Intrinsic PARG inhibitor sensitivity is mimicked by TIMELESS haploinsufficiency and rescued by nucleoside supplementation.

A subset of cancer cells are intrinsically sensitive to inhibitors targeting PARG, the poly(ADP-ribose) glycohydrolase that degrades PAR chains. Sensitivity is accompanied by persistent DNA replication stress, and can be induced by inhibition of TIMELESS, a replisome accelerator. However, the nature of the vulnerability responsible for intrinsic sensitivity remains undetermined. To understand PARG activity dependency, we analysed Timeless model systems and intrinsically sensitive ovarian cancer cells. We show that nucleoside supplementation rescues all phenotypes associated with PARG inhibitor sensitivity, including replisome speed and fork stalling, S-phase completion and mitotic entry, proliferation dynamics and clonogenic potential. Importantly nucleoside supplementation restores PARG inhibitor resistance despite the continued presence of PAR chains, indicating that sensitivity does not correlate with PAR levels. In addition, we show that inhibition of thymidylate synthase, an enzyme required for dNTP homeostasis, induces PARG-dependency. Together, these observations suggest that PARG inhibitor sensitivity reflects an inability to control replisome speed and/or maintain helicase-polymerase coupling in response to nucleotide imbalances.

Introduction of one-view tomosynthesis in population-based mammography screening: Impact on detection rate, interval cancer rate and false-positive rate.

OBJECTIVE: To assess performance endpoints of a combination of digital breast tomosynthesis (DBT) and full-field digital mammography (FFDM) compared with FFDM only in breast cancer screening. MATERIALS AND METHODS: This was a prospective population-based screening study, including eligible (50-69 years) women attending the Capital Region Mammography Screening Program in Denmark. All attending women were offered FFDM. A subgroup was consecutively allocated to a screening room with DBT. All FFDM and DBT underwent independent double reading, and all women were followed up for 2 years after screening date or until next screening date, whichever came first. RESULTS: 6353 DBT + FFDM and 395 835 FFDM were included in the analysis and were undertaken in 196 267 women in the period from 1 November 2012 to 12 December 2018. Addition of DBT increased sensitivity: 89.9% (95% confidence interval (CI): 81.0-95.5) for DBT + FFDM and 70.1% (95% CI: 68.6-71.6) for FFDM only, p < 0.001. Specificity remained similar: 98.2% (95% CI: 97.9-98.5) for DBT + FFDM and 98.3% (95% CI: 98.2-98.3) for FFDM only, p = 0.9. Screen-detected cancer rate increased statistically significantly: 11.18/1000 for DBT + FFDM and 6.49/1000 for FFDM only, p < 0.001. False-positive rate was unchanged: 1.75% for DBT + FFDM and 1.73% for FFDM only, p = 0.9. Positive predictive value for recall was 39.0% (95% CI: 31.9-46.5) for DBT + FFDM and 27.3% (95% CI: 26.4-28.2), for FFDM only, p < 0.0005. The interval cancer rate decreased: 1.26/1000 for DBT + FFDM and 2.76/1000 for FFDM only, p = 0.02. CONCLUSION: DBT + FFDM yielded a statistically significant increase in cancer detection and program sensitivity.

Magnetic fields reveal signatures of triplet-pair multi-exciton photoluminescence in singlet fission.

The photophysical processes of singlet fission and triplet fusion have numerous emerging applications. They involve the separation of a photo-generated singlet exciton into two dark triplet excitons and the fusion of two dark triplet excitons into an emissive singlet exciton, respectively. The role of the excimer state and the nature of the triplet-pair state in these processes have been a matter of contention. Here we analyse the room temperature time-resolved emission of a neat liquid singlet fission chromophore and show that it exhibits three spectral components: two that correspond to the bright singlet and excimer states and a third component that becomes more prominent during triplet fusion. This spectrum is enhanced by magnetic fields, confirming its origins in the recombination of weakly coupled triplet pairs. It is thus attributed to a strongly coupled triplet pair state. These observations unite the view that there is an emissive intermediate in singlet fission and triplet fusion, distinct from the broad, unstructured excimer emission.

