The development of multicellular, sexually reproductive organisms starts with the fusion of a spermatozoa and an oocyte that created a diploid zygote (depicted in Figure 1). Then, the zygote divides, forming a cluster of undifferentiated cells (i.e., blastomeres) known as the morula. The first step of lineage differentiation occurs during the subsequent blastocyst stage in which the embryoblast (i.e., the inner cell mass) and the trophectoderm are formed [1,2]. The trophectoderm develops into all extra-embryonic structures, whereas the embryoblast, which consists of pluripotent embryonic stem (ES) cells, develops into all embryonic structures [2]. ES cells are transiently present, can self-renew and give rise to all three embryonic germ layers during gastrulation (i.e., the ecto-, meso-, and endoderm), which ultimately will give rise to all cell lineages of the adult organism [2]. Additionally, ES cells give rise to the primordial germ cells (PGC) in the embryonic yolk sac, which subsequently migrate toward developing gonads to give rise to the germ line [2,3] (see Figure 1).

The pluripotent nature of ES cells is characterized by the expression of pluripotency markers including but not limited to OCT4, NANOG, SOX2, and REX1 (Figure 1) [4]. In contrast, somatic cells are differentiated and thus lineage-restricted and unipotent [5]. Therefore, these cells seldomly give rise to other progeny than the identity of the cells themselves [5]. In addition, due to their capacity to self-renew, ES cells can be cultured indefinitely in vitro while retaining their embryonic state as well as a stable genome. This immortality is facilitated by active telomerase, which is an enzyme that extends telomeric repeats that are otherwise lost due to the end-replication problem after each cell cycle (approximately 50–150 base pairs are lost per cycle) [6,7] (Figure 1). Conversely, somatic cells have a restricted lifespan due to the lack of active telomerase and induce cellular senescence when a critical telomere length is reached [6,7]. Furthermore, ES cells also regulate the cell cycle differently as these lack Cyclin D expression yet continuously express Cyclin A and E, which leads to a significantly shortened G₁-phase when compared to somatic cells [8]. This characteristic also facilitates the tendency for ES cells to initiate apoptosis rather than cell cycle arrest and DNA repair when DNA damage occurs, the latter of which occurs predominantly in the G₁ phase after cell cycle arrest [9,10]. The preference for apoptosis in these early embryonic cells is considered a failsafe mechanism to preserve the genetic integrity of their multitudinous progeny. In rare occasions, ES cells can initiate DNA repair; however, these cells then opt for error-free homologous recombination (HR) to repair double-strand breaks, whereas somatic cells predominantly employ error-prone non-homologous end joining (NHEJ) [9]. Finally, a zygote is known to lose its inherited DNA methylation pattern immediately after fertilization and subsequent changes in DNA methylation occur during early embryonic development as various cell lineages arise. Thus, DNA methylation patterns between ES and somatic cells vary significantly [11,12].

Of note, a discovery made over a decade ago demonstrated that differentiated somatic human cells (e.g., adult human fibroblasts) could be reprogrammed to a pluripotent state reminiscent of ES cells (i.e., nuclear reprogramming) [13]. These cells are known as induced pluripotent stem cells (iPSCs) and, in theory, they can also be maintained in a pluripotent state indefinitely. iPSCs are originally generated in vitro by overexpressing a cocktail of four essential genes (OCT3/4, c-MYC, SOX2, and KLF4) in the differentiated cells [14].

Moreover, many of the characteristics of ES cells are strongly conserved in germ cell tumors (GCTs), which represent a heterogeneous cluster of solid tumors that originate from both ES cells and PGCs [3]. Of note, the parallels observed between ES cells and GCTs do not apply to the teratomas, which is a GCT subtype that consists of fully differentiated cells and harbor significant differences in their (epi)genetic makeup and clinical response (i.e., inherent chemotherapeutic resistance) compared to other GCT subtypes [3,15,16,17]. An example of the similarities observed between ES cells and GCTs is that the latter also express multiple pluripotency markers including OCT3/4 and NANOG and have active telomerase activity, again with the exception of teratoma [3]. Moreover, it is widely accepted that in response to DNA damage, GCTs also favor apoptosis, which is thought to underlie their unique sensitivity to platinum-based chemotherapeutics, including cisplatin (a key component of GCT treatment) [9,10,18]. This is supported by additional evidence demonstrating that GCTs have low expression of genes involved in cell cycle arrest and a deregulated G1-S phase checkpoint, thus potentially also preventing the activation of DNA repair pathways [18,19,20,21,22,23]. Moreover, the apoptotic response of ES cells and GCTs in response to DNA damage has been attributed to the well-known P53 pathway; however, at least in GCTs, it remains a topic of debate [18,21,24,25,26,27,28,29,30,31,32,33,34,35,36,37]. The P53 pathway has been dubbed the guardian of the genome due to its integral role in maintaining genomic integrity among all cell types. This pathway acts in response to cellular stress and lies at the apex of a plethora of downstream signaling pathways including cell cycle arrest, DNA repair, and apoptosis. The deregulation of this pathway has been observed in cancer and is strongly associated to many aspects of tumorigenesis. For instance, GCTs most often express high levels of wild-type (WT) TP53, and it has been suggested that the P53 pathway is (partially) responsible for their characteristic to enter apoptosis in response to platinum-based chemotherapeutics (e.g., cisplatin) which is reminiscent of ES cells in response to DNA damage [10,18,21,23,25,26,27,28,29,33,34,35,36,37]. In addition, it has been suggested that the P53 pathway is deregulated through multiple mechanisms in GCTs that have acquired resistance to chemotherapy, including cisplatin [15,18,38,39,40,41,42].

