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| Map of TERT and CLPTM1L region. Recombination hotspots in the CEU population (red line), as well as 1000G combined recombination rate (blue line) across the TERT/CLPTM1L region are shown relative to the CLPTM1L and TERT genes, as well as the grouping of ten highly correlated sequence variants strongly associated with risk of pancreatic, testicular, and lung cancers in the region closest to CLPTM1L. (a) Chromatin interaction analysis paired-end (ChIA-PET) sequencing data from the K562 chronic myeloid leukaemia cell line using an antibody against RNA polymerase II generated by the ENCODE project (https://www.encodeproject.org/) is shown. For each of the ten strongly associated variants, layered H3K4Me1, H3K4Me3, and H3K27Ac chromatin immunoprecipiation (ChIP-seq), DNAse I hypersensitivity sequencing (DNase) and transcription factor ChIP-seq (TF-ChIP-Seq) data from the ENCODE project are shown (b) as displayed by the UCSC Genome Browser (lower panels).
Source publication
Genome wide association studies (GWAS) have mapped multiple independent cancer susceptibility loci to chr5p15.33. Here, we show that fine-mapping of pancreatic and testicular cancer GWAS within one of these loci (Region 2 in CLPTM1L) focuses the signal to nine highly correlated SNPs. Of these, rs36115365-C associated with increased pancreatic and t...
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... lower than the previously published association signal marked by rs401681, with rs451360 being the smallest (P ¼ 2.0 Â 10 À 10 for rs451360; P ¼ 3.7 Â 10 À 7 for rs401681; Supplementary Table 1) 18 . This SNP is highly correlated with eight other SNPs (r 2 40.60, 1000G EUR population) that collectively mark Region 2 in pancreatic cancer (Fig. 1). Fine- mapping of Region 2 for testicular germ cell tumours (TGCT) and lung cancer revealed that the strongest SNP for each was among this group of nine SNPs (rs35953391 for TGCT, P ¼ 1.08 Â 10 À 9 ; and rs37004 for lung cancer, P ¼ 1.18 Â 10 À 13 ; Supplementary Table 1). Conditional analysis for the most significant SNP across each ...
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... to that of the other nine variants (rs3030832, P ¼ 8.25 Â 10 À 10 , OR¼ 1.28 95% CI 1.18-1.39; Supplementary Table 2) indicating that this indel variant should likewise be considered a candidate functional risk variant. Overall, these ten variants extend across the entire length of CLPTM1L, from the promoter to B6 kb downstream of the gene (Fig. 1). Three variants, rs36115365, rs380145 and rs27071, are located within potential gene regulatory regions, annotated by the ENCODE project ( Fig. 1, Supplementary Fig. ...
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... variant should likewise be considered a candidate functional risk variant. Overall, these ten variants extend across the entire length of CLPTM1L, from the promoter to B6 kb downstream of the gene (Fig. 1). Three variants, rs36115365, rs380145 and rs27071, are located within potential gene regulatory regions, annotated by the ENCODE project ( Fig. 1, Supplementary Fig. ...
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... SNP is located in-between the 5 0 end of TERT (B18 kb upstream) and 3 0 end of CLPTM1L (B5 kb downstream), a region that overlaps active histone modification marks and multiple transcription factor binding sites according to ENCODE data ( Fig. 1, Supplementary Fig. 1). The region harbouring rs36115365 demonstrated an allele-specific increase in luciferase reporter activity as compared to empty vector that was consistent across all eight cancer cell lines tested (Fig. 3, Supplementary Fig. 4), including those from pancreas (PANC-1 and MIA PaCa-2, average fold change for C versus G allele 1.38, range ...
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... fourth protein, ZNF740, was also found to preferentially bind the C variant in both cell lines using mixed poly-dAdT and poly-dIdC competitors ( Supplementary Fig. 9, bottom panels). We sought to verify whether any of these four proteins differentially bound the C-allele by using antibodies against these proteins in conjunction with EMSAs for rs36115365 (Fig. 5b, Supplementary Figs 10 and 11). Only the antibody against ZNF148 consistently resulted in loss of C allele-specific banding in pancreatic (PANC-1; Fig. 5b), as well as testis (NTERA-2) and lung cancer (A549) lines ( Supplementary Fig. 10). ...