ADP-ribosylome analysis reveals homogeneous DNA-damage-induced serine ADP-ribosylation across wild-type and BRCA-mutant breast cancer cell lines.

ADP-ribosylation (ADPr) signaling plays a crucial role in DNA damage response. Inhibitors against the main enzyme catalyzing ADPr after DNA damage, poly(ADP-ribose) polymerase 1 (PARP1), are used to treat patients with breast cancer harboring BRCA1/2 mutations. However, resistance to PARP inhibitors (PARPi) is a major obstacle in treating patients. To understand the role of ADPr in PARPi sensitivity, we use liquid chromatography-tandem mass spectrometry (LC-MS/MS) to analyze ADPr in six breast cancer cell lines exhibiting different PARPi sensitivities. We identify 1,632 sites on 777 proteins across all cell lines, primarily on serine residues, with site-specific overlap of targeted residues across DNA-damage-related proteins across all cell lines, demonstrating high conservation of serine ADPr-signaling networks upon DNA damage. Furthermore, we observe site-specific differences in ADPr intensities in PARPi-sensitive BRCA mutants and unique ADPr sites in PARPi-resistant BRCA-mutant HCC1937 cells, which have low poly(ADP-ribose) glycohydrolase (PARG) levels and longer ADPr chains on PARP1.

PARP14 and PARP9/DTX3L regulate interferon-induced ADP-ribosylation.

PARP-catalysed ADP-ribosylation (ADPr) is important in regulating various cellular pathways. Until recently, PARP-dependent mono-ADP-ribosylation has been poorly understood due to the lack of sensitive detection methods. Here, we utilised an improved antibody to detect mono-ADP-ribosylation. We visualised endogenous interferon (IFN)-induced ADP-ribosylation and show that PARP14 is a major enzyme responsible for this modification. Fittingly, this signalling is reversed by the macrodomain from SARS-CoV-2 (Mac1), providing a possible mechanism by which Mac1 counteracts the activity of antiviral PARPs. Our data also elucidate a major role of PARP9 and its binding partner, the E3 ubiquitin ligase DTX3L, in regulating PARP14 activity through protein-protein interactions and by the hydrolytic activity of PARP9 macrodomain 1. Finally, we also present the first visualisation of ADPr-dependent ubiquitylation in the IFN response. These approaches should further advance our understanding of IFN-induced ADPr and ubiquitin signalling processes and could shed light on how different pathogens avoid such defence pathways.

Super Resolution Ultrasound Imaging using the Erythrocytes: I: Density Images.

A new approach for vascular super resolution imaging using the erythrocytes as targets (SURE imaging) is described and investigated. SURE imaging does not require fragile contrast agent bubbles, making it possible to use the maximum allowable mechanical index for ultrasound scanning for an increased penetration depth. A synthetic aperture ultrasound sequence was employed with 12 virtual sources using a 10 MHz GE L8-18i-D linear array hockey stick probe. The axial resolution was 1.20λ,(185.0µm) and the lateral resolution was 1.50λ,(231.3µm). Field IIpro simulations were conducted on 12.5 µm radius vessel pairs with varying separations. A vessel pair with a separation of 70 µm could be resolved, indicating a SURE image resolution below half a wavelength. A Verasonics research scanner was used for the in vivo experiments to scan the kidneys of Sprague-Dawley rats for up to 46 s to visualize their microvasculature by processing from 0.1 up to 45 s of data for SURE imaging, and for 46.8 s for super resolution (SR) imaging with a SonoVue contrast agent. Afterward, the renal vasculature was filled with the ex vivo micro-CT contrast agent Microfil, excised, and scanned in a micro-CT scanner at both a 22.6 µm voxel size for 11 hours, and for 20 hours in a 5 µm voxel size for validating the SURE images. Comparing the SURE and micro-CT images revealed that vessels with a diameter of 28 µm, five times smaller than the ultrasound wavelength, could be detected, and the dense grid of microvessels in the full kidney was shown for scan times between 1 to 10 s. The vessel structure in the cortex was also similar for the SURE and SR images. Fourier ring correlation indicated a resolution capability of 29 µm. SURE images are acquired in seconds rather than minutes without any patient preparation or contrast injection, making the method translatable to clinical use.

