Multi-method analysis of microplastic distribution by flood frequency and local topography in Rhine floodplains.

Rivers are important transport pathways for microplastics into the ocean, but they can also be potential sinks due to microplastic deposition in the sediments of the river bed and adjacent floodplains. In particular, floods can (re)mobilise microplastics from sediments and floodplains, (re)deposit and relocate them depending on the floodplain topography. The knowledge about fluvial microplastic input to floodplains, their spatial distribution and their fate in floodplain soils is limited. To investigate this topic, we sampled soil at a depth of 5-20 cm along three transects in three different Rhine floodplains. We analysed the soil samples in tandem with pyrolysis GC/MS and ATR- & µ-FPA-FTIR for their microplastic abundance and mass concentrations. To study the influence of flood frequency on the microplastic abundance in the three floodplains, we fitted a hydrodynamic flood model (MIKE 21, DHI, Hørsholm, Denmark) and related the results to the respective spatial microplastic distribution. We found similar microplastic distribution patterns in each floodplain. The highest microplastic abundance (8516-70,124 microplastics kg-1) and mass concentration (46.2-141.6 mg kg-1) were consistently found in the farthest transects from the Rhine in a topographical depression. This microplastic distribution pattern is detectable with both, pyrolysis GC/MS and FTIR. The strongest correlation between the results of both methods was found for small, abundant microplastic particles. Our results suggest that the spatial distribution of microplastics in floodplains is related to the combination of flood frequency and local topography, that ought to be explicitly considered in future studies conducted in floodplains. Finally, our results indicate that pyrolysis GC/MS and FTIR data are comparable under certain conditions, which may help in the decision for the analytical method and sampling design in future studies.

Dual truncation of tau by caspase-2 accelerates its CHIP-mediated degradation.

Intraneuronal aggregates of the microtubule binding protein Tau are a hallmark of different neurodegenerative diseases including Alzheimer's disease (AD). In these aggregates, Tau is modified by posttranslational modifications such as phosphorylation as well as by proteolytic cleavage. Here we identify a novel Tau cleavage site at aspartate 65 (D65) that is specific for caspase-2. In addition, we show that the previously described cleavage site at D421 is also efficiently processed by caspase-2, and both sites are cleaved in human brain samples. Caspase-2-generated Tau fragments show increased aggregation potential in vitro, but do not accumulate in vivo after AAV-mediated overexpression in mouse hippocampus. Interestingly, we observe that steady-state protein levels of caspase-2 generated Tau fragments are low in our in vivo model despite strong RNA expression, suggesting efficient clearance. Consistent with this hypothesis, we find that caspase-2 cleavage significantly improves the recognition of Tau by the ubiquitin E3 ligase CHIP, leading to increased ubiquitination and faster degradation of Tau fragments. Taken together our data thus suggest that CHIP-induced ubiquitination is of particular importance for the clearance of caspase-2 generated Tau fragments in vitro and in vivo.

Protein import motor complex reacts to mitochondrial misfolding by reducing protein import and activating mitophagy.

Mitophagy is essential to maintain mitochondrial function and prevent diseases. It activates upon mitochondria depolarization, which causes PINK1 stabilization on the mitochondrial outer membrane. Strikingly, a number of conditions, including mitochondrial protein misfolding, can induce mitophagy without a loss in membrane potential. The underlying molecular details remain unclear. Here, we report that a loss of mitochondrial protein import, mediated by the pre-sequence translocase-associated motor complex PAM, is sufficient to induce mitophagy in polarized mitochondria. A genome-wide CRISPR/Cas9 screen for mitophagy inducers identifies components of the PAM complex. Protein import defects are able to induce mitophagy without a need for depolarization. Upon mitochondrial protein misfolding, PAM dissociates from the import machinery resulting in decreased protein import and mitophagy induction. Our findings extend the current mitophagy model to explain mitophagy induction upon conditions that do not affect membrane polarization, such as mitochondrial protein misfolding.

Flooding frequency and floodplain topography determine abundance of microplastics in an alluvial Rhine soil.

