Aggregation of the Tau protein into fibrils defines progression of neurodegenerative diseases, including Alzheimer’s Disease. The molecular basis for potentially toxic reactions of Tau aggregates is poorly understood. Here we show that π-stacking by Arginine side-chains drives protein binding to Tau fibrils. We mapped an aggregation-dependent interaction pattern of Tau. Fibrils recruit specifically aberrant interactors characterised by intrinsically disordered regions of atypical sequence features. Arginine residues are key to initiate these aberrant interactions. Crucial for scavenging is the guanidinium group of its side chain, not its charge, indicating a key role of π-stacking chemistry for driving aberrant fibril interactions. Remarkably, despite the non-hydrophobic interaction mode, the molecular chaperone Hsp90 can modulate aberrant fibril binding. Together, our data present a molecular mode of action for derailment of protein-protein interaction by neurotoxic fibrils.

Oncotarget | https://doi.org/10.18632/oncotarget.27367 Boxiao Ding, Alexander Haidurov, Ayesha Chawla, Anita Parmigiani, Gerarda van de Kamp, Alexandra Dalina, Fang Yuan, Jun Hee Lee, Peter M. Chumakov, Steven R....

Radiotherapy induces tumor cell killing by generating DNA double strand breaks (DSBs). The effectiveness of radiotherapy is significantly influenced by the repair of DSBs, which counteracts this lethal effect. Current investigations are focused on determining whether non-homologous end joining (NHEJ) or homologous recombination is the predominant repair pathway following proton and photon radiation. In this study, we examined the response of FaDu cells, a head and neck squamous cell carcinoma model, to spread-out Bragg peak (SOBP) proton and photon radiation combined with mild hyperthermia (42 °C for one hour) to induce homologous recombination deficiency or NHEJ inhibition by AZD7648. Hyperthermia resulted in stronger radiosensitization after proton radiation (SR = 1.53) compared to photon radiation (SR = 1.32). Conversely, NHEJ inhibition did not produce a significant differential effect between photon and proton radiation. This indicates a greater reliance on homologous recombination following proton radiation compared to photon radiation. We found that the number of DSBs formed after photon versus proton irradiation is comparable. Interestingly, the homologous recombination protein Rad51 accumulated more frequently at DSBs following proton irradiation than photon irradiation. These findings support the hypothesis that cells rely more on homologous recombination to repair proton-induced DNA damage compared to photon-induced DNA damage. As clinically applied hyperthermia enhances the therapeutic effect of photon irradiation by, among other factors, inducing homologous recombination deficiency, our results suggests that hyperthermia could be more effective in combination with proton irradiation than photon irradiation.

DNA double strand breaks (DSBs) are critical for the efficacy of radiotherapy as they lead to cell death if not repaired. DSBs caused by ionizing radiation (IR) initiate histone modifications and accumulate DNA repair proteins, including 53BP1, which forms distinct foci at damage sites and serves as a marker for DSBs. DSB repair primarily occurs through Non-Homologous End Joining (NHEJ) and Homologous Recombination (HR). NHEJ directly ligates DNA ends, employing proteins such as DNA-PK_cs, while HR, involving proteins such as Rad54, uses a sister chromatid template for accurate repair and functions in the S and G2 phases of the cell cycle. Both pathways are crucial, as illustrated by the IR sensitivity in cells lacking DNA-PK_cs or Rad54. We generated mouse embryonic stem (mES) cells which are knockout (KO) for DNA-PK_cs and Rad54 to explore the combined role of HR and NHEJ in DSB repair. We found that cells lacking both DNA-PK_cs and Rad54 are hypersensitive to X-ray radiation, coinciding with impaired 53BP1 focus resolution and a more persistent G2 phase cell cycle block. Additionally, mES cells deficient in DNA-PK_cs or both DNA-PK_cs and Rad54 exhibit an increased nuclear size approximately 18–24 h post-irradiation. To further explore the role of Rad54 in the absence of DNA-PK_cs, we generated DNA-PK_cs KO mES cells expressing GFP-tagged wild-type (WT) or ATPase-defective Rad54 to track the Rad54 foci over time post-irradiation. Cells lacking DNA-PK_cs and expressing ATPase-defective Rad54 exhibited a similar phenotypic response to IR as those lacking both DNA-PK_cs and Rad54. Despite a strong G2 phase arrest, live-cell imaging showed these cells eventually progress through mitosis, forming micronuclei. Additionally, mES cells lacking DNA-PK_cs showed increased Rad54 foci over time post-irradiation, indicating an enhanced reliance on HR for DSB repair without DNA-PK_cs. Our findings underscore the essential roles of HR and NHEJ in maintaining genomic stability post-IR in mES cells. The interplay between these pathways is crucial for effective DSB repair and cell cycle progression, highlighting potential targets for enhancing radiotherapy outcomes.