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... sought to verify whether any of these four proteins differentially bound the C-allele by using antibodies against these proteins in conjunction with EMSAs for rs36115365 (Fig. 5b, Supplementary Figs 10 and 11). Only the antibody against ZNF148 consistently resulted in loss of C allele-specific banding in pancreatic (PANC-1; Fig. 5b), as well as testis (NTERA-2) and lung cancer (A549) lines ( Supplementary Fig. 10). Furthermore, EMSAs using recombinant purified ZNF148 protein demonstrated specific binding of ZNF148 to the C allele of rs36115365 (Fig. 5c). ...
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... of the kruppel-like family 35 , is predicted to bind to a consensus DNA-recognition motif created by the C-allele of rs36115365 ( Fig. 5d). To further establish the binding of ZNF148 to rs36115365 and surrounding genomic region, we performed chromatin-immunoprecipitation (ChIP) for ZNF148 followed by quantitative PCR, noting an enrichment of binding at rs36115365 in pancreatic and lung cancer cell lines homozygous and heterozygous for rs36115365-C as compared to background and the surrounding area ( Fig. 5e, Supplementary Fig. 12a-f). We also assessed allelic enrichment in the immunoprecipitates and noted a significant enrichment of the C allele as compared to the G allele in A549 cells (1.51 fold, P ¼ 0.01; t-test; Supplementary Fig. 12g), with Panc 05.04 cells showing a nonsignificant trend in the same direction (1.12 fold, P ¼ 0.06; t-test; Supplementary Fig. 12g). ...
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... further establish the binding of ZNF148 to rs36115365 and surrounding genomic region, we performed chromatin-immunoprecipitation (ChIP) for ZNF148 followed by quantitative PCR, noting an enrichment of binding at rs36115365 in pancreatic and lung cancer cell lines homozygous and heterozygous for rs36115365-C as compared to background and the surrounding area ( Fig. 5e, Supplementary Fig. 12a-f). We also assessed allelic enrichment in the immunoprecipitates and noted a significant enrichment of the C allele as compared to the G allele in A549 cells (1.51 fold, P ¼ 0.01; t-test; Supplementary Fig. 12g), with Panc 05.04 cells showing a nonsignificant trend in the same direction (1.12 fold, P ¼ 0.06; t-test; Supplementary Fig. 12g). ...
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... further establish the binding of ZNF148 to rs36115365 and surrounding genomic region, we performed chromatin-immunoprecipitation (ChIP) for ZNF148 followed by quantitative PCR, noting an enrichment of binding at rs36115365 in pancreatic and lung cancer cell lines homozygous and heterozygous for rs36115365-C as compared to background and the surrounding area ( Fig. 5e, Supplementary Fig. 12a-f). We also assessed allelic enrichment in the immunoprecipitates and noted a significant enrichment of the C allele as compared to the G allele in A549 cells (1.51 fold, P ¼ 0.01; t-test; Supplementary Fig. 12g), with Panc 05.04 cells showing a nonsignificant trend in the same direction (1.12 fold, P ¼ 0.06; t-test; Supplementary Fig. 12g). ...
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... ¼ 2.0 Â 10 À 4 -0.012; t-test; Fig. 6a, Supplementary Figs 13 and 14), consistent with a role for ZNF148 in regulating TERT expression. In contrast, siRNA-mediated knockdown of VEZF1 (ZNF161), ZNF281 and ZNF740 showed no effect on expression of either TERT or CLPTM1L (Supplementary Fig. 15). ...
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... Fig. 6a, Supplementary Figs 13 and 14), consistent with a role for ZNF148 in regulating TERT expression. In contrast, siRNA-mediated knockdown of VEZF1 (ZNF161), ZNF281 and ZNF740 showed no effect on expression of either TERT or CLPTM1L (Supplementary Fig. 15). We next sought to assess if ZNF148-mediated regulation of TERT expression was accompanied by effects on telomerase activity and telomere length. ...
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... next sought to assess if ZNF148-mediated regulation of TERT expression was accompanied by effects on telomerase activity and telomere length. Knockdown of ZNF148 via PTGS resulted in reduced telomerase activity in A549 and MIA PaCa-2 cells (Fig. 6b), as well as in NTERA-2 and UACC903 cells (Supplementary Fig. 16). This reduction was similar to that observed via siRNA-mediated depletion of TERT itself, or by transcriptional gene silencing (TGS, siRNA3) to target the gene regulatory element encompassing rs36115365. ...
