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Alt-EJ is an error-prone DNA double-strand break (DSBs) repair pathway coming to the fore when first-line repair pathways, c-NHEJ and HR, are defective or fail. It is thought to benefit from DNA end-resection—a process whereby 3′ single-stranded DNA-tails are generated—initiated by the CtIP/MRE11-RAD50-NBS1 (MRN) complex and extended by EXO1 or the BLM/DNA2 complex. The connection between alt-EJ and resection remains incompletely characterized. Alt-EJ depends on the cell cycle phase, is at maximum in G₂-phase, substantially reduced in G₁-phase and almost undetectable in quiescent, G₀-phase cells. The mechanism underpinning this regulation remains uncharacterized. Here, we compare alt-EJ in G₁- and G₀-phase cells exposed to ionizing radiation (IR) and identify CtIP-dependent resection as the key regulator. Low levels of CtIP in G₁-phase cells allow modest resection and alt-EJ, as compared to G₂-phase cells. Strikingly, CtIP is undetectable in G₀-phase cells owing to APC/C-mediated degradation. The suppression of CtIP degradation with bortezomib or CDH1-depletion rescues CtIP and alt-EJ in G₀-phase cells. CtIP activation in G₀-phase cells also requires CDK-dependent phosphorylation by any available CDK but is restricted to CDK4/6 at the early stages of the normal cell cycle. We suggest that suppression of mutagenic alt-EJ in G₀-phase is a mechanism by which cells of higher eukaryotes maintain genomic stability in a large fraction of non-cycling cells in their organisms. CA Iliakis

This thesis contribute towards the understanding of labor economics and applied econometrics; the thesis is made up of three chapters.The first chapter explores the causal effect of parents’ social capital on the intergenerational occupational inertia in addition to individuals’ labor market outcomes. A new data extract was constructed by re-weighting and combining the Panel Study of Income Dynamics (PSID) and the Survey of Income and Program Participation (SIPP) to correct the selection biases induced by children’s endogenous moving behaviors post-graduation. By exploiting the recent technological revolution and the resulting changes in occupational skill compositions measured by Dictionary of Titles (DOT) and its successor O*NET, it became possible to isolate the effect of inherited social capital from inherited human capital through a regression discontinuity design. Besides, a correction of the selection bias induced by the social capital advantage through children’s occupational switching patterns after the first jobs was made. The results indicate that around 30% of individuals choose the same occupation as their parents for their first job; such people rely more on their parents’ social connections in job hunting. Also, they enjoy a positive wage premium of about 5% of the percentile ranks of annual labor income for entry-level jobs but this positive effect fades away in the long-run.The second chapter studies the estimation and inference of nonlinear econometric model when the economic variables are contained in different datasets. We show that the unknown structural parameters of interest can be possibly uniquely identified if there are some common conditioning variables in different datasets. The identification result is constructive, which enables us to estimate the unknown parameters based on a simple minimum distance (MD) estimator. We study the asymptotic properties of the MD estimator and provide inference procedure. A simple model specification test on the key identification conditions is also provided.The third chapter provides an application example of the method developed in the second chapter. It is a long-standing problem in the empirical research that the economic variables are contained in different datasets. One well-accepted solution to this problem is the imputation method, which serves as a crucial step in the seminal work, Blundell, Pistaferri, and Preston (2008) studied the dynamic relationship between consumption and income, with consumption data from CEX and income data from PSID. In this chapter, we first prove that the imputation method is biased because they are significantly different from those based on true data, which is the newly available PSID from 1999 which includes both consumption and income data. Furthermore, we investigate the finite sample performance of our new method with this new PSID data and show that our method delivers comparable results with those based on the true data. We conclude that the imputation gives largely biased estimation compared to the real data results and the new estimator developed in Chapter 2 performs better.The three chapters share the same interests in the long-lasting question that how we can deal with the situation in which the economic variables or the study population is contained in different datasets. The first chapter starts off from the simplest scenario that the data setis complete in terms of variables but biased in terms of representativeness. The other two chapters deal with the other more difficult and more usual case that the data set is incomplete in terms of economic variables. We not only contribute methodologically by providinga new estimator but also implement the method in an important application case and discuss the implications.

