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BMP Inhibition in Seminomas Initiates Acquisition of Pluripotency via NODAL Signaling Resulting in Reprogramming to an Embryonal Carcinoma.

Nettersheim D, Jostes S, Sharma R, Schneider S, Hofmann A, Ferreira HJ, Hoffmann P, Kristiansen G, Esteller MB, Schorle H - PLoS Genet. (2015)

Bottom Line: Analysis of expression and DNA methylation dynamics during transition of TCam-2 revealed that many pluripotency- and reprogramming-associated genes were upregulated while seminoma-markers were downregulated.We demonstrate that inhibition of BMP signaling is the initial event in reprogramming, resulting in activation of the pluripotency-associated genes and NODAL signaling.In parallel, DNMT3B-driven de novo methylation silences seminoma-associated genes and epigenetically fixes the EC state.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pathology, Department of Developmental Pathology, University Medical School, Bonn, Germany.

ABSTRACT
Type II germ cell cancers (GCC) can be subdivided into seminomas and non-seminomas. Seminomas are similar to carcinoma in situ (CIS) cells, the common precursor of type II GCCs, with regard to epigenetics and expression, while embryonal carcinomas (EC) are totipotent and differentiate into teratomas, yolk-sac tumors and choriocarcinomas. GCCs can present as seminomas with a non-seminoma component, raising the question if a CIS gives rise to seminomas and ECs at the same time or whether seminomas can be reprogrammed to ECs. In this study, we utilized the seminoma cell line TCam-2 that acquires an EC-like status after xenografting into the murine flank as a model for a seminoma to EC transition and screened for factors initiating and driving this process. Analysis of expression and DNA methylation dynamics during transition of TCam-2 revealed that many pluripotency- and reprogramming-associated genes were upregulated while seminoma-markers were downregulated. Changes in expression level of 53 genes inversely correlated to changes in DNA methylation. Interestingly, after xenotransplantation 6 genes (GDF3, NODAL, DNMT3B, DPPA3, GAL, AK3L1) were rapidly induced, followed by demethylation of their genomic loci, suggesting that these 6 genes are poised for expression driving the reprogramming. We demonstrate that inhibition of BMP signaling is the initial event in reprogramming, resulting in activation of the pluripotency-associated genes and NODAL signaling. We propose that reprogramming of seminomas to ECs is a multi-step process. Initially, the microenvironment causes inhibition of BMP signaling, leading to induction of NODAL signaling. During a maturation phase, a fast acting NODAL loop stimulates its own activity and temporarily inhibits BMP signaling. During the stabilization phase, a slow acting NODAL loop, involving WNTs re-establishes BMP signaling and the pluripotency circuitry. In parallel, DNMT3B-driven de novo methylation silences seminoma-associated genes and epigenetically fixes the EC state.

No MeSH data available.


Related in: MedlinePlus

Meta-analysis of Gex and 5mC data of xenografted TCam-2 and GCC tissues.(A, B) Venn diagrams summarizing commonly up- (A) and downregulated (B) genes between seminoma, TCam-2 xenografted for 1 and 6 weeks and ECs. Corresponding data is given in (E and H in S1 Data). Genes recorded in duplicates (due to multiple probes on the array) were included only once. (C) Expression intensities of indicated genes in TCam-2 xenografted for 6 weeks and the EC group as fold change versus appropriate controls (TCam-2 in vitro /seminoma tissues). Genes were categorized (color coded) based on [48] [18] [35]. (D) 5mC levels of indicated genes in seminomas, parental and xenografted TCam-2 as well as EC cell lines (2102EP, NEC8, 833KE, SuSa) as found by 450k microarray analysis. (E) BDCP analysis of 5mC data of genes and samples indicated in (D).
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pgen.1005415.g003: Meta-analysis of Gex and 5mC data of xenografted TCam-2 and GCC tissues.(A, B) Venn diagrams summarizing commonly up- (A) and downregulated (B) genes between seminoma, TCam-2 xenografted for 1 and 6 weeks and ECs. Corresponding data is given in (E and H in S1 Data). Genes recorded in duplicates (due to multiple probes on the array) were included only once. (C) Expression intensities of indicated genes in TCam-2 xenografted for 6 weeks and the EC group as fold change versus appropriate controls (TCam-2 in vitro /seminoma tissues). Genes were categorized (color coded) based on [48] [18] [35]. (D) 5mC levels of indicated genes in seminomas, parental and xenografted TCam-2 as well as EC cell lines (2102EP, NEC8, 833KE, SuSa) as found by 450k microarray analysis. (E) BDCP analysis of 5mC data of genes and samples indicated in (D).

Mentions: Now, we wanted to understand whether the changes in 5mC correlate to changes in Gex. A Pearson’s correlation of the microarray data identified 601 genes, showing inverse correlation between 5mC and Gex (S1B Data). A BDPC methylation cluster analysis of these 601 genes demonstrates that the transplanted cells cluster to the parental TCam-2 cell line up to 2 weeks after transplantation. Thereafter, they cluster more to the 2102EP to become highly similar after 6 weeks after transplantation (Fig 3B).


