Checkpoint-Dependent and -Independent Roles of Swi3 in Replication Fork Recovery and Sister Chromatid Cohesion in Fission Yeast

Multiple genome maintenance processes are coordinated at the replication fork to preserve genomic integrity. How eukaryotic cells accomplish such a coordination is unknown. Swi1 and Swi3 form the replication fork protection complex and are involved in various processes including stabilization of replication forks, activation of the Cds1 checkpoint kinase and establishment of sister chromatid cohesion in fission yeast. However, the mechanisms by which the Swi1–Swi3 complex achieves and coordinates these tasks are not well understood. Here, we describe the identification of separation-of-function mutants of Swi3, aimed at dissecting the molecular pathways that require Swi1–Swi3. Unlike swi3 deletion mutants, the separation-of-function mutants were not sensitive to agents that stall replication forks. However, they were highly sensitive to camptothecin that induces replication fork breakage. In addition, these mutants were defective in replication fork regeneration and sister chromatid cohesion. Interestingly, unlike swi3-deleted cell, the separation-of-functions mutants were proficient in the activation of the replication checkpoint, but their fork regeneration defects were more severe than those of checkpoint mutants including cds1Δ, chk1Δ and rad3Δ. These results suggest that, while Swi3 mediates full activation of the replication checkpoint in response to stalled replication forks, Swi3 activates a checkpoint-independent pathway to facilitate recovery of collapsed replication forks and the establishment of sister chromatid cohesion. Thus, our separation-of-function alleles provide new insight into understanding the multiple roles of Swi1-Swi3 in fork protection during DNA replication, and into understanding how replication forks are maintained in response to different genotoxic agents

State-Dependent Fragility Curves for Aftershock Seismic Risk Assessment of Japanese Steel Frames

Probabilistic seismic risk assessment of civil infrastructures has been at tracting attention in Japan, especially after recent mega - earthquakes with a long - lasting series of aftershocks capable of accumulating building damage; e.g., the 2011 Tohoku earthquake. To this aim , it is valuable to be able to assess the failure probabil ity of a particular structure and its evolution in time due to sequential earthquake events, which may cause a difficulty for stakeholders to perform consistent decision making to warrant business continuity. This kind of risk analysis may require state - de pendent fragility curves, which in the study were developed for a Japanese steel frame . To construct the curves , a numerical model of a t hree - story steel moment - resisting frame was first constructed and calibrated acc ording to the results of shake table te st s for a typical Japanese steel structure . This model was subsequently transformed in an equivalent single degree of freedom (ESDOF) system , based on the results of the nonlinear static (pushover) analysis. The probabilistic damage model was then construc ted via nonlinear dynamic analys i s of the ESDOF system . The spectral acceleration of the elastic period of the ESDOF system was selected as the ground motion intensity measure while the drift angle was selected as response measure. All the records used in the analysis were selected from the Japanese strong - motion network, K - N et. Finally , the state - dependent fragility curves were developed for five levels of damage: As - New (AN), Immediate Occupancy (IO), Life Safety (LS), Collapse Prevention (CP) and Failure ( F ). The limit state value for each damage state (DS) was set in compliance with the results of the shake table tes t s . After computing the damage state probability due to the mainshock, t he time - variant aftershock risk of the steel structure was quantifie d integrating the developed state - dependent fragility curves with the seismic hazard, followin g a Markov chain model already available in the literature, which makes use of aftershock probabilistic seismic hazard analysis (APSHA) . Hazard was computed assum ing that the structure was located in Osaka, a site that may be affected by a mega - earthquake at the Nankai Trough subduction - zone . In particular, in the considered exercise, the most probable damage state due to the considered mainshock scenario was found to be IO, followed, in probability terms, by LS, F, AN, and CP. Given the probability distribution of the mainshock - induced damage, the daily evolution of aftershock damage was computed, and it was found that the most likely DS after two m onths since the mainshock was still IO followed by F, LS, CP and AN