In line with their estrogen

In line with their estrogen responsiveness and activation of the 12-O-tetradecanoyl phorbol-13-acetate Supplier at puberty, the CD24+ CD49fhi SCA-1+ cells were found to be enriched in TEBs. TEBs are highest in the pubertal animal and are considered the proliferative unit of the developing gland (Russo and Russo, 1978a, 1978b, 1980). This agrees with our data showing that the SCA-1+ TEBS are abundant and actively cycling at puberty. TEBs have been associated with the carcinogen sensitivity of the mammary gland, yet whether the stem cells or the proliferative nature of the TEBs is responsible is unclear (Russo et al., 1979). Our data indicate that the proliferating SCA-1+ subset of MaSCs within the TEBs are likely mediating the carcinogen sensitivity. Here, we have characterized an ER-positive and estrogen-sensitive population within the CD24+ CD49fhi MaSC-enriched compartment. Mammary repopulating activity, while present, does not appear to be a predominant function, instead their abundance in the young mammary gland and estrogen sensitivity indicate a role in pubertal mammary development and hormonal sensitivity early in life.
Experimental Procedures
Author Contributions
Acknowledgments G.D., Australian Postgradutate Scholarship; K.B., NBCF ECR Fellowship (ECF 11-01), NHMRC New Investigator grant (APP1044661), and VCA ECR fellowship (ECSG08_07); R.L.A., NBCF Senior Fellowship; K.S.K., Division of Intramural Research/NIEHS [1ZIAESO70065]; G.P.R., NHMRC fellowship. We thank Dr. Carl Walkey (St Vincents Institute) and A/Prof Steve Lane (QIMR Berghofer) for their helpful discussions about cell-cycle-specific staining, the flow core facility at Monash University, and the FACS facility at Peter Mac as well as Monash Micro Imaging Facility and Peter Mac microscopy groups for provision of instrumentation, training, and general support.
Introduction The ability to generate induced pluripotent stem cells (iPSCs) has been heralded to have great potential in regenerative medicine and research (Yu et al., 2007; Takahashi and Yamanaka, 2006). However, this potential is currently under debate, due to evidence that iPSCs can acquire DNA damage and genomic instability during the reprogramming process (Ruiz et al., 2015; Liang and Zhang, 2013; Gore et al., 2011; Mayshar et al., 2010). In fact, even just expressing the reprogramming factors, regardless of the methodology used to generate them, causes DNA damage, mainly by replication stress (Ruiz et al., 2015; Gonzalez et al., 2013; Tilgner et al., 2013). It is critical, however, to obtain “safe” iPSCs that are genetically identical to their parent cells for clinical use. An essential prerequisite for this is to obtain a thorough understanding about how the DNA repair machinery acts in these cells. Several pieces of evidence suggest that pluripotent stem cells need more active DNA repair pathways than somatic differentiated cells (Rocha et al., 2013). Supporting this view, members of the DNA damage response (DDR) have been shown to prevent genomic instability in iPSCs (Hong et al., 2009; Kawamura et al., 2009; Li et al., 2009). Indeed, proteins involved in the repair of DNA double-strand breaks (DSBs), in both homologous recombination (HR) and non-homologous end joining (NHEJ), have a relevant role in reprogramming efficiency (Gonzalez et al., 2013; Ruiz et al., 2013; Tilgner et al., 2013). HR is required for an error-free repair of DSBs, using homologous sequences (normally from the sister chromatid) (Heyer et al., 2010), and for restarting replication forks stalled during replication stress (Petermann and Helleday, 2010). In contrast, NHEJ competes with HR for DSB repair in a more error-prone pathway (Gomez-Cabello et al., 2013; Huertas, 2010; Lieber, 2008). DNA end resection is a key event that regulates the DSB repair pathway choice between NHEJ and HR. This mechanism generates single-strand DNA (ssDNA) by 5′ to 3′ degradation at both sides of a break (Huertas and Jackson, 2009; Jackson and Bartek, 2009). Although resected DNA is an obligate substrate for HR, it blocks NHEJ (Heyer et al., 2010). CtIP is a major player in the decision between HR and NHEJ as it allows for ssDNA formation, precluding binding of the NHEJ machinery to DNA breaks (Huertas, 2010). DNA end resection is highly regulated by multiple signals, including cell-cycle-dependent CtIP phosphorylation (Sartori et al., 2007). Cells depleted of CtIP fail to repair DNA DSBs by HR, are sensitive to DNA damaging agents, and accumulate chromosomal aberrations in response to DNA damage (Sartori et al., 2007; Huertas and Jackson, 2009).