BMP Inhibition in Seminomas Initiates Acquisition of Pluripotency via NODAL Signaling Resulting in Reprogramming to an Embryonal Carcinoma.

Nettersheim D, Jostes S, Sharma R, Schneider S, Hofmann A, Ferreira HJ, Hoffmann P, Kristiansen G, Esteller MB, Schorle H - PLoS Genet. (2015)

Meta-analysis of Gex and 5mC data of xenografted TCam-2 and GCC tissues.(A, B) Venn diagrams summarizing commonly up- (A) and downregulated (B) genes between seminoma, TCam-2 xenografted for 1 and 6 weeks and ECs. Corresponding data is given in (E and H in S1 Data). Genes recorded in duplicates (due to multiple probes on the array) were included only once. (C) Expression intensities of indicated genes in TCam-2 xenografted for 6 weeks and the EC group as fold change versus appropriate controls (TCam-2 in vitro /seminoma tissues). Genes were categorized (color coded) based on [48] [18] [35]. (D) 5mC levels of indicated genes in seminomas, parental and xenografted TCam-2 as well as EC cell lines (2102EP, NEC8, 833KE, SuSa) as found by 450k microarray analysis. (E) BDCP analysis of 5mC data of genes and samples indicated in (D).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4520454&req=5

pgen.1005415.g003: Meta-analysis of Gex and 5mC data of xenografted TCam-2 and GCC tissues.(A, B) Venn diagrams summarizing commonly up- (A) and downregulated (B) genes between seminoma, TCam-2 xenografted for 1 and 6 weeks and ECs. Corresponding data is given in (E and H in S1 Data). Genes recorded in duplicates (due to multiple probes on the array) were included only once. (C) Expression intensities of indicated genes in TCam-2 xenografted for 6 weeks and the EC group as fold change versus appropriate controls (TCam-2 in vitro /seminoma tissues). Genes were categorized (color coded) based on [48] [18] [35]. (D) 5mC levels of indicated genes in seminomas, parental and xenografted TCam-2 as well as EC cell lines (2102EP, NEC8, 833KE, SuSa) as found by 450k microarray analysis. (E) BDCP analysis of 5mC data of genes and samples indicated in (D).
Mentions: Now, we wanted to understand whether the changes in 5mC correlate to changes in Gex. A Pearson’s correlation of the microarray data identified 601 genes, showing inverse correlation between 5mC and Gex (S1B Data). A BDPC methylation cluster analysis of these 601 genes demonstrates that the transplanted cells cluster to the parental TCam-2 cell line up to 2 weeks after transplantation. Thereafter, they cluster more to the 2102EP to become highly similar after 6 weeks after transplantation (Fig 3B).

Bottom Line: Analysis of expression and DNA methylation dynamics during transition of TCam-2 revealed that many pluripotency- and reprogramming-associated genes were upregulated while seminoma-markers were downregulated.We demonstrate that inhibition of BMP signaling is the initial event in reprogramming, resulting in activation of the pluripotency-associated genes and NODAL signaling.In parallel, DNMT3B-driven de novo methylation silences seminoma-associated genes and epigenetically fixes the EC state.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pathology, Department of Developmental Pathology, University Medical School, Bonn, Germany.

ABSTRACT
Type II germ cell cancers (GCC) can be subdivided into seminomas and non-seminomas. Seminomas are similar to carcinoma in situ (CIS) cells, the common precursor of type II GCCs, with regard to epigenetics and expression, while embryonal carcinomas (EC) are totipotent and differentiate into teratomas, yolk-sac tumors and choriocarcinomas. GCCs can present as seminomas with a non-seminoma component, raising the question if a CIS gives rise to seminomas and ECs at the same time or whether seminomas can be reprogrammed to ECs. In this study, we utilized the seminoma cell line TCam-2 that acquires an EC-like status after xenografting into the murine flank as a model for a seminoma to EC transition and screened for factors initiating and driving this process. Analysis of expression and DNA methylation dynamics during transition of TCam-2 revealed that many pluripotency- and reprogramming-associated genes were upregulated while seminoma-markers were downregulated. Changes in expression level of 53 genes inversely correlated to changes in DNA methylation. Interestingly, after xenotransplantation 6 genes (GDF3, NODAL, DNMT3B, DPPA3, GAL, AK3L1) were rapidly induced, followed by demethylation of their genomic loci, suggesting that these 6 genes are poised for expression driving the reprogramming. We demonstrate that inhibition of BMP signaling is the initial event in reprogramming, resulting in activation of the pluripotency-associated genes and NODAL signaling. We propose that reprogramming of seminomas to ECs is a multi-step process. Initially, the microenvironment causes inhibition of BMP signaling, leading to induction of NODAL signaling. During a maturation phase, a fast acting NODAL loop stimulates its own activity and temporarily inhibits BMP signaling. During the stabilization phase, a slow acting NODAL loop, involving WNTs re-establishes BMP signaling and the pluripotency circuitry. In parallel, DNMT3B-driven de novo methylation silences seminoma-associated genes and epigenetically fixes the EC state.

No MeSH data available.


Related in: MedlinePlus