Rivers are major pathways for the transport of microplastics towards the oceans, and many studies focus on microplastic abundance in fluvial ecosystems. Although flooding strongly affects transport of microplastics, knowledge about the potential input via floodwaters, spatial distribution, and fate of microplastics in adjacent floodplains remains very limited. In this study, we suggest that local topography and flood frequency could influence the abundance of microplastics in floodplains. Based on this concept, we took soil samples in a Rhine River floodplain in two different depths (0-5 cm and 5-20 cm) along three transects with increasing distance to the river and analysed the abundance of microplastics via FTIR spectroscopy. Flood frequency of the transects was estimated by a combination of hydrodynamic modelling with MIKE 21 (DHI, Hørsholm Denmark) and analysis of time series of water levels. Microplastic abundance per kg dry soil varied between 25,502 to 51,119 particles in the top 5 cm and 25,616 to 84,824 particles in the deeper soil (5-20 cm). The results of our study indicate that local topography and resulting flooding patterns are responsible for the amount of microplastics found at the respective transect. Differences in soil properties, vegetation cover and signs of earthworm activity in the soil profile seem to be related to microplastic migration and accumulation in the deeper soil. The interdisciplinary approach we used in our work can be applied to other floodplains to elucidate the respective processes. This information is essentially important both for locating potential microplastic sinks for process-informed sampling designs and to identify areas of increased bioavailability of microplastics for proper ecological risk assessment.

Robust LC3B lipidation analysis by precisely adjusting autophagic flux.

Autophagic flux can be quantified based on the accumulation of lipidated LC3B in the presence of late-stage autophagy inhibitors. This method has been widely applied to identify novel compounds that activate autophagy. Here we scrutinize this approach and show that bafilomycin A1 (BafA) but not chloroquine is suitable for flux quantification due to the stimulating effect of chloroquine on non-canonical LC3B-lipidation. Significant autophagic flux increase by rapamycin could only be observed when combining it with BafA concentrations not affecting basal flux, a condition which created a bottleneck, rather than fully blocking autophagosome-lysosome fusion, concomitant with autophagy stimulation. When rapamycin was combined with saturating concentrations of BafA, no significant further increase of LC3B lipidation could be detected over the levels induced by the late-stage inhibitor. The large assay window obtained by this approach enables an effective discrimination of autophagy activators based on their cellular potency. To demonstrate the validity of this approach, we show that a novel inhibitor of the acetyltransferase EP300 activates autophagy in a mTORC1-dependent manner. We propose that the creation of a sensitized background rather than a full block of autophagosome progression is required to quantitatively capture changes in autophagic flux.

A Novel Approach for Quantifying the Pharmacological Activity of T-Cell Engagers Utilizing In Vitro Time Course Experiments and Streamlined Data Analysis.

CD3-bispecific antibodies are a new class of immunotherapeutic drugs against cancer. The pharmacological activity of CD3-bispecifics is typically assessed through in vitro assays of cancer cell lines co-cultured with human peripheral blood mononuclear cells (PBMCs). Assay results depend on experimental conditions such as incubation time and the effector-to-target cell ratio, which can hinder robust quantification of pharmacological activity. In order to overcome these limitations, we developed a new, holistic approach for quantification of the in vitro dose-response relationship. Our experimental design integrates a time-independent analysis of the dose-response across different time points as an alternative to the static, "snap-shot" analysis based on a single time point commonly used in dose-response assays. We show that the potency values derived from static in vitro experiments depend on the incubation time, which leads to inconsistent results across multiple assays and compounds. We compared the potency values from the time-independent analysis with a model-based approach. We find comparably accurate potency estimates from the model-based and time-independent analyses and that the time-independent analysis provides a robust quantification of pharmacological activity. This approach may allow for an improved head-to-head comparison of different compounds and test systems and may prove useful for supporting first-in-human dose selection.

On which common ground to build? Transferable knowledge across cases in transdisciplinary sustainability research.

