Transcription Factor TFAP2C Regulates Major Programs Required for Murine Fetal Germ Cell Maintenance and Haploinsufficiency Predisposes to Teratomas in Male Mice

Jana Schemmer,Marcos J Araúzo-Bravo,Daniel Nettersheim,Andreas Zimmer,Astrid Becker,Hubert Schorle,Sabine Schäfer,Natalie Haas,Susanne N Weber,Dawid Eckert,Stefan Schlatt

doi:10.1371/journal.pone.0071113

Abstract

Maintenance and maturation of primordial germ cells is controlled by complex genetic and epigenetic cascades, and disturbances in this network lead to either infertility or malignant aberration. Transcription factor TFAP2C has been described to be essential for primordial germ cell maintenance and to be upregulated in several human germ cell cancers. Using global gene expression profiling, we identified genes deregulated upon loss of Tfap2c in embryonic stem cells and primordial germ cell-like cells. We show that loss of Tfap2c affects many aspects of the genetic network regulating germ cell biology, such as downregulation of maturation markers and induction of markers indicative for somatic differentiation, cell cycle, epigenetic remodeling and pluripotency. Chromatin-immunoprecipitation analyses demonstrated binding of TFAP2C to regulatory regions of deregulated genes (Sfrp1, Dmrt1, Nanos3, c-Kit, Cdk6, Cdkn1a, Fgf4, Klf4, Dnmt3b and Dnmt3l) suggesting that these genes are direct transcriptional targets of TFAP2C in primordial germ cells. Since Tfap2c deficient primordial germ cell-like cells display cancer related deregulations in epigenetic remodeling, cell cycle and pluripotency control, the Tfap2c-knockout allele was bred onto 129S2/Sv genetic background. There, mice heterozygous for Tfap2c develop with high incidence germ cell cancer resembling human pediatric germ cell tumors. Precursor lesions can be observed as early as E16.5 in developing testes displaying persisting expression of pluripotency markers. We further demonstrate that mice with a heterozygous deletion of the TFAP2C target gene Nanos3 are also prone to develop teratomas. These data highlight TFAP2C as a critical and dose-sensitive regulator of germ cell fate.

Highlights

Germ cell cancers (GCCs) are usually diagnosed between the age of 20–40 years and are the most common cancer type of young men [1]
In order to analyze the molecular consequences of Tfap2c deficiency in primordial germ cells (PGCs) we took advantage of protocols allowing PGC-like cells (PGCLCs) derivation from embryonic stem cells through an epiblastlike cell (EpiLC) intermediate [24]
The embryonic stem cells (ESCs)-lines were transiently transfected with a plasmid encoding the Cre-Protein mediating the deletion of the Tfap2cflox/flox alleles resulting in Blimp1mVenus/ Tfap2c2/2 ESCs (#1-Tfap2c2/2 and #2-Tfap2c2/2) (Fig. S1 A, B, C)

Summary

Introduction

Germ cell cancers (GCCs) are usually diagnosed between the age of 20–40 years and are the most common cancer type of young men [1]. In infants and pre-pubertal adolescents, teratomas and yolk-sac tumors (Type I GCC) are detected in gonads, cranium or along the body midline. These tumors characteristically consist of tissues of all three germ layers and are generally benign in nature, with rare malignant transformation. TFAP2C (Tcfap2c/AP-2c) is a member of the activator protein-2 (AP-2) family, which comprises of five closely related members, namely Tfap2a-e. They are characterized by a highly conserved DNA-binding and dimerization motif at the C-terminus. High levels of TFAP2C protein were observed in precursor lesions of GCC (carcinoma in situ (CIS)) and classical seminomas [19;20]

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Results

Conclusion