Super Resolution Ultrasound Imaging Using the Erythrocytes: II: Velocity Images.

Super resolution ultrasound imaging using the erythrocytes (SURE) has recently been introduced. The method uses erythrocytes as targets instead of fragile microbubbles (MBs). The abundance of erythrocyte scatterers makes it possible to acquire SURE data in just a few seconds compared to several minutes in ultrasound localization microscopy (ULM) using MBs. A high number of scatterers can reduce the acquisition time, however, the tracking of uncorrelated and high-density scatterers is quite challenging. This paper hypothesizes that it is possible to detect and track erythrocytes as targets to obtain vascular flow images. A SURE tracking pipeline is used with modules for beamforming, recursive synthetic aperture imaging, motion estimation, echo canceling, peak detection, and recursive nearest neighbor tracker. The SURE tracking pipeline is capable of distinguishing the flow direction and separating tubes of a simulated Field II phantom with 125 to 25 µm wall-to-wall tube distances, as well as a 3D-printed hydrogel micro-flow phantom with 100 to 60 µm wall-to-wall channel distances. The comparison of an in-vivo SURE scan of a Sprague-Dawley rat kidney with ULM and micro-CT scans with voxel sizes of 26.5µm and 5µm demonstrated consistent findings. A microvascular structure composed of 16 vessels exhibited similarities across all imaging modalities. The flow direction and velocity profiles in the SURE scan were found to be concordant with those from ULM.

RNF212B E3 ligase is essential for crossover designation and maturation during male and female meiosis in the mouse.

Meiosis, a reductional cell division, relies on precise initiation, maturation, and resolution of crossovers (COs) during prophase I to ensure the accurate segregation of homologous chromosomes during metaphase I. This process is regulated by the interplay of RING-E3 ligases such as RNF212 and HEI10 in mammals. In this study, we functionally characterized a recently identified RING-E3 ligase, RNF212B. RNF212B colocalizes and interacts with RNF212, forming foci along chromosomes from zygonema onward in a synapsis-dependent and DSB-independent manner. These consolidate into larger foci at maturing COs, colocalizing with HEI10, CNTD1, and MLH1 by late pachynema. Genetically, RNF212B foci formation depends on Rnf212 but not on Msh4, Hei10, and Cntd1, while the unloading of RNF212B at the end of pachynema is dependent on Hei10 and Cntd1. Mice lacking RNF212B, or expressing an inactive RNF212B protein, exhibit modest synapsis defects, a reduction in the localization of pro-CO factors (MSH4, TEX11, RPA, MZIP2) and absence of late CO-intermediates (MLH1). This loss of most COs by diakinesis results in mostly univalent chromosomes. Double mutants for Rnf212b and Rnf212 exhibit an identical phenotype to that of Rnf212b single mutants, while double heterozygous demonstrate a dosage-dependent reduction in CO number, indicating a functional interplay between paralogs. SUMOylome analysis of testes from Rnf212b mutants and pull-down analysis of Sumo- and Ubiquitin-tagged HeLa cells, suggest that RNF212B is an E3-ligase with Ubiquitin activity, serving as a crucial factor for CO maturation. Thus, RNF212 and RNF212B play vital, yet overlapping roles, in ensuring CO homeostasis through their distinct E3 ligase activities.

Retron-Eco1 assembles NAD+-hydrolyzing filaments that provide immunity against bacteriophages.