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... further assess the role of ZNF148 in regulating TERT expression and activity, we performed rescue experiments after depletion of endogenous ZNF148 using an siRNA targeting the 3 0 -UTR of ZNF148. Overexpression of exogenous ZNF148 lacking the 3 0 -UTR indeed rescued both TERT expression and telomerase activity in A549 and MIA PaCa-2 cells (Supplementary Fig. 17). Consistent with these data, depletion of either ZNF148 or TERT, or alternatively targeting the rs36115365 regulatory region in both A549 and MIA PaCa-2 cells all resulted in similar reductions of telomere length (Fig. 6c). ...
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... TERT. Here, we identify rs36115365 as a functional SNP in this region and provide a plausible biological explanation underlying risk, featuring altered TERT, but not CLPTM1L, expression. Fine-mapping of Region 2 using GWAS data from pancreatic, lung and testicular cancer confirmed significant association with this small set of tightly linked SNPs (Fig. 1). Little signal remained within Region 2 after ...
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... pulled-down DNA was assayed by nine SYBR Green qPCR amplicons for enrichment of target sites using primers listed in Supplementary Table 4. A TaqMan genotyping assay for rs36115365 (C_470504_10, Life Technologies) was used to quantify the C and G alleles in immunoprecipitated DNA samples in seven independent experiments ( Supplementary Fig. 12g). A paired two sided T-test was applied to C-and G-allele signals (normalized to input DNA) in order to assess significance of enrichment of the C versus G allele at rs36115365. ...
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... specificity of the ZNF148 antibody was tested by western blot analysis with and without siRNA mediated knockdown of ZNF148. GAPDH (ab37168, 1 mg ml À 1 , Abcam) was used as a loading control ( Supplementary Fig. 18). ...
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... assess possible off-target effects for the ZNF148 siRNAs we also purchased each of the four siRNAs from the SMARTpool separately and tested their effects on ZNF148 and TERT expression. All four siRNAs inhibited both ZNF148 and TERT expression indicating that off-target effects are not likely to explain our findings ( Supplementary Fig. 14). Transfection, RNA purification, cDNA generation and expression analysis procedures were as described above for the region-targeted siRNA assay, except that RNA was isolated 72 h after transfection. ...
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... Among these proteins, we identified 8 multifunctional transcription factors, including YY1, GATA2, MAZ, NFIC, TCF7L2, ELF2, ZNF281, and ZNF148 (Fig. 4G, 4H). These 8 proteins can exhibit both positive and negative control over a substantial number of cellular genes by binding to their respective transcription start sites [32][33][34][35][36][37][38][39][40][41][42][43][44][45][46][47] . ...
Poly-ADP-ribosylation (PARylation) is a reversible posttranslational modification that occurs in higher eukaryotes. While thousands of PARylated substrates have been identified, the specific biological functions of most PARylated proteins remain elusive. PARylation stoichiometry is a critical parameter to assess the potential functions of a PARylated protein. Here, we developed a large-scale strategy to measure the absolute stoichiometries of protein PARylation. By integrating mild cell lysis, boronate enrichment and carefully designed titration experiments, we were able to determine the PARylation stoichiometries for a total of 235 proteins. This approach enables the capture of all PARylation events on various amino acid acceptors. We revealed that PARylation occupancy spans over three orders of magnitude. However, most PARylation events occur at low stoichiometric values (median 0.578%). Notably, we observed that high stoichiometry PARylation (>1%) predominantly targets proteins involved in transcription regulation and chromatin remodeling. Thus, our study provides a systems-scale, quantitative view of PARylation stoichiometries under genotoxic conditions, which serves as invaluable resources for future functional studies of this important protein posttranslational modification.
... Despite the absence of significant allelic effects after multiple testing corrections, rs36115365 exhibited nominal significance in the combined data of three lung-related cell lines (MPRA: log 2 FoldChange = 0.12, P nominal = 0.03). The block was recognized for its association with TL 46-48 and its influence on the onset of various tumors 9,49,50 . The effect of block 4, however, was found to be independent of TL, with the underlying genetic mechanisms remaining elusive. ...
