 23 publications

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“…34 We further show that DNA methylation of neurofilament genes is not specific to breast cancers but occurs in other types of cancers as well, i.e., in pancreas, colon, and gastric cancers. These results and the recent findings that showed NEFH inactivation in association with promoter methylation in esophageal squamous cell and hepatocellular carcinomas as well as in Ewing sarcomas 19,21,25 suggest that neurofilament genes may be epigenetically downregulated in a wide range of malignancies and that these gene family may have importance in tumorigenesis. Using proliferation and colony formation assays, we show that NEFH possesses tumor cell growth suppressive activity in breast cancer cells.…”

Section: Discussionsupporting

confidence: 65%

“…[22][23][24] Furthermore, aberrant expression and methylation of neurofilament genes were recently detected in esophageal squamous cell, renal cell and hepatocellular carcinoma as well as in Ewing sarcoma. 19,21,25,26 In the present study we used a comprehensive approach to study the epigenetic inactivation of neurofilament genes in breast cancer and the functional significance of these events.…”

Section: Introductionmentioning

confidence: 99%

“…14,15 Neurofilaments have been implicated in the pathology of neurological diseases, [16][17][18] but recent data also point to a role in the pathology of cancer. [19][20][21] For example, NEFL and NEFM are both located within the 8p21 chromosome region and LOH of this region has been described in several cancers including breast cancer. [22][23][24] Furthermore, aberrant expression and methylation of neurofilament genes were recently detected in esophageal squamous cell, renal cell and hepatocellular carcinoma as well as in Ewing sarcoma.…”

Section: Introductionmentioning

confidence: 99%

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Epigenetic silencing of neurofilament genes promotes an aggressive phenotype in breast cancer

et al. 2015

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Neurofilament heavy polypeptide (NEFH) has recently been identified as a candidate DNA hypermethylated gene within the functional breast cancer hypermethylome. NEFH exists in a complex with neurofilament medium polypeptide (NEFM) and neurofilament light polypeptide (NEFL) to form neurofilaments, which are structural components of the cytoskeleton in mature neurons. Recent studies reported the deregulation of these proteins in several malignancies, suggesting that neurofilaments may have a role in other cell types as well. Using a comprehensive approach, we studied the epigenetic inactivation of neurofilament genes in breast cancer and the functional significance of this event. We report that DNA methylation-associated silencing of NEFH, NEFL, and NEFM in breast cancer is frequent, cancer-specific, and correlates with clinical features of disease progression. DNA methylationmediated inactivation of these genes occurs also in multiple other cancer histologies including pancreas, gastric, and colon. Restoration of NEFH function, the major subunit of the neurofilament complex, reduces proliferation and growth of breast cancer cells and arrests them in Go/G1 phase of the cell cycle along with a reduction in migration and invasion. These findings suggest that DNA methylation-mediated silencing of the neurofilament genes NEFH, NEFM, and NEFL are frequent events that may contribute to the progression of breast cancer and possibly other malignancies.

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Section: Discussionsupporting

confidence: 65%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Epigenetic silencing of neurofilament genes promotes an aggressive phenotype in breast cancer

et al. 2015

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“…MUG-Chor1 was maintained in IMDM/RPMI (4:1) with 2 mM L-Glutamine, 1% ITS and 1% PenStrep at 37 C, 5% CO 2 . The cell lines were treated with 5-aza-2 0 -deoxycitidine (5-azaDC) over 5 d as described in Alholle et al 27 RT-PCR, bisulfite modification and COBRA Superscript III cDNA synthesis kit (Invitrogen) was used to create cDNA from 1 mg RNA. RT-PCR was performed as described previously.…”

Section: Chordoma Cell Linesmentioning

confidence: 99%

Genome-wide DNA methylation profiling of recurrent and non-recurrent chordomas

et al. 2015

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Chordomas are an aggressive rare type of malignant bone tumors arising from the remnant of the notochord. Chordomas occur mainly in vertebral bones and account for 1-4% of malignant bone tumors. Management and treatment of chordomas are difficult as they are resistant to conventional chemotherapy; therefore, they are mainly treated with surgery and radiation therapy. In this study, we performed DNA methylation profiling of 26 chordomas and normal nucleus pulposus samples plus UCH-1 chordoma cell line using the Illumina Infinium HumanMethylation450 BeadChips. Combined bisulfite restriction analysis and bisulfite sequencing was used to confirm the methylation data. Gene expression was analyzed using RT-PCR before and after 5-aza-2'-deoxycytidine (5-azaDC) treatment of chordoma cell lines. Analysis of the HumanMethylation450 BeadChip data led to the identification of 8,819 loci (2.9%) that were significantly differentially methylated (>0.2 average b-value difference) between chordomas and nucleus pulposus samples (adjusted P < 0.05). Among these, 5,868 probes (66.5%) were hypomethylated, compared to 2,951 (33.5%) loci that were hypermethylated in chordomas compared to controls. From the 2,951 differentially hypermethylated probes, 33.3% were localized in the promoter region (982 probes) and, among these, 104 probes showed cancer-specific hypermethylation. Ingenuity Pathway Analysis indicates that the cancer-specific differentially methylated loci are involved in various networks including cancer disease, nervous system development and function, cell death and survival, cellular growth, cellular development, and proliferation. Furthermore, we identified a subset of probes that were differentially methylated between recurrent and non-recurrent chordomas. BeadChip methylation data was confirmed for these genes and gene expression was shown to be upregulated in methylated chordoma cell lines after treatment with 5-azaDC. Understanding epigenetic changes in chordomas may provide insights into chordoma tumorigenesis and development of epigenetic biomarkers.