Retrons are toxin-antitoxin systems protecting bacteria against bacteriophages via abortive infection. The Retron-Eco1 antitoxin is formed by a reverse transcriptase (RT) and a non-coding RNA (ncRNA)/multi-copy single-stranded DNA (msDNA) hybrid that neutralizes an uncharacterized toxic effector. Yet, the molecular mechanisms underlying phage defense remain unknown. Here, we show that the N-glycosidase effector, which belongs to the STIR superfamily, hydrolyzes NAD+ during infection. Cryoelectron microscopy (cryo-EM) analysis shows that the msDNA stabilizes a filament that cages the effector in a low-activity state in which ADPr, a NAD+ hydrolysis product, is covalently linked to the catalytic E106 residue. Mutations shortening the msDNA induce filament disassembly and the effector's toxicity, underscoring the msDNA role in immunity. Furthermore, we discovered a phage-encoded Retron-Eco1 inhibitor (U56) that binds ADPr, highlighting the intricate interplay between retron systems and phage evolution. Our work outlines the structural basis of Retron-Eco1 defense, uncovering ADPr's pivotal role in immunity.

Singlet fission in TIPS-anthracene thin films.

Singlet fission is an exciton multiplication process that allows for the conversion of one singlet exciton into two triplet excitons. Organic semiconductors, such as acenes and their soluble bis(triisopropylsilylethynyl) (TIPS) substituted counterparts, have played a major role in elucidating the understanding of the underlying mechanisms of singlet fission. Despite this, one prominent member of the acene family that has received little experimental attention to date is TIPS-anthracene, even with computational studies suggesting potential high singlet fission yields in the solid state. Here, time-resolved spectroscopic and magneto-photoluminescence measurements were performed on spin-cast films of TIPS-anthracene, showing evidence for singlet fission. A singlet fission yield of 19% (out of 200%) is estimated from transient absorption spectroscopy. Kinetic modeling of the magnetic field effect on photoluminescence suggests that fast rates of triplet dissociation lead to a low magnetic photoluminescence effect and that non-radiative decay of both the S1 and 1(TT) states is the cause for the low triplet yield.

Prognostic relevance of radiological findings on early postoperative MRI for 187 consecutive glioblastoma patients receiving standard therapy.

Several prognostic factors are known to influence survival for patients treated with IDH-wildtype glioblastoma, but unknown factors may remain. We aimed to investigate the prognostic implications of early postoperative MRI findings. A total of 187 glioblastoma patients treated with standard therapy were consecutively included. Patients either underwent a biopsy or surgery followed by an early postoperative MRI. Progression-free survival (PFS) and overall survival (OS) were analysed for known prognostic factors and MRI-derived candidate factors: resection status as defined by the response assessment in neuro-oncology (RANO)-working group (no contrast-enhancing residual tumour, non-measurable contrast-enhancing residual tumour, or measurable contrast-enhancing residual tumour) with biopsy as reference, contrast enhancement patterns (no enhancement, thin linear, thick linear, diffuse, nodular), and the presence of distant tumours. In the multivariate analysis, patients with no contrast-enhancing residual tumour or non-measurable contrast-enhancing residual tumour on the early postoperative MRI displayed a significantly improved progression-free survival compared with patients receiving only a biopsy. Only patients with non-measurable contrast-enhancing residual tumour showed improved overall survival in the multivariate analysis. Contrast enhancement patterns were not associated with survival. The presence of distant tumours was significantly associated with both poor progression-free survival and overall survival and should be considered incorporated into prognostic models.

DNAJC9 prevents CENP-A mislocalization and chromosomal instability by maintaining the fidelity of histone supply chains.