Genome-wide association studies have identified thousands of genetic variants associated with non-small cell lung cancer (NSCLC), however, it is still challenging to determine the causal variants and to improve disease risk prediction. Here, we applied massively parallel reporter assays to perform NSCLC variant-to-function mapping at scale. A total of 1249 candidate variants were evaluated, and 30 potential causal variants within 12 loci were identified. Accordingly, we proposed three genetic architectures underlying NSCLC susceptibility: multiple causal variants in a single haplotype block (e.g. 4q22.1), multiple causal variants in multiple haplotype blocks (e.g. 5p15.33), and a single causal variant (e.g. 20q11.23). We developed a modified polygenic risk score using the potential causal variants from Chinese populations, improving the performance of risk prediction in 450,821 Europeans from the UK Biobank. Our findings not only augment the understanding of the genetic architecture underlying NSCLC susceptibility but also provide strategy to advance NSCLC risk stratification.
... Several reports have identified NPPA as an independent prognostic indicator for breast cancer (17,18). Furthermore, ZNF740 has been shown to potentially engage in the regulation of the cell cycle, cellular adhesion and tumor proliferation (19)(20)(21)(22), demonstrating a significant overlap of >20% with TP53 target genes (23). Nevertheless, the precise roles and molecular mechanisms of ZNF740 in HCC remain unclear. ...
Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related death, and efficient treatments to facilitate recovery and enhance long-term outcomes are lacking. Zinc finger proteins (ZNFs), known as the largest group of transcription factors, have gained interest for their roles in HCC by stimulating the transcription of well-known tumor-causing genes. However, the specific roles and molecular mechanisms of ZNF740 in HCC remain unknown. The present study performed bioinformatics analysis and RNA-sequencing analysis of differentially expressed genes in HCC, detected ZNF740 expression levels in HCC using reverse transcription-quantitative PCR, western blotting and immunohistochemistry, and explored the effects of ZNF740 on the progression of liver cancer in vitro and in vivo using cellular functionality assays and cell-derived xenografts. In addition, a dual-luciferase reporter assay was performed to analyze the binding of ZNF740 with the METTL3 promoter. Furthermore, cell functionality experiments were performed to analyze whether ZNF740 promotes the proliferation of liver cancer cells in a METTL3-dependent manner. Bioinformatics and immunoprecipitation assays were further used to analyze the molecular mechanism of ZNF740 in liver cancer. The present study demonstrated that ZNF740 expression was upregulated in HCC. Mechanistically, overexpressed ZNF740 interacted with the methyltransferase-like 3 (METTL3) promoter and increased METTL3 expression, leading to the stabilization of hypoxia-inducible factor-1A (HIF1A) mRNA in an N6-methyladenosine/YTH N6-methyladenosine RNA-binding protein 1-dependent manner. Eventually, the ZNF740/METTL3/HIF1A signaling axis may facilitate the proliferation, invasion and metastasis of liver cancer via METTL3/HIF-1A signaling. The present findings revealed the important role of ZNF740 and suggested a potential therapeutic approach that might improve clinical therapies for liver cancer.
... Overall, these findings suggest that the same biological pathway has opposite effects on the susceptibility to different tumor types. This interpretation is supported by functional characterization of rs36115365, a variant on 5p15.33, which was found to have similar cis-regulatory effects on TERT in multiple cancers cell lines from different cancers, but was associated with a higher risk of pancreatic and testicular cancer and a lower risk of lung cancer [33]. Alternatively, a causal variant may differently influence cis-gene regulation and/or alter different biological pathways depending on the cell or tissue of origin [34]. ...
Background
Genome-wide association studies (GWAS) have identified multiple common breast cancer susceptibility variants. Many of these variants have differential associations by estrogen receptor (ER) status, but how these variants relate with other tumor features and intrinsic molecular subtypes is unclear.
Methods
Among 106,571 invasive breast cancer cases and 95,762 controls of European ancestry with data on 173 breast cancer variants identified in previous GWAS, we used novel two-stage polytomous logistic regression models to evaluate variants in relation to multiple tumor features (ER, progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) and grade) adjusting for each other, and to intrinsic-like subtypes.
Results
Eighty-five of 173 variants were associated with at least one tumor feature (false discovery rate < 5%), most commonly ER and grade, followed by PR and HER2. Models for intrinsic-like subtypes found nearly all of these variants (83 of 85) associated at p < 0.05 with risk for at least one luminal-like subtype, and approximately half (41 of 85) of the variants were associated with risk of at least one non-luminal subtype, including 32 variants associated with triple-negative (TN) disease. Ten variants were associated with risk of all subtypes in different magnitude. Five variants were associated with risk of luminal A-like and TN subtypes in opposite directions.