High linear energy transfer (LET) radiation is more harmful than low LET radiation because it deposits energy in a concentrated manner, resulting in clustered DNA damage (CDD). Double strand breaks (DSBs) are among the most damaging types of DNA damage, and if not repaired, they may trigger cell death. DSBs can be repaired through three mechanisms: non-homologous end joining (NHEJ), homologous recombination (HR), and theta-mediated end joining (TMEJ). This study aimed to assess how these pathways contribute to repairing DSBs induced by low LET X-ray and proton radiation, and high LET alpha-particle radiation. We used mouse embryonic stem (mES) cells lacking key repair proteins to examine clonogenic survival and the formation and resolution of 53BP1 foci, a DNA damage marker, after exposure to X-ray, proton, and alpha-particle radiation. The results showed increased sensitivity to X-ray and proton radiation in NHEJ, HR, and TMEJ repair-deficient cell lines compared to wild-type cells, with similar trends for both radiation types. Notably, Rad54-deficient cells showed slower resolution of 53BP1 foci after proton exposure, indicating increased reliance on HR for repairing proton-induced DSBs. Clonogenic survival assays revealed a relative biological effectiveness (RBE) of 4.6-5.8 for alpha-particles compared to protons and X-rays, confirming that alpha-particles are more effective at causing cell death. Our findings suggest that TMEJ is important for repairing DSBs caused by alpha-particles. This study highlights differences in repairing low LET versus high LET DNA damage, offering new insights for radiation biology and therapeutic strategies.

DNA crosslink-inducing drugs are widely used in clinical settings for treatment of solid tumors. Double strand breaks (DSBs) that arise during interstrand crosslink (ICL) repair are crucial determinants of the therapeutic response, as they lead to cell death if not repaired. DSBs can be repaired through non-homologous end joining (NHEJ), theta-mediated end joining (TMEJ), and homologous recombination (HR). HR is considered a major pathway for repairing DSBs induced during ICL repair. In this study, we examine the roles of NHEJ, TMEJ, and HR in ICL repair using mouse embryonic stem (mES) cells. We show that DNA-PK_cs-deficient mES cells are resistant to the crosslinkers mitomycin C (MMC), cisplatin and carboplatin, contrasting with the increased sensitivity observed in mES cells lacking Rad54. Furthermore, the absence of DNA-PK_cs correlates with enhanced HR activity, as evidenced by an increased number of Rad54 foci following MMC treatment. The combined knock-outof DNA-PK_cs and Rad54 reduces sensitivity to crosslinkers compared to cells lacking only Rad54, suggesting the involvement of another DSB repair pathway besides HR. We found that TMEJ deficiency can sensitize cells to cisplatin, particularly in those lacking NHEJ and HR repair. This suggests that TMEJ contributes to cell survival following cisplatin treatment. In clinical settings, higher PRKDC expression correlates with poorer survival, while elevated RAD54L and POLQ expression correlates with better survival in cisplatin-treated cervical and head and neck cancers. These findings reflect the opposing roles of NHEJ versus HR and TMEJ in replication-associated DSB repair, as observed in vitro.