The centromeric histone H3 variant CENP-A is overexpressed in many cancers. The mislocalization of CENP-A to noncentromeric regions contributes to chromosomal instability (CIN), a hallmark of cancer. However, pathways that promote or prevent CENP-A mislocalization remain poorly defined. Here, we performed a genome-wide RNAi screen for regulators of CENP-A localization which identified DNAJC9, a J-domain protein implicated in histone H3-H4 protein folding, as a factor restricting CENP-A mislocalization. Cells lacking DNAJC9 exhibit mislocalization of CENP-A throughout the genome, and CIN phenotypes. Global interactome analysis showed that DNAJC9 depletion promotes the interaction of CENP-A with the DNA-replication-associated histone chaperone MCM2. CENP-A mislocalization upon DNAJC9 depletion was dependent on MCM2, defining MCM2 as a driver of CENP-A deposition at ectopic sites when H3-H4 supply chains are disrupted. Cells depleted for histone H3.3, also exhibit CENP-A mislocalization. In summary, we have defined novel factors that prevent mislocalization of CENP-A, and demonstrated that the integrity of H3-H4 supply chains regulated by histone chaperones such as DNAJC9 restrict CENP-A mislocalization and CIN.

Concerted SUMO-targeted ubiquitin ligase activities of TOPORS and RNF4 are essential for stress management and cell proliferation.

Protein SUMOylation provides a principal driving force for cellular stress responses, including DNA-protein crosslink (DPC) repair and arsenic-induced PML body degradation. In this study, using genome-scale screens, we identified the human E3 ligase TOPORS as a key effector of SUMO-dependent DPC resolution. We demonstrate that TOPORS promotes DPC repair by functioning as a SUMO-targeted ubiquitin ligase (STUbL), combining ubiquitin ligase activity through its RING domain with poly-SUMO binding via SUMO-interacting motifs, analogous to the STUbL RNF4. Mechanistically, TOPORS is a SUMO1-selective STUbL that complements RNF4 in generating complex ubiquitin landscapes on SUMOylated targets, including DPCs and PML, stimulating efficient p97/VCP unfoldase recruitment and proteasomal degradation. Combined loss of TOPORS and RNF4 is synthetic lethal even in unstressed cells, involving defective clearance of SUMOylated proteins from chromatin accompanied by cell cycle arrest and apoptosis. Our findings establish TOPORS as a STUbL whose parallel action with RNF4 defines a general mechanistic principle in crucial cellular processes governed by direct SUMO-ubiquitin crosstalk.

Simulation-based education in ultrasound - diagnostic and interventional abdominal focus. / Simulationsbasierte Ausbildung im Ultraschall ­ Fokus auf der diagnostischen und interventionellen Abdomen-Sonografie.

Simulation-based training (SBT) is increasingly acknowledged worldwide and has become a popular tool for ultrasound education. Ultrasound simulation involves the use of technology and software to create a virtual training setting. Simulation-based training allows healthcare professionals to learn, practice, and improve their ultrasound imaging skills in a safe learning-based environment. SBT can provide a realistic and focused learning experience that creates a deep and immersive understanding of the complexity of ultrasound, including enhancing knowledge and confidence in specific areas of interest. Abdominal ultrasound simulation is a tool to increase patient safety and can be a cost-efficient training method. In this paper, we provide an overview of various types of abdominal ultrasound simulators, and the benefits, and challenges of SBT. We also provide examples of how to develop SBT programs and learning strategies including mastery learning. In conclusion, the growing demand for medical imaging increases the need for healthcare professionals to start using ultrasound simulators in order to keep up with the rising standards.

VCF1 is a p97/VCP cofactor promoting recognition of ubiquitylated p97-UFD1-NPL4 substrates.