The current view of the involvement of PI3-kinases in checkpoint responses after DNA damage is that ATM is the key regulator of G₁-, S-or G₂-phase checkpoints, that ATR is only partly involved in the regulation of S-and G₂-phase checkpoints and that DNA-PKcs is not involved in checkpoint regulation. However, further analysis of the contributions of these kinases to checkpoint responses in cells exposed to ionizing radiation (IR) recently uncovered striking integrations and interplays among ATM, ATR and DNA-PKcs that adapt not only to the phase of the cell cycle in which cells are irradiated, but also to the load of DNA double-strand breaks (DSBs), presumably to optimize their processing. Specifically, we found that low IR doses in G₂-phase cells activate a G₂-checkpoint that is regulated by epistatically coupled ATM and ATR. Thus, inhibition of either kinase suppresses almost fully its activation. At high IR doses, the epistatic ATM/ATR coupling relaxes, yielding to a cooperative regulation. Thus, single-kinase inhibition suppresses partly, and only combined inhibition suppresses fully G₂-checkpoint activation. Interestingly, DNA-PKcs integrates with ATM/ATR in G₂-checkpoint control, but functions in its recovery in a dose-independent manner. Strikingly, irradiation during S-phase activates, independently of dose, an exclusively ATR-dependent G₂ checkpoint. Here, ATM couples with DNA-PKcs to regulate checkpoint recovery. In the present work, we extend these studies and investigate organization and functions of these PI3-kinases in the activation of the G₁ checkpoint in cells irradiated either in the G₀ or G₁ phase. We report that ATM is the sole regulator of the G₁ checkpoint after exposure to low IR doses. At high IR doses, ATM remains dominant, but contributions from ATR also become detectable and are associated with limited ATM/ATR-dependent end resection at DSBs. Under these conditions, only combined ATM + ATR inhibition fully abrogates checkpoint and resection. Contributions of DNA-PKcs and CHK2 to the regulation of the G₁ checkpoint are not obvious in these experiments and may be masked by the endpoint employed for checkpoint analysis and perturbations in normal progression through the cell cycle of cells exposed to DNA-PKcs inhibitors. The results broaden our understanding of organization throughout the cell cycle and adaptation with increasing IR dose of the ATM/ATR/DNA-PKcs module to regulate checkpoint responses. They emphasize notable similarities and distinct differences between G₁-, G₂-and S-phase checkpoint regulation that may guide DSB processing decisions. CA Iliakis

OA Förderung 2022 The load of DNA double-strand breaks (DSBs) induced in the genome of higher eukaryotes by different doses of ionizing radiation (IR) is a key determinant of DSB repair pathway choice, with homologous recombination (HR) and ATR substantially gaining ground at doses below 0.5 Gy. Increased resection and HR engagement with decreasing DSB-load generate a conundrum in a classical non-homologous end-joining (c-NHEJ)-dominated cell and suggest a mechanism adaptively facilitating resection. We report that ablation of DNA-PKcs causes hyper-resection, implicating DNA-PK in the underpinning mechanism. However, hyper-resection in DNA-PKcs-deficient cells can also be an indirect consequence of their c-NHEJ defect. Here, we report that all tested DNA-PKcs mutants show hyper-resection, while mutants with defects in all other factors of c-NHEJ fail to do so. This result rules out the model of c-NHEJ versus HR competition and the passive shift from c-NHEJ to HR as the causes of the increased resection and suggests the integration of DNA-PKcs into resection regulation. We develop a model, compatible with the results of others, which integrates DNA-PKcs into resection regulation and HR for a subset of DSBs. For these DSBs, we propose that the kinase remains at the break site, rather than the commonly assumed autophosphorylation-mediated removal from DNA ends.