Proteostasis failure and cellular senescence in long-term cultured postmitotic rat neurons

Cellular senescence, a stress‐induced irreversible cell cycle arrest, has been defined for mitotic cells and is implicated in aging of replicative tissues. Age‐related functional decline in the brain is often attributed to a failure of protein homeostasis (proteostasis), largely in postmitotic neurons, which accordingly is a process distinct by definition from senescence. It is nevertheless possible that proteostasis failure and cellular senescence have overlapping molecular mechanisms. Here, we identify postmitotic cellular senescence as an adaptive stress response to proteostasis failure. Primary rat hippocampal neurons in long‐term cultures show molecular changes indicative of both senescence (senescence‐associated β‐galactosidase, p16, and loss of lamin B1) and proteostasis failure relevant to Alzheimer's disease. In addition, we demonstrate that the senescent neurons exhibit resistance to stress. Importantly, treatment of the cultures with an mTOR antagonist, protein synthesis inhibitor, or chemical compound that reduces the amount of protein aggregates relieved the proteotoxic stresses as well as the appearance of senescence markers. Our data propose mechanistic insights into the pathophysiological brain aging by establishing senescence as a primary cell‐autonomous neuroprotective response

spRap1 and spRif1, recruited to telomeres by Taz1, are essential for telomere function in fission yeast

AbstractTelomeres are essential for genome integrity. scRap1 (S. cerevisiae Rap1) directly binds to telomeric DNA [1–3] and regulates telomere length and telomere position effect (TPE) [4–6] by recruiting two different groups of proteins to its RCT (Rap1 C-terminal) domain [7]. The first group, Rif1 and Rif2, regulates telomere length [8, 9]. The second group, Sir3 and Sir4 [10], is involved in heterochromatin formation [11–13]. On the other hand, human TRF1 and TRF2, as well as their fission yeast homolog, Taz1, directly bind to telomeric DNA [14–16] and negatively regulate telomere length [16–20]. Taz1 also plays important roles in TPE and meiosis [16, 20, 21]. Human Rap1, the ortholog of scRap1, negatively regulates telomere length and appears to be recruited to telomeres by interacting with TRF2 [7]. Here, we describe two novel fission yeast proteins, spRap1 (S. pombe Rap1) and spRif1 (S. pombe Rif1), which are orthologous to scRap1 and scRif1, respectively. spRap1 and spRif1 are independently recruited to telomeres by interacting with Taz1. The rap1 mutant is severely defective in telomere length control, TPE, and telomere clustering toward the spindle pole body (SPB) at the premeiotic horsetail stage, indicating that spRap1 has critical roles in these telomere functions. The rif1 mutant also shows some defects in telomere length control and meiosis. Our results indicate that Taz1 provides binding sites for telomere regulators, spRap1 and spRif1, which perform the essential telomere functions. This study establishes the similarity of telomere organization in fission yeast and humans

The CST complex facilitates cell survival under oxidative genotoxic stress

Genomic DNA is constantly exposed to a variety of genotoxic stresses, and it is crucial for organisms to be equipped with mechanisms for repairing the damaged genome. Previously, it was demonstrated that the mammalian CST (CTC1-STN1-TEN1) complex, which was originally identified as a single-stranded DNA-binding trimeric protein complex essential for telomere maintenance, is required for survival in response to hydroxyurea (HU), which induces DNA replication fork stalling. It is still unclear, however, how the CST complex is involved in the repair of diverse types of DNA damage induced by oxidizing agents such as H₂O₂. STN1 knockdown (KD) sensitized HeLa cells to high doses of H₂O₂. While H₂O₂ induced DNA strand breaks throughout the cell cycle, STN1 KD cells were as resistant as control cells to H₂O₂ treatment when challenged in the G1 phase of the cell cycle, but they were sensitive when exposed to H₂O₂ in S/G2/M phase. STN1 KD cells showed a failure of DNA synthesis and RAD51 foci formation upon H₂O₂ treatment. Chemical inhibition of RAD51 in shSTN1 cells did not exacerbate the sensitivity to H₂O₂, implying that the CST complex and RAD51 act in the same pathway. Collectively, our results suggest that the CST complex is required for maintaining genomic stability in response to oxidative DNA damage, possibly through RAD51-dependent DNA repair/protection mechanisms

Loss of linker histone H1 in cellular senescence

Cellular senescence is a tumor-suppressing mechanism that is accompanied by characteristic chromatin condensation called senescence-associated heterochromatic foci (SAHFs). We found that individual SAHFs originate from individual chromosomes. SAHFs do not show alterations of posttranslational modifications of core histones that mark condensed chromatin in mitotic chromosomes, apoptotic chromatin, or transcriptionally inactive heterochromatin. Remarkably, SAHF-positive senescent cells lose linker histone H1 and exhibit increased levels of chromatin-bound high mobility group A2 (HMGA2). The expression of N-terminally enhanced green fluorescent protein (EGFP)–tagged histone H1 induces premature senescence phenotypes, including increased levels of phosphorylated p53, p21, and hypophosphorylated Rb, and a decrease in the chromatin-bound endogenous histone H1 level but not in p16 level accumulation or SAHF formation. However, the simultaneous ectopic expression of hemagglutinin-tagged HMGA2 and N-terminally EGFP-tagged histone H1 leads to significant SAHF formation (P < 0.001). It is known that histone H1 and HMG proteins compete for a common binding site, the linker DNA. These results suggest that SAHFs are a novel type of chromatin condensation involving alterations in linker DNA–binding proteins

Using a Variational Autoencoder to Learn Valid Search Spaces of Safely Monitored Autonomous Robots for Last-Mile Delivery

The use of autonomous robots for delivery of goods to customers is an exciting new way to provide a reliable and sustainable service. However, in the real world, autonomous robots still require human supervision for safety reasons. We tackle the real-world problem of optimizing autonomous robot timings to maximize deliveries, while ensuring that there are never too many robots running simultaneously so that they can be monitored safely. We assess the use of a recent hybrid machine-learning-optimization approach COIL (constrained optimization in learned latent space) and compare it with a baseline genetic algorithm for the purposes of exploring variations of this problem. We also investigate new methods for improving the speed and efficiency of COIL. We show that only COIL can find valid solutions where appropriate numbers of robots run simultaneously for all problem variations tested. We also show that when COIL has learned its latent representation, it can optimize 10% faster than the GA, making it a good choice for daily re-optimization of robots where delivery requests for each day are allocated to robots while maintaining safe numbers of robots running at once