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Telomere binding protein Stn1 forms the CST (Cdc13/CTC1-STN1-TEN1) complex in budding yeast and mammals. Likewise, fission yeast Stn1 and Ten1 form a complex indispensable for telomere protection. We have previously reported that stn1-1, a high-temperature sensitive mutant, rapidly loses telomere DNA at the restrictive temperature due to frequent failure of replication fork progression at telomeres and subtelomeres, both containing repetitive sequences. It is unclear, however, whether Stn1 is required for maintaining other repetitive DNAs such as ribosomal DNA. In this study, we have demonstrated that stn1-1 cells, even when grown at the permissive temperature, exhibited dynamic rearrangements in the telomere-proximal regions of subtelomere and ribosomal DNA repeats. Furthermore, Rad52 and γH2A accumulation was observed at ribosomal DNA repeats in the stn1-1 mutant. The phenotypes exhibited by the stn1-1 allele were largely suppressed in the absence of Reb1, a replication fork barrier-forming protein, suggesting that Stn1 is involved in the maintenance of the arrested replication forks. Collectively, we propose that Stn1 maintains the stability of repetitive DNAs at subtelomeres and rDNA regions.

We introduce a goal-aware extension of responsibility-sensitive safety (RSS), a recent methodology for rule-based safety guarantee for automated driving systems (ADS). Making RSS rules guarantee goal achievement -- in addition to collision avoidance as in the original RSS -- requires complex planning over long sequences of manoeuvres. To deal with the complexity, we introduce a compositional reasoning framework based on program logic, in which one can systematically develop RSS rules for smaller subscenarios and combine them to obtain RSS rules for bigger scenarios. As the basis of the framework, we introduce a program logic dFHL that accommodates continuous dynamics and safety conditions. Our framework presents a dFHL-based workflow for deriving goal-aware RSS rules; we discuss its software support, too. We conducted experimental evaluation using RSS rules in a safety architecture. Its results show that goal-aware RSS is indeed effective in realising both collision avoidance and goal achievement., 33 pages, 18 figures, 1 table. Accepted for publication in IEEE Transactions on Intelligent Vehicles

A common approach in testing automated and autonomous driving systems (ADS) consists in running the ADS in a simulator where driving and environmental conditions are specified in terms of scenarios. An important aspect in ADS testing is to cover different driving situations in which the autonomous car must perform different types of maneuvers. In this paper, we consider the path planner of our industry partner; the path planner is responsible for deciding the path that must be followed by the autonomous car. A path is characterized by the driving characteristics (as forward acceleration, lateral acceleration, curvature, and so on) that are needed, at each time point, to implement it. For different driving characteristics, a good test suite should contain a scenario for which the path planner chooses a path that requires the application of the selected driving characteristics for a non-negligible period of time: this means that the characteristics are relevant in that path. With such a test suite, engineers can observe the different types of decision taken by the path planner, and so possibly better assess its correctness. In the paper, we introduce the notion of patterns of driving characteristics, to characterize their interaction (i.e., simultaneous or not) and measure their duration. Exploiting this definition, we propose two search-based approaches (for single and pairs of driving characteristics) to find scenarios in which such patterns occur and their duration is maximized. Experimental results show that the approaches are effective in finding scenarios for which the path planner generates paths where the different driving characteristics occur in terms of the specified pattern.

Recent years have witnessed substantial progress in understanding tumor heterogeneity and the process of tumor progression; however, the entire process of the transition of tumors from a benign to metastatic state remains poorly understood. In the present study, we performed a prospective cancer genome-sequencing analysis by employing an experimental carcinogenesis mouse model of squamous cell carcinoma to systematically understand the evolutionary process of tumors. We surgically collected a part of a lesion of each tumor and followed the progression of these tumors in vivo over time. Comparative time-series analysis of the genomes of tumors with different fates, i.e., those that eventually metastasized and regressed, suggested that these tumors acquired and inherited different mutations. These findings suggest that despite the occurrence of an intra-tumor selection event for malignant alteration during the transformation from early- to late-stage papilloma, the fate determination of tumors might be determined at an even earlier stage.

The CST complex is a phylogenetically conserved protein complex consisting of CTC1/Cdc13, Stn1 and Ten1 that protects telomeres on linear chromosomes. Deletion of the fission yeast homologs stn1 and ten1 results in complete telomere loss; however, the precise function of Stn1 is still largely unknown. Here, we have isolated a high-temperature sensitive stn1 allele (termed stn1-1). stn1-1 cells abruptly lost telomeric sequence almost completely at the restrictive temperature. The loss of chromosomal DNA happened without gradual telomere shortening, and extended to 30 kb from the ends of chromosomes. We found transient and modest single-stranded G-strand exposure, but did not find any evidence of checkpoint activation in stn1-1 at the restrictive temperature. When we probed neutral-neutral 2D gels for subtelomere regions, we found no Y-arc-shaped replication intermediates in cycling cells. We conclude that the loss of telomere and subtelomere DNAs in stn1-1 cells at the restrictive temperature is caused by very frequent replication fork collapses specifically in subtelomere regions. Furthermore, we identified two independent suppressor mutants of the high-temperature sensitivity of stn1-1: a multi-copy form of pmt3 and a deletion of rif1. Collectively, we propose that fission yeast Stn1 primarily safeguards the semi-conservative DNA replication at telomeres and subtelomeres.