To support societal problem solving, transdisciplinary research (TDR) uses knowledge co-production focusing on relevance and validity in a studied case and its particular social-ecological context. In the first instance, the resulting situated knowledge seems to be restricted to these single cases. However, if some of the knowledge generated in TDR could be used in other research projects, this would imply that there is a body of knowledge representing this special type of research. This study used a qualitative approach based on the methodology of grounded theory to empirically examine what knowledge is considered transferable to other cases, if any. 30 leaders of 12 Swiss-based TDR projects in the field of sustainable development were interviewed, representing both academia and practice. The transferable knowledge we found consists of the following: (1) Transdisciplinary principles, (2) transdisciplinary approaches, (3) systematic procedures, (4) product formats, (5) experiential know-how, (6) framings and (7) insights, data and information. The discussion of TDR has predominantly been focusing on transdisciplinary principles and approaches. In order to take knowledge co-production in TDR beyond an unmanageable field of case studies, more efforts in developing and critically discussing transferable knowledge of the other classes are needed, foremost systematic procedures, product formats and framings.

A Maternal-Effect Toxin Affects Epithelial Differentiation and Tissue Mechanics in Caenorhabditis elegans.

Selfish genetic elements that act as post-segregation distorters cause lethality in non-carrier individuals after fertilization. Two post-segregation distorters have been previously identified in Caenorhabditis elegans, the peel-1/zeel-1 and the sup-35/pha-1 elements. These elements seem to act as modification-rescue systems, also called toxin/antidote pairs. Here we show that the maternal-effect toxin/zygotic antidote pair sup-35/pha-1 is required for proper expression of apical junction (AJ) components in epithelia and that sup-35 toxicity increases when pathways that establish and maintain basal epithelial characteristics, die-1, elt-1, lin-26, and vab-10, are compromised. We demonstrate that pha-1(e2123) embryos, which lack the antidote, are defective in epidermal morphogenesis and frequently fail to elongate. Moreover, seam cells are frequently misshaped and mispositioned and cell bond tension is reduced in pha-1(e2123) embryos, suggesting altered tissue material properties in the epidermis. Several aspects of this phenotype can also be induced in wild-type embryos by exerting mechanical stress through uniaxial loading. Seam cell shape, tissue mechanics, and elongation can be restored in pha-1(e2123) embryos if expression of the AJ molecule DLG-1/Discs large is reduced. Thus, our experiments suggest that maternal-effect toxicity disrupts proper development of the epidermis which involves distinct transcriptional regulators and AJ components.

Design and Development of a Chemical Probe for Pseudokinase Ca2+/calmodulin-Dependent Ser/Thr Kinase.

CASK (Ca2+/calmodulin-dependent Ser/Thr kinase) is a member of the MAGUK (membrane-associated guanylate kinase) family that functions as neurexin kinases with roles implicated in neuronal synapses and trafficking. The lack of a canonical DFG motif, which is altered to GFG in CASK, led to the classification as a pseudokinase. However, functional studies revealed that CASK can still phosphorylate substrates in the absence of divalent metals. CASK dysfunction has been linked to many diseases, including colorectal cancer, Parkinson's disease, and X-linked mental retardation, suggesting CASK as a potential drug target. Here, we exploited structure-based design for the development of highly potent and selective CASK inhibitors based on 2,4-diaminopyrimidine-5-carboxamides targeting an unusual pocket created by the GFG motif. The presented inhibitor design offers a more general strategy for the development of pseudokinase ligands that harbor unusual sequence motifs. It also provides a first chemical probe for studying the biological roles of CASK.

Right ventricular function and vasoactive peptides for early prediction of bronchopulmonary dysplasia.