Genomic changes during development serve as the driving force of evolution; however, these may also negatively impact multiple cell lineages with possible detrimental effects on both the individual and their offspring. Therefore, protection of the genomic integrity is paramount for a species to survive and commences during early embryonic development [23]. For example, the oocyte expresses and deposits multiple mRNAs that protect the zygote against DNA damage during the first cellular divisions [43]. As mentioned before, there have been several indications of a robust protective mechanism in ES cells including the removal of mutated ES cells from the stem cell pool through apoptosis, which is a characteristic that is seemingly conserved in GCTs [7,12,44]. Alternatively, ES cells may also be instructed to differentiate through transcriptional inhibition of the pluripotency factor NANOG, which also effectively removes ES cells from the stem cell pool [45]. These mechanisms in combination with lacking a G₁ checkpoint are thought to result in a 100-fold lower frequency of accumulating mutations in at least murine ES cells compared to mouse embryonic fibroblasts, which resemble the somatic cells [9,23]. Conversely to ES cells and GCTs, somatic cells utilize different pathways (DNA repair rather than apoptosis) to protect their genome that enable these to survive while risking an increased mutational load [9,23].

As mentioned before, the P53 pathway is essential in maintaining genomic integrity. Central to this pathway is the P53 protein that is regarded as a tumor suppressor and originates from the TP53 gene. Several key discoveries regarding P53 are depicted in Figure 2, as well as its relevance in GCTs [22,23,24,25,26,27,28,29,30,31,32,33,34,35,36,37,38,46,47,48,49,50,51,52,53,54,55,56,57,58,59]. In response to cellular stress, the P53 protein is activated and accumulates in the cell after which it mainly functions as a transcription factor that transactivates a plethora of downstream targets [49]. These target genes are key players in one of many downstream pathways including cell cycle arrest, DNA repair, apoptosis, senescence, and autophagy [51,60,61]. The P53 protein counts 393 amino acids (AA) and consists of multiple domains, starting with a transactivation domain (TAD) at the amino-terminus (N-terminus), a directly neighboring proline-rich domain (PRD), a large DNA-binding domain (DBD), a tetramerization domain (TD), and a carboxy-terminus (C-terminus) regulatory domain (REG) [62,63] (depicted in Figure 3). Once activated and accumulated in the cell, the P53 protein functions and binds DNA as a tetramer, which is facilitated by the TD [64]. When tetramerized, the DBD contains three loops, L1, L2, and L3 respectively [65]. Both L2 and L3 are bound by a zinc ion, effectively linking both loops and enabling L3 to bind to the minor groove of the DNA, while a helix in the DBD is complementary to the major groove [65,66].

There are many different forms of cellular stress such as DNA damage that activate upstream regulators of p53, leading to the activation of the p53 pathway. For example, Ataxia-telangiectasia mutated (ATM) kinase serves as an important DNA damage sensor, as it recognizes and binds to double-strand DNA breaks and subsequently activates the P53 pathway, ultimately resulting in the activation of NHEJ or HR DNA repair mechanisms [55,67]. ATM phosphorylates histone H2AX, which is a variant of histone H2A that recruits molecules responsible for H3K9 methylation [55]. In turn, methylated H3K9 leads to the acetylation and activation of ATM [55]. Among other substrates, ATM phosphorylates and activates Checkpoint Kinase 1 and 2 (CHK1 and CHK2, respectively) [55]. In turn, CHK1 and 2 function as transcriptional activators by phosphorylating and activating the P53 protein [68]. The subsequent section will further outline several downstream targets and their related pathways including cell cycle arrest, senescence, apoptosis, and autophagy (also summarized in Figure 4).