Genomic rearrangements (gross chromosomal rearrangements, GCRs) threatens genome integrity and cause cell death or tumor formation. At the terminus of linear chromosomes, a telomere-binding protein complex, called shelterin, ensures chromosome stability by preventing chromosome end-to-end fusions and regulating telomere length homeostasis. As such, shelterin-mediated telomere functions play a pivotal role in suppressing GCR formation. However, it remains unclear whether the shelterin proteins play any direct role in inhibiting GCR at non-telomeric regions. Here, we have established a GCR assay for the first time in fission yeast and measured GCR rates in various mutants. We found that fission yeast cells lacking shelterin components Taz1 or Rap1 (mammalian TRF1/2 or RAP1 homologues, respectively) showed higher GCR rates compared to wild-type, accumulating large chromosome deletions. Genetic dissection of Rap1 revealed that Rap1 contributes to inhibiting GCRs via two independent pathways. The N-terminal BRCT-domain promotes faithful DSB repair, as determined by I-SceI-mediated DSB-induction experiments; moreover, association with Poz1 mediated by the central Poz1-binding domain regulates telomerase accessibility to DSBs, leading to suppression of de novo telomere additions. Our data highlight unappreciated functions of the shelterin components Taz1 and Rap1 in maintaining genome stability, specifically by preventing non-telomeric GCRs., Author summary Tips of chromosomes, telomeres, are bound and protected by a telomere-binding protein complex called shelterin. Most previous studies focused on shelterin’s telomere-specific role, and its general role in genome maintenance has not been explored extensively. In this study, we first set up an assay measuring the spontaneous formation rate per cell division of gross chromosomal rearrangements (GCRs) in fission yeast. We found that the rate of GCRs is elevated in mutants defective for shelterin components Taz1 or Rap1. Detailed genetic experiments revealed unexpectedly that Taz1 and Rap1 have a novel role in repairing DNA double-strand breaks (DSBs) and suppressing GCRs at non-telomeric regions. Given that shelterin components are conserved between fission yeast and humans, future studies are warranted to test whether shelterin dysfunction leads to genome-wide GCRs, which are frequently observed in cancers.

More and more software practitioners are tackling towards industrial applications of artificial intelligence (AI) systems, especially those based on machine learning (ML). However, many of existing principles and approaches to traditional systems do not work effectively for the system behavior obtained by training not by logical design. In addition, unique kinds of requirements are emerging such as fairness and explainability. To provide clear guidance to understand and tackle these difficulties, we present an analysis on what quality concepts we should evaluate for AI systems. We base our discussion on ISO/IEC 25000 series, known as SQuaRE, and identify how it should be adapted for the unique nature of ML and $\textit{Ethics guidelines for trustworthy AI}$ from European Commission. We thus provide holistic insights for quality of AI systems by incorporating the ML nature and AI ethics to the traditional software quality concepts.

International audience; From smart buildings to medical devices to smart nations, software systems increasingly integrate computation, networking, and interaction with the physical environment. These systems are known as Cyber-Physical Systems (CPS). While these systems open new opportunities to deliver improved quality of life for people and reinvigorate computing, their engineering is a difficult problem given the level of heterogeneity and dynamism they exhibit. While progress has been made, we argue that complexity is now at a level such that existing approaches need a major rethink to define principles and associated techniques for CPS. In this paper, we identify research challenges when modelling, analysing and engineering CPS. We focus on three key topics: theoretical foundations of CPS, self-adaptation methods for CPS, and exemplars of CPS serving as a research vehicle shared by a larger community. For each topic, we present an overview and suggest future research directions, thereby focusing on selected challenges. This paper is one of the results of the Shonan Seminar 118 on Modelling and Analysing Resilient Cyber-Physical Systems, which took place in December 2018.

Telomeres maintain the integrity of chromosome ends and telomere length is an important marker of aging. The epidemiological studies suggested that many types of stress including psychosocial stress decrease telomere length. However, it remains unknown how various stresses induce telomere shortening. Here, we report that the stress-responsive transcription factor ATF7 mediates TNF-α–induced telomere shortening. ATF7 and telomerase, an enzyme that elongates telomeres, are localized on telomeres via interactions with the Ku complex. In response to TNF-α, which is induced by various stresses including psychological stress, ATF7 was phosphorylated by p38, leading to the release of ATF7 and telomerase from telomeres. Thus, a decrease of ATF7 and telomerase on telomeres in response to stress causes telomere shortening, as observed in ATF7-deficient mice. These findings give credence to the idea that various types of stress might shorten telomere.