BACKGROUND: To assess the prognostic value of early echocardiographic indices of right ventricular function and vasoactive peptides for prediction of bronchopulmonary dysplasia (BPD) or death in very preterm infants. METHODS: Prospective study involving 294 very preterm infants (median [IQR] gestational age 28.4 [26.4-30.4] weeks, birth weight 1065 [800-1380] g), of whom 57 developed BPD (oxygen supplementation at 36 weeks postmenstrual age) and 10 died. Tricuspid annular plane systolic excursion (TAPSE), right ventricular index of myocardial performance (RIMP), plasma concentrations of mid-regional pro-atrial natriuretic peptide (MR-proANP) and C-terminal pro-endothelin-1 (CT-proET1) were measured on day 7 of life. RESULTS: RIMP was significantly increased (median [IQR] 0.3 [0.23-0.38] vs 0.22 [0.15-0.29]), TAPSE decreased (median [IQR] 5.0 [5.0-6.0] vs 6.0 [5.4-7.0] mm), MR-proANP increased (median [IQR] 784 [540-936] vs 353 [247-625] pmol/L), and CT-proET1 increased (median [IQR] 249 [190-345] vs 199 [158-284] pmol/L) in infants who developed BPD or died, as compared to controls. All variables showed significant but weak correlations with each other (rS -0.182 to 0.359) and predicted BPD/death with similar accuracy (areas under receiver operator characteristic curves 0.62 to 0.77). Multiple regression revealed only RIMP and birth weight as independent predictors of BPD or death. CONCLUSIONS: Vasoactive peptide concentrations and echocardiographic assessment employing standardized measures, notably RIMP, on day 7 of life are useful to identify preterm infants at increased risk for BPD or death.

Development of a Selective Dual Discoidin Domain Receptor (DDR)/p38 Kinase Chemical Probe.

Discoidin domain receptors 1 and 2 (DDR1/2) play a central role in fibrotic disorders, such as renal and pulmonary fibrosis, atherosclerosis, and various forms of cancer. Potent and selective inhibitors, so-called chemical probe compounds, have been developed to study DDR1/2 kinase signaling. However, these inhibitors showed undesired activity on other kinases such as the tyrosine protein kinase receptor TIE or tropomyosin receptor kinases, which are related to angiogenesis and neuronal toxicity. In this study, we optimized our recently published p38 mitogen-activated protein kinase inhibitor 7 toward a potent and cell-active dual DDR/p38 chemical probe and developed a structurally related negative control. The structure-guided design approach used provided insights into the P-loop folding process of p38 and how targeting of non-conserved amino acids modulates inhibitor selectivity. The developed and comprehensively characterized DDR/p38 probe, 30 (SR-302), is a valuable tool for studying the role of DDR kinase in normal physiology and in disease development.

Differential Thresholds of Proteasome Activation Reveal Two Separable Mechanisms of Sensory Organ Polarization in C. elegans.

Cephalization is a major innovation of animal evolution and implies a synchronization of nervous system, mouth, and foregut polarization to align alimentary tract and sensomotoric system for effective foraging. However, the underlying integration of morphogenetic programs is poorly understood. Here, we show that invagination of neuroectoderm through de novo polarization and apical constriction creates the mouth opening in the Caenorhabditis elegans embryo. Simultaneously, all 18 juxta-oral sensory organ dendritic tips become symmetrically positioned around the mouth: While the two bilaterally symmetric amphid sensilla endings are towed to the mouth opening, labial and cephalic sensilla become positioned independently. Dendrite towing is enabled by the pre-polarized sensory amphid pores intercalating into the leading edge of the anteriorly migrating epidermal sheet, while apical constriction-mediated cell-cell re-arrangements mediate positioning of all other sensory organs. These two processes can be separated by gradual inactivation of the 26S proteasome activator, RPN-6.1. Moreover, RPN-6.1 also shows a dose-dependent requirement for maintenance of coordinated apical polarization of other organs with apical lumen, the pharynx, and the intestine. Thus, our data unveil integration of morphogenetic programs during the coordination of alimentary tract and sensory organ formation and suggest that this process requires tight control of ubiquitin-dependent protein degradation.

Health care provided to recent asylum-seeking and non-asylum-seeking pediatric patients in 2016 and 2017 at a Swiss tertiary hospital - a retrospective study.

