TICB-1043; No. of Pages 7 Opinion Stem cells: balancing resistance and sensitivity to DNA damage 1 2,3 1 Julia C. Liu , Paul H. Lerou , and Galit Lahav 1 Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA 2 Department of Pediatric Newborn Medicine, Brigham and Women’s Hospital, Boston, MA 02115, USA 3 Department of Medicine, Division of Genetics, Brigham and Women’s Hospital, Boston, MA 02115, USA Embryonic stem cells (ESCs) are known to be very sen- they must block the propagation of mutations to their sitive to DNA damage and undergo rapid apoptosis even progeny cells, because fixed mutations can have long-term after low-damage doses. By contrast, adult stem cells functional consequences including predisposition to malig- show variable sensitivity to damage. Here we describe nancy. On the other hand, stem cells have a responsibility the multiple pathways that have been proposed to affect to maintain existing tissue organization. Here we describe the sensitivity of stem cells to damage, including prox- the current understanding of the DNA damage response in imity to the apoptotic threshold (mitochondrial priming) hESCs, in particular how their sensitivity is maintained and the p53 signaling pathway, through activation of close to the apoptotic threshold. We also discuss the vari- transcription or direct interaction with proapoptotic ability in sensitivity of adult stem cells to DNA damage and proteins in the cytoplasm. We also discuss which cellular propose multiple regulatory networks that may affect the factors might connect mitochondrial priming with plur- sensitivity of adult stem cells in different tissues. ipotency and the potential therapeutic advances that can be achieved by better understanding of the molecular Multiple distinct mechanisms sensitize hESCs to DNA mechanisms leading to sensitivity or resistance of em- damage bryonic or adult stem cells from different tissues. Many studies have shown that hESCs have higher rates of apoptosis after DNA damage than differentiated cells Stem cells must respond appropriately to DNA damage [4–6], yet this phenomenon remains incompletely under- A major component of cellular aging is the detrimental stood. Several distinct mechanisms have been proposed to accumulation of mutations in the cell’s DNA. Mutations explain how the regulatory networks that control apoptosis that facilitate deregulated proliferation or predispose cells might have unique functions in hESCs (Figure 1). Among to acquire further mutations are often associated with the these mediators is the tumor suppressor protein p53. Well development of cancer [1]. Cellular stress, such as DNA known as a regulator of cell fate decisions in somatic cells damage, contributes to the accrual of such deleterious [10], p53 is also induced after various types of DNA damage mutations and therefore cells mount responses to guard in hESCs. Different classes of DNA lesions, including DNA against genomic instability [2]. Different cell types favor double-strand breaks (DSBs) and single-stranded DNA different responses to cellular stress, ranging from cell (ssDNA), can be generated by various damaging agents, cycle arrest and DNA repair to senescence or apoptosis such as g-ray irradiation and UV irradiation, respectively [3]. ESCs, both human (hESCs) and mouse, are known to [11]. Apoptosis of hESCs in response to both DSBs and be acutely sensitive to DNA damage [4–8]. As an important ssDNA was shown to be dependent on p53 [4,5]. However, cell population that gives rise to all of the tissues in the multiple mechanisms have been proposed to explain how body, ESCs are expected to undergo apoptosis after dam- p53 induces the intrinsic (mitochondrial) apoptotic path- age to prevent damaged cells from compromising the ge- way (Box 1) in hESCs, including p53 transcriptional acti- nomic integrity of the population. Conversely, adult stem vation of proapoptotic target genes, direct interaction of cells are more resistant to cell death after DNA damage, p53 with mitochondrial proteins, and p53-mediated trans- possibly to prevent uncontrolled apoptosis that might com- location of the proapoptotic protein Bax. promise tissue and organ structure. However, this resis- p53 activates the transcription of multiple genes in- tance may lead to genomic instability if damage-induced volved in apoptosis, such as BAX, PUMA, NOXA, and mutations are not properly repaired [9]. Hence, stem cells APAF1 [12]. On treatment with the DNA-damaging drug are caught between two opposing needs. On the one hand, etoposide [13], p53 was found to translocate to the nucleus of hESCs and to upregulate PUMA [5]. Independently of its Corresponding author: Lahav, G. ([email protected]). transcriptional activity, p53 can promote apoptosis by Keywords: apoptosis; hESCs; mitochondrial priming; p53. directly interacting with mitochondrial proteins [14]. It 0962-8924/$ – see front matter was shown in hESCs treated with UV irradiation that ß 2014 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.tcb.2014.03.002 p53-dependent apoptosis occurred without transcription of p53 target genes; instead, p53 associated with the mitochondria [4]. It is currently unclear why in some cases p53 acts as a transcription factor whereas in others it Trends in Cell Biology xx (2014) 1–7 1 TICB-1043; No. of Pages 7 Opinion Trends in Cell Biology xxx xxxx, Vol. xxx, No. x ( 2) P rot i ein p53 nte – on ra p ip c ro cr o t (3) Mitochondrial ns n e a in Tr priming ) (1 Pro– Pro- Mitochondria Apoptosis Nucleus e Bax (4) Bax n Golgi TRENDS in Cell Biology Figure 1. Multiple pathways contribute to human embryonic stem cell (hESC) sensitivity to DNA damage. (1) The tumor suppressor protein p53 acts as a transcription factor, upregulating the expression of proapoptotic target genes. The proapoptotic protein products proceed to induce apoptosis via the mitochondrial pathway. (2) p53 can act independently of transcription to directly interact with cytoplasmic proapoptotic and antiapoptotic proteins, thereby promoting apoptosis. (3) High mitochondrial priming, implying a high ratio of proapoptotic to antiapoptotic proteins, maintains hESCs close to the apoptotic threshold. (4) The apoptotic protein Bax is localized in its active conformation at the Golgi apparatus in some hESCs, poised to induce rapid apoptosis after damage-induced translocation to the mitochondria. activates apoptosis through interacting with mitochondri- when transcription was inhibited, suggesting that al- al proteins. The specific mechanism of activation might be though p53 transcriptional activity is functional, it is not dependent on the type of DNA damage, although further required for apoptosis [16]. Differentiated cells showed studies are required to better understand the specific similar induction of apoptotic targets, suggesting that relationship between the type of damage and p53 function p53 transcriptional activity does not distinguish hESCs in hESCs. Recently, the proapoptotic protein Bax was from differentiated cells. Looking further, p53 was ob- shown to be a major activator of apoptosis in hESCs served in the cytoplasm of hESCs and a cytoplasm-confined dependent on p53. Unlike differentiated cells, hESCs were p53 mutant confirmed that cytoplasmic p53 alone is able to found to maintain Bax in its active conformation at the induce apoptosis. However, cytoplasmic p53 was also pres- Golgi apparatus in basal conditions, whereas previously ent in differentiated cells. These results suggest that, Bax activation was thought to be exclusive to apoptotic although p53 signaling appears similar in hESCs and cells [15]. In hESCs damaged with the DNA-damaging differentiated cells, the internal environment of the two drug etoposide, Bax rapidly localizes to the mitochondria cell types may differ, explaining their differential sensitiv- and initiates apoptosis. Although Bax translocation was ity to DNA damage. Indeed, DNA damage sensitivity in shown to be dependent on p53, the exact mechanism hESCs was shown to correlate with a cell-intrinsic proper- generating the translocation remains unclear. Notably, ty termed mitochondrial priming [16]. Mitochondrial prim- the basal level of active Bax varies substantially among ing is determined by the balance between proapoptotic and different hESC lines and is not detectable in the H1 cell antiapoptotic Bcl-2 family proteins and can predict a cell’s line. Nonetheless, H1 cells show the typical hESC sensi- sensitivity to DNA-damaging drugs [17]. It can be mea- tivity to DNA damage [16], suggesting that basally acti- sured via Bcl-2 homology domain 3 (BH3) profiling, which vated Bax is not required for induction of apoptosis, and quantifies the magnitude of mitochondrial outer mem- hence the other mechanisms presented here can be suffi- brane permeabilization (MOMP) on exposure to a panel cient to induce apoptosis (Figure 1). of proapoptotic promiscuously interacting BH3 peptides. The specific mechanisms leading to DNA damage-in- BH3 profiling revealed that hESCs exhibited higher mito- duced apoptosis in hESCs were recently investigated by chondrial priming than their differentiated progeny and simultaneously probing the DNA damage response in hence were closer to the apoptotic threshold [16]. Boosting differentiated cells and comparing it with the response the priming of differentiated cells using ABT-263, an in- in their differentiated progeny [16]. Consistent with previ- hibitor against antiapoptotic Bcl-2 family proteins, led to ous findings, activation of apoptosis