Conclusion
This report demonstrates a high level of complexity in the etiology heterogeneity of breast cancer susceptibility variants and can inform investigations of subtype-specific risk prediction.
... The 5p15.33 TERT/CLPTM1L locus is an extensively characterized multi-cancer risk locus, [18][19][20][21] and over 10 tumor types are associated with this risk region, including carcinomas from ER-negative breast, colon, lung, pancreas, prostate, kidney, ovary, head and neck, esophagus, and endometrium, as well as germ cell tumor, cutaneous melanoma, and glioma. Importantly, up to ten independent risk loci have now been identified within this genomic region, encompassing both CLPTM1L and TERT. ...
... 48 Telomere maintenance has been extensively linked to cancer, and TERT reactivation is a possible mechanism by which tumor cells could maintain abnormal cell survival and promote cancer development. 49 Several lines of evidence support a role of TERT in rs452384 allele-specific UM risk: (1) the GTEx database indicates a correlation between rs452384 and TERT expression in oesophagus; (2) 5p15.33 risk SNPs (including some within CLPTM1L introns) in lung carcinomas and others are associated with TERT, but not CLPTM1L, expression; 19,50 and (3) TERT SNPs play a significant role in predisposition to many solid tumors, including cutaneous melanoma. 20,21,51 As previously mentioned, TERT mRNA is barely detectable in UM tumors, 17,52 making it challenging to assess for genotype-expression correlations. ...
The TERT/CLPTM1L risk locus on chromosome 5p15.33 is a pleiotropic cancer risk locus in which multiple independent risk alleles have been identified, across well over ten cancer types. We previously conducted a genome-wide association study in uveal melanoma (UM), which uncovered a role for the TERT/CLPTM1L risk locus in this intraocular tumor and identified multiple highly correlated risk alleles. Aiming to unravel the biological mechanisms in UM of this locus, which contains a domain enriched in active chromatin marks and enhancer elements, we demonstrated the allele-specific enhancer activity of this risk region using reporter assays. In UM, we identified the functional variant rs452384, of which the C risk allele is associated with higher gene expression, increased CLPTM1L expression in UM tumors, and a longer telomere length in peripheral blood mononuclear cells. Electrophoretic mobility shift assays and quantitative mass spectrometry identified NKX2.4 as an rs452384-T-specific binding protein, whereas GATA4 preferentially interacted with rs452384-C. Knockdown of NKX2.4 but not GATA4 resulted in increased TERT and CLPTM1L expression. In summary, the UM risk conferred by the 5p locus is at least partly due to rs452384, for which NKX2.4 presents strong differential binding activity and regulates CLPTM1L and TERT expression. Altogether, our work unraveled some of the complex regulatory mechanisms at the 5p15.33 susceptibility region in UM, and this might also shed light on shared mechanisms with other tumor types affected by this susceptibility region.
... To determine TERT promoter methylation patterns associated with active or silent histone marks in these cell lines, ChIP DNA was bisulfite converted using an EZ DNA Methylation-Gold Kit (Zymo Research) and PCR amplified for 45 cycles at 95 • C for 15 s, 63 • C for 20 s, and 68 • C for 40 s using primers spanning the region from 1,295,224 to 1,295,546 of the TERT promoter with 5 -GGAAAGGAAGGGGAGGGGTTGGGAG-3 and TERT allele-specific expression in cell lines was measured in rs2736098 or rs2853690 (ThermoFisher; Waltham, MA, USA) heterozygotes using ddPCR TaqMan SNP assays compatible with cDNA of spliced mRNA. Heterozygous sample gDNA allele ratios were quantified by ddPCR [29]. Allele-specific expression was then calculated as follows: allele1 or 2 gDNA ratio = (allele1 or 2 gDNA droplet counts)/(total gDNA droplet counts) allele1 or 2 cDNA ratio = (allele1 or 2 cDNA droplet counts)/(total cDNA droplet counts) normalized allele1 or 2 cDNA ratio = (allele1 or 2 cDNA ratio)/(allele1 or 2 gDNA ratio) allele-specific expression1 or 2 = (normalized allele1 or 2 cDNA ratio)/(normalized allele1 cDNA ratio + normalized allele2 cDNA ratio) Samples with mono-allelic expression had ≥9:1 allele-specific expression ratio [5,30,31] as measured by at least 10 positive signal droplets [31,32]. ...