BACKGROUND: Asylum-seeking children represent an increasing and vulnerable group of patients whose health needs are largely unmet. Data on the health care provision to asylum-seeking children in European contexts is scarce. In this study we compare the health care provided to recent asylum-seeking and non-asylum-seeking children at a Swiss tertiary hospital. METHODS: We performed a cross-sectional retrospective study in a pediatric tertiary care hospital in Basel, Switzerland. All patients and visits from January 2016 to December 2017 were identified, using administrative and medical electronic health records. The asylum-seeking status was systematically assessed and the patients were allocated accordingly in the two study groups. RESULTS: A total of 202,316 visits by 55,789 patients were included, of which asylum-seeking patients accounted for 1674 (1%) visits by 439 (1%) individuals. The emergency department recorded the highest number of visits in both groups with a lower proportion in asylum-seeking compared to non-asylum-seeking children: 19% (317/1674) and 32% (64,315/200,642) respectively. The median number of visits per patient was 1 (IQR 1-2) in the asylum-seeking and 2 (IQR 1-4) in the non-asylum-seeking children. Hospital admissions were more common in asylum-seeking compared to non-asylum-seeking patients with 11% (184/1674) and 7% (14,692/200,642). Frequent visits (> 15 visits per patient) accounted for 48% (807/1674) of total visits in asylum-seeking and 25% (49,886/200,642) of total visits in non-asylum-seeking patients. CONCLUSIONS: Hospital visits by asylum-seeking children represented a small proportion of all visits. The emergency department had the highest number of visits in all patients but was less frequently used by asylum-seeking children. Frequent care suggests that asylum-seeking patients also present with more complex diseases. Further studies are needed, focusing on asylum-seeking children with medical complexity.

Serum Neurofilament Levels in Children With Febrile Seizures and in Controls.

OBJECTIVE: Neuroaxonal damage is reflected by serum neurofilament light chain (sNfL) values in a variety of acute and degenerative diseases of the brain. The aim of this study was to investigate the impact of febrile and epileptic seizures on sNfL, serum copeptin, and prolactin levels in children compared with children with febrile infections without convulsions. METHODS: A prospective cross-sectional study was performed in children aging 6 months to 5 years presenting with fever (controls, n = 61), febrile seizures (FS, n = 78), or epileptic seizures (ES, n = 16) at our emergency department. sNfL, copeptin, and prolactin were measured within a few hours after the event in addition to standard clinical, neurophysiological, and laboratory assessment. All children were followed up for at least 1 year after presentation concerning recurrent seizures. RESULTS: Serum copeptin values were on average 4.1-fold higher in FS and 3.2-fold higher in ES compared with controls (both p < 0.01). Serum prolactin values were on average 1.3-fold higher in FS compared with controls ( p < 0.01) and without difference between ES and controls. There was no significant difference of mean sNfL values (95% CI) between all three groups, FS 21.7 pg/ml (19.6-23.9), ES 17.7 pg/ml (13.8-21.6), and controls 23.4 pg/ml (19.2-27.4). In multivariable analysis, age was the most important predictor of sNfL, followed by sex and C reactive protein. Neither the duration of seizures nor the time elapsed from seizure onset to blood sampling had an impact on sNfL. None of the three biomarkers were related to recurrent seizures. SIGNIFICANCE: Serum neurofilament light is not elevated during short recovery time after FS when compared with children presenting febrile infections without seizures. We demonstrate an age-dependent decrease of sNfL from early childhood until school age. In contrast to sNfL levels, copeptin and prolactin serum levels are elevated after FS.

Reasons for admission in asylum-seeking and non-asylum-seeking patients in a paediatric tertiary care centre.