The hexameric AAA+ ATPase p97/VCP functions as an essential mediator of ubiquitin-dependent cellular processes, extracting ubiquitylated proteins from macromolecular complexes or membranes by catalyzing their unfolding. p97 is directed to ubiquitylated client proteins via multiple cofactors, most of which interact with the p97 N-domain. Here, we discover that FAM104A, a protein of unknown function also named VCF1 (VCP/p97 nuclear Cofactor Family member 1), acts as a p97 cofactor in human cells. Detailed structure-function studies reveal that VCF1 directly binds p97 via a conserved α-helical motif that recognizes the p97 N-domain with unusually high affinity, exceeding that of other cofactors. We show that VCF1 engages in joint p97 complex formation with the heterodimeric primary p97 cofactor UFD1-NPL4 and promotes p97-UFD1-NPL4-dependent proteasomal degradation of ubiquitylated substrates in cells. Mechanistically, VCF1 indirectly stimulates UFD1-NPL4 interactions with ubiquitin conjugates via its binding to p97 but has no intrinsic affinity for ubiquitin. Collectively, our findings establish VCF1 as an unconventional p97 cofactor that promotes p97-dependent protein turnover by facilitating p97-UFD1-NPL4 recruitment to ubiquitylated targets.

Biomedical laboratory scientists and technicians in digital pathology - Is there a need for professional development?

Objective: Digital pathology (DP) is moving into Danish pathology departments at high pace. Conventionally, biomedical laboratory scientists (BLS) and technicians have prepared tissue sections for light microscopy, but workflow alterations are required for the new digital era with whole slide imaging (WSI); digitally assisted image analysis (DAIA) and artificial intelligence (AI). We aim to explore the role of BLS in DP and assess a potential need for professional development. Methods: We investigated the roles of BLS in the new digital era through qualitative interviews at Danish Pathology Departments in 2019/2020 before DP implementation (supported by a questionnaire); and in 2022 after DP implementation. Additionally, senior lecturers from three Danish University Colleges reported on how DP was integrated into the 2023 bachelor's degree educational curricula for BLS students. Results: At some Danish pathology departments, BLS were involved in the implementation process of DP and their greatest concerns were lack of physical laboratory requirements (69%) and implementation strategies (63%). BLS were generally positive towards working with DP, however, some expressed concern about extended working hours for scanning. Work-task transfers from pathologists were generally greeted positively from both management and pathologists; however, at follow-up interviews after DP implementation, job transfers had not been effectuated. At Danish university colleges, DP had been integrated systematically in the curricula for BLS students, especially WSI. Conclusion: Involving BLS in DP implementation and development may benefit the process, as BLS have a hands-on workflow perspective with a focus on quality assurance. Several new work opportunities for BLS may occur with DP including WSI, DAIA and AI, and therefore new qualifications are warranted, which must be considered in future undergraduate programmes for BLS students or postgraduate programmes for BLS.

Surgical stress response in robot-assisted versus laparoscopic surgery for colon cancer (SIRIRALS): randomized clinical trial.

BACKGROUND: Evidence for the routine use of robotic technology and its impact on short-term outcomes in colon cancer surgery is lacking. The aim of this study was to compare the surgically induced systemic stress response and clinical and patient-reported outcomes for patients undergoing robot-assisted or laparoscopic colon cancer surgery. METHODS: In this double-blinded superiority RCT completed between August 2021 and March 2023, patients with stage 1-3 colon cancer were randomized in a 1 : 1 ratio to undergo either robot-assisted or laparoscopic colon cancer surgery. The primary outcome was changes in the systemic stress response, characterized by C-reactive protein expression in the first three postoperative days. Secondary outcomes were intraoperative and postoperative complications and patient-reported outcomes. The latter included quality of recovery-15 and pain intensity using a visual analogue scale. RESULTS: In total, 128 patients were screened for potential inclusion in this study; 50 patients (25 in the robot-assisted group and 25 in the laparoscopic group) were included in the final follow-up and analysis. The postoperative C-reactive protein response was higher on the first postoperative day in the laparoscopic group (mean difference = 19.88 mg/l, 95% c.i. 3.89-35.86; P = 0.045). No statistically significant differences were noted for C-reactive protein expression on the second and third postoperative days. CONCLUSION: Adopting robot-assisted surgery for stage 1-3 colon cancer is associated with a reduction in the surgical stress response. REGISTRATION NUMBER: NCT04687384 (http://www.clinicaltrials.gov).