... Our cancer cell line analysis was different in several ways from many previous TERT promoter methylation and expression studies. First, because TERT is often regulated at the allele level [5,29], and in most cancer cells not all TERT alleles are transcribed [5,25], we looked at the two major allelic methylation patterns from each cell line separately, instead of averaging them together [24,37]. This prevented masking of the methylation characteristics of the active allele due to mixing with those of the silent allele. ...
... Third, we searched for both TERT promoter point mutations and TERT structural rearrangements to identify TERT genetically altered samples; sometimes promoter mutations are taken into account while rearrangements are overlooked. Finally, we measured decitabine's effect on TERT allelespecific expression levels instead of total TERT mRNA expression levels [20,29,36,37,[49][50][51]. This approach revealed previously unappreciated findings in the methylation profiles and responses to demethylating agents between wild-type and genetically altered cancer cell lines. ...
Background:
TERT promoter methylation, located several hundred base pairs upstream of the transcriptional start site, is cancer specific and correlates with increased TERT mRNA expression and poorer patient outcome. Promoter methylation, however, is not mutually exclusive to TERT activating genetic alterations, as predicted for functionally redundant mechanisms. To annotate the altered patterns of TERT promoter methylation and their relationship with gene expression, we applied a Pacific Biosciences-based, long-read, bisulfite-sequencing technology and compared the differences in the methylation marks between wild-type and mutant cancers in an allele-specific manner.
Results:
We cataloged TERT genetic alterations (i.e., promoter point mutations or structural variations), allele-specific promoter methylation patterns, and allele-specific expression levels in a cohort of 54 cancer cell lines. In heterozygous mutant cell lines, the mutant alleles were significantly less methylated than their silent, mutation-free alleles (p < 0.05). In wild-type cell lines, by contrast, both epialleles were equally methylated to high levels at the TERT distal promoter, but differentially methylated in the proximal regions. ChIP analysis showed that epialleles with the hypomethylated proximal and core promoter were enriched in the active histone mark H3K4me2/3, whereas epialleles that were methylated in those regions were enriched in the repressive histone mark H3K27me3. Decitabine therapy induced biallelic expression in the wild-type cancer cells, whereas the mutant cell lines were unaffected.
Conclusions:
Long-read bisulfite sequencing analysis revealed differences in the methylation profiles and responses to demethylating agents between TERT wild-type and genetically altered cancer cell lines. The causal relation between TERT promoter methylation and gene expression remains to be established.
... The locus at 5p15.33 is a well-known multi-cancer locus (52,53) and was associated with lung cancer across different populations (12,14,16). The most robust signal is tagged by rs2736100, which was consistently observed in European and East Asian populations among smokers and never-smokers. ...
... A multi-cancer meta-analysis including lung can-cer in this locus identified six independent regions (52). Functional characterization of one of them identified rs36115365 as a functional variant (tagged by rs37004) which is located in an intergenic region between TERT (∼18 kb telomeric from the SNP) and CLPTM1L (Cleft Lip and Palate Transmembrane Protein 1-Like; ∼5 kb centromeric from the SNP) (53). The lung cancer-protective C allele of rs36115365 preferentially recruits a transcription factor, ZNF148, which increases TERT expression, but not CLPTM1L, in lung cancer cell lines. ...
Fourteen years after the first genome-wide association study (GWAS) of lung cancer was published, approximately forty-five genomic loci have now been significantly associated with lung cancer risk. While functional characterization was performed for several of these loci, a comprehensive summary of current molecular understanding of lung cancer risk has been lacking. Further, many novel computational and experimental tools now became available to accelerate the functional assessment of disease-associated variants, moving beyond locus-by-locus approaches. In this review, we first highlight the heterogeneity of lung cancer GWAS findings across histological subtypes, ancestries, and smoking status, which poses unique challenges to follow-up studies. We then summarize the published lung cancer post-GWAS studies for each risk-associated locus to assess the current understanding of biological mechanisms beyond the initial statistical association. We further summarize strategies for GWAS functional follow-up studies considering cutting-edge functional genomics tools and providing a catalog of available resources relevant to lung cancer. Overall, we aim to highlight the importance of integrating computational and experimental approaches to draw biological insights from the lung cancer GWAS results beyond association.
... The high degree of freedom noted in these positions (Fig. 3) implies that when designing a promotor sequence, it is advisable to examine all positions that interact or are predicted to interact with the protein by the phosphate backbone rather than the nitrogenous base. Fang et al. showed that a single SNP in a DNA binding recognition site can influence transcription factor binding, thereby affecting gene regulation 39 . Thus, our engineering of synthetic promoters is also relevant for natural variations among humans. ...