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“…Here we used dysfunctional telomeres to identify PHF11 (plant homeodomain finger 11) as a DDR factor. PHF11 is frequently methylated in Ewing's sarcoma (Alholle et al 2013), codeleted with a cluster of genes in CLL (Parker et al 2011), and deleted in 10%-20% of prostate cancers (cBioPortal for Cancer Genomics, http://www.cbioportal .org). PHF11 is most abundant in lymphocytes and is thought to act as a transcription factor that promotes class switching in the IgE locus in B cells (Clarke et al 2008;Ikari et al 2014).…”

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confidence: 99%

PHF11 promotes DSB resection, ATR signaling, and HR

et al. 2017

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Resection of double-strand breaks (DSBs) plays a critical role in their detection and appropriate repair. The 3 ′ ssDNA protrusion formed through resection activates the ATR-dependent DNA damage response (DDR) and is required for DSB repair by homologous recombination (HR). Here we report that PHF11 (plant homeodomain finger 11) encodes a previously unknown DDR factor involved in 5 ′ end resection, ATR signaling, and HR. PHF11 was identified based on its association with deprotected telomeres and localized to sites of DNA damage in S phase. Depletion of PHF11 diminished the ATR signaling response to telomere dysfunction and genome-wide DNA damage, reduced end resection at sites of DNA damage, resulted in compromised HR and misrejoining of S-phase DSBs, and increased the sensitivity to DNA-damaging agents. PHF11 interacted with the ssDNA-binding protein RPA and was found in a complex with several nucleases, including the 5 ′ dsDNA exonuclease EXO1. Biochemical experiments demonstrated that PHF11 stimulates EXO1 by overcoming its inhibition by RPA, suggesting that PHF11 acts (in part) by promoting 5 ′ end resection at RPA-bound sites of DNA damage. These findings reveal a role for PHF11 in DSB resection, DNA damage signaling, and DSB repair.[Keywords: PHF11; EXO1; RPA; ATR; DSB; resection; homologous recombination] Supplemental material is available for this article.Received October 9, 2016; revised version accepted December 22, 2016.Nuclear double-strand breaks (DSBs) activate the ATM and ATR kinase-dependent DNA damage response (DDR) pathways (for review, see Ciccia and Elledge 2010). Whereas the ATM kinase responds to the presence of dsDNA ends, activation of the ATR kinase requires the presence of ssDNA that is bound by RPA. In addition, activation of the ATR kinase requires the loading of the 9-1-1 complex on the double-stranded-single-stranded junction formed after 5 ′ end resection. Resection of the 5 ′ end of DSBs is therefore a critical step in the activation of ATR signaling. DSB resection is also required for the initiation of several DNA repair pathways, including homologous recombination (HR) and single-strand annealing (SSA) (for review, see Symington and Gautier 2011). HR can restore the original DNA sequence using the sister chromatid as a template for repair. After 5 ′ end resection, HR is initiated by the BRCA2-mediated loading of the Rad51 recombinase onto the ssDNA. In the absence of HR, S-phase DSBs can become a substrate for more error-prone DSB repair, including SSA and classical or alternative nonhomologous end-joining (NHEJ). DSB resection is initiated through the agency of BRCA1, the MRE11/RAD50/NBS1 (MRN) complex, and CtIP (for review, see Cejka 2015). The outcome is a ssDNA end with a short (∼20-nucleotide [nt]) 3 ′ overhang that is a substrate for further nucleolytic attack by EXO1, a dsDNA exonuclease that degrades only the 5 ′ strand to leave a 3 ′ overhang that can be thousands of nucleotides in length. In addition, an overlapping pathway involving the DNA2 nuclease acting in conjun...

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Functional epigenetic approach identifies frequently methylated genes in Ewing sarcoma

Cited by 39 publications

References 23 publications

Epigenetic silencing of neurofilament genes promotes an aggressive phenotype in breast cancer

Epigenetic silencing of neurofilament genes promotes an aggressive phenotype in breast cancer

Genome-wide DNA methylation profiling of recurrent and non-recurrent chordomas

PHF11 promotes DSB resection, ATR signaling, and HR

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