A quantitative and site-specific atlas of the citrullinome reveals widespread existence of citrullination and insights into PADI4 substrates.

Despite the importance of citrullination in physiology and disease, global identification of citrullinated proteins, and the precise targeted sites, has remained challenging. Here we employed quantitative-mass-spectrometry-based proteomics to generate a comprehensive atlas of citrullination sites within the HL60 leukemia cell line following differentiation into neutrophil-like cells. We identified 14,056 citrullination sites within 4,008 proteins and quantified their regulation upon inhibition of the citrullinating enzyme PADI4. With this resource, we provide quantitative and site-specific information on thousands of PADI4 substrates, including signature histone marks and transcriptional regulators. Additionally, using peptide microarrays, we demonstrate the potential clinical relevance of certain identified sites, through distinct reactivities of antibodies contained in synovial fluid from anti-CCP-positive and anti-CCP-negative people with rheumatoid arthritis. Collectively, we describe the human citrullinome at a systems-wide level, provide a resource for understanding citrullination at the mechanistic level and link the identified targeted sites to rheumatoid arthritis.

Thermal Disorder-Induced Strain and Carrier Localization Activate Reverse Halide Segregation.

The reversal of halide ions is studied under various conditions. However, the underlying mechanism of heat-induced reversal remains unclear. This work finds that dynamic disorder-induced localization of self-trapped polarons and thermal disorder-induced strain (TDIS) can be co-acting drivers of reverse segregation. Localization of polarons results in an order of magnitude decrease in excess carrier density (polaron population), causing a reduced impact of the light-induced strain (LIS - responsible for segregation) on the perovskite framework. Meanwhile, exposing the lattice to TDIS exceeding the LIS can eliminate the photoexcitation-induced strain gradient, as thermal fluctuations of the lattice can mask the LIS strain. Under continuous 0.1 W cm⁻2 illumination (upon segregation), the strain disorder is estimated to be 0.14%, while at 80 °C under dark conditions, the strain is 0.23%. However, in situ heating of the segregated film to 80 °C under continuous illumination (upon reversal) increases the total strain disorder to 0.25%, where TDIS is likely to have a dominant contribution. Therefore, the contribution of entropy to the system's free energy is likely to dominate, respectively. Various temperature-dependent in situ measurements and simulations further support the results. These findings highlight the importance of strain homogenization for designing stable perovskites under real-world operating conditions.

Profiling ubiquitin signalling with UBIMAX reveals DNA damage- and SCFß-Trcp1-dependent ubiquitylation of the actin-organizing protein Dbn1.

Ubiquitin widely modifies proteins, thereby regulating most cellular functions. The complexity of ubiquitin signalling necessitates unbiased methods enabling global detection of dynamic protein ubiquitylation. Here, we describe UBIMAX (UBiquitin target Identification by Mass spectrometry in Xenopus egg extracts), which enriches ubiquitin-conjugated proteins and quantifies regulation of protein ubiquitylation under precise and adaptable conditions. We benchmark UBIMAX by investigating DNA double-strand break-responsive ubiquitylation events, identifying previously known targets and revealing the actin-organizing protein Dbn1 as a major target of DNA damage-induced ubiquitylation. We find that Dbn1 is targeted for proteasomal degradation by the SCFß-Trcp1 ubiquitin ligase, in a conserved mechanism driven by ATM-mediated phosphorylation of a previously uncharacterized ß-Trcp1 degron containing an SQ motif. We further show that this degron is sufficient to induce DNA damage-dependent protein degradation of a model substrate. Collectively, we demonstrate UBIMAX's ability to identify targets of stimulus-regulated ubiquitylation and reveal an SCFß-Trcp1-mediated ubiquitylation mechanism controlled directly by the apical DNA damage response kinases.