Cancer immunotherapies are highly potent and are gaining wide clinical usage. However, severe side effects require focusing effector immune cell activities on the tumor microenvironment (TME). We recently developed a chimeric antigen receptor tumor-induced vector (CARTIV), a synthetic promoter activated by TME factors. To improve CARTIV functions including background, activation levels, and synergism, we screened a library of promoters with variations in key positions. Here, we present a screening method involving turning ON/OFF stimulating TNFα and IFNγ cytokines, followed by sequential cell sorting. Sequencing of enriched promoters identified seventeen candidates, which were cloned and whose activities were then validated, leading to the identification of two CARTIVs with lower background and higher induction. We further combined a third hypoxia element with the two-factor CARTIV, demonstrating additional modular improvement. Our study presents a method of fine-tuning synthetic promoters for desired immunotherapy needs.
... Overall, these findings suggest that the same biological pathway has opposite effects on the susceptibility to different tumor types. This interpretation is supported by functional characterization of rs36115365, a variant on 5p15.33, which was found to have similar cis-regulatory effects on TERT in multiple cancers cell lines from different cancers, but was associated with a higher risk of pancreatic and testicular cancer and a lower risk of lung cancer [33]. Alternatively, a causal variant may differently influence cis-gene regulation and/or alter different biological pathways depending on the cell or tissue of origin [34]. ...
Background Genome-wide association studies (GWAS) have identified multiple common breast cancer susceptibility variants. Many of these variants have differential associations by estrogen receptor (ER) status, but how these variants relate with other tumor features and intrinsic molecular subtypes is unclear. Methods Among 106,571 invasive breast cancer cases and 95,762 controls of European ancestry with data on 173 breast cancer variants identified in previous GWAS, we used novel two-stage polytomous logistic regression models to evaluate variants in relation to multiple tumor features (ER, progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) and grade) adjusting for each other, and to intrinsic-like subtypes. Results Eighty-five of 173 variants were associated with at least one tumor feature (false discovery rate < 5%), most commonly ER and grade, followed by PR and HER2. Models for intrinsic-like subtypes found nearly all of these variants (83 of 85) associated at p < 0.05 with risk for at least one luminal-like subtype, and approximately half (41 of 85) of the variants were associated with risk of at least one non-luminal subtype, including 32 variants associated with triple-negative (TN) disease. Ten variants were associated with risk of all subtypes in different magnitude. Five variants were associated with risk of luminal A-like and TN subtypes in opposite directions. Conclusion This report demonstrates a high level of complexity in the etiology heterogeneity of breast cancer susceptibility variants and can inform investigations of subtype-specific risk prediction.
... Given the extremely aggressive behavior and poor prognosis of both melanoma and PC, the identification of individuals at risk for one or both of these cancers and their close surveillance is of utmost importance. Hence, further research has led to the detection of multiple genetic factors that modify the risk of melanoma and PC in p16-Leiden mutation carriers, such as melanocortin 1 receptor gene (MC1R) variants that influence the risk of melanoma (28,29), rs36115365-C, a single-nucleotide polymorphism which controls TERT expression and is associated with increased risk of PC and decreased risk of melanoma (30,31), mutations in glutathione S-transferase genes GSTM1 and GSTT1 (32), as well as in the vitamin D receptor gene that appear to have a slight protective effect against melanoma (33). ...
Multiple primary cancers may occur in the same patient, with a prevalence that follows an ascendant trend. Their development is dictated by a complex interplay between a variety of factors, both patient-dependent and external. The case of a 38-year-old female patient diagnosed and treated for pancreatic cancer (PC) is presented in whom the digital dermoscopic monitoring of melanocytic nevi revealed a marked change of two nevi that acquired rapidly highly atypical features. They were surgically excised and the histopathological examination revealed two completely excised dysplastic compound nevi. Clinicians should be aware of the strong association between dysplastic nevus syndrome and PC, a malignancy associated with an extremely poor prognosis. Familial atypical multiple mole melanoma syndrome (FAMMM) predisposes to the development of melanoma, pancreatic cancer and other neoplasms. The common genetic background of PC and hereditary melanoma is discussed and the importance of regular skin checkup and screening for PC in these patients is underlined.