BACKGROUND: In the last decade an increasing number of asylum-seeking children arrived in Europe and local healthcare systems have been challenged to adapt to their health needs. The aim of this study was to compare the spectrum of disease and management of asylum-seeking and non-asylum-seeking children requiring hospital admission. METHODS: This was a retrospective cohort study including health data from recently arrived asylum-seeking and non-asylum-seeking children admitted between January 2016 and December 2017. Data were collected using electronic administrative and medical records. RESULTS: Of 11,794 admissions of 9407 patients, 149 (1%) were asylum-seeking and 11,645 (99%) from non-asylum-seeking children. In asylum-seeking children the median age was 4 years (interquartile range [IQR] 0–13) with 61% males and in non-asylum-seeking children 4 years (IQR 0–11) years with 56% males. Respiratory infections accounted for 17–19% of admissions in both groups. Rare infectious diseases were more frequent in asylum-seeking children (15 vs 7%; difference in proportions 0.08, 95% confidence interval [CI] 0.02–0.14; p <0.001,). Injuries were more frequent in non-asylum-seeking children (22 vs 13%; difference in proportions 0.09, 95% CI 0.04–0.014; p <0.01). Admissions for mental health disorders were infrequent but more common in asylum-seeking children (6 vs 3%; difference in proportions 0.03, 95% CI −0.01 – 0.07; p = 0.02) Prescription of analgesics was lower in asylum-seeking than non-asylum-seeking children (3.4 vs 6.5 accounting units per admission). Antibiotic prescription was comparable in both groups. CONCLUSION: Asylum-seeking children represent a small number of total admissions. Age distribution and main reason for admission being diseases of the respiratory system were comparable in asylum-seeking and non-asylum-seeking children. Rare infections and mental health disorders are important diseases in asylum-seeking children and require special attention and training of staff working with paediatric asylum seekers.

Cellular quality control by the ubiquitin-proteasome system and autophagy.

To achieve homeostasis, cells evolved dynamic and self-regulating quality control processes to adapt to new environmental conditions and to prevent prolonged damage. We discuss the importance of two major quality control systems responsible for degradation of proteins and organelles in eukaryotic cells: the ubiquitin-proteasome system (UPS) and autophagy. The UPS and autophagy form an interconnected quality control network where decision-making is self-organized on the basis of biophysical parameters (binding affinities, local concentrations, and avidity) and compartmentalization (through membranes, liquid-liquid phase separation, or the formation of aggregates). We highlight cellular quality control factors that delineate their differential deployment toward macromolecular complexes, liquid-liquid phase-separated subcellular structures, or membrane-bound organelles. Finally, we emphasize the need for continuous promotion of quantitative and mechanistic research into the roles of the UPS and autophagy in human pathophysiology.

Planar Asymmetries in the C. elegans Embryo Emerge by Differential Retention of aPARs at Cell-Cell Contacts.

Formation of the anteroposterior and dorsoventral body axis in Caenorhabditis elegans depends on cortical flows and advection of polarity determinants. The role of this patterning mechanism in tissue polarization after formation of cell-cell contacts is not fully understood. Here, we demonstrate that planar asymmetries are established during left-right symmetry breaking: Centripetal cortical flows asymmetrically and differentially advect anterior polarity determinants (aPARs) from contacts to the medial cortex, resulting in their unmixing from apical myosin. Contact localization and advection of PAR-6 requires balanced CDC-42 activation, while asymmetric retention and advection of PAR-3 can occur independently of PAR-6. Concurrent asymmetric retention of PAR-3, E-cadherin/HMR-1 and opposing retention of antagonistic CDC-42 and Wnt pathway components leads to planar asymmetries. The most obvious mark of planar asymmetry, retention of PAR-3 at a single cell-cell contact, is required for proper cytokinetic cell intercalation. Hence, our data uncover how planar polarity is established in a system without the canonical planar cell polarity pathway through planar asymmetric retention of aPARs.

Mechanical stress induces a scalable switch in cortical flow polarization during cytokinesis.

During animal development, cells need to sense and adapt to mechanical forces from their environment. Ultimately, these forces are transduced through the actomyosin cortex. How the cortex simultaneously responds to and creates forces during cytokinesis is not well understood. Here we show that, under mechanical stress, cortical actomyosin flow can switch polarization during cytokinesis in the C. elegans embryo. In unstressed embryos, longitudinal cortical flow contributes to contractile ring formation, while rotational cortical flow is additionally induced in uniaxially loaded embryos, i.e. embryos compressed between two plates. Rotational flow depends on astral microtubule signals and is required for the redistribution of the actomyosin cortex in loaded embryos. Rupture of longitudinally aligned cortical fibers during cortex rotation releases tension, initiates orthogonal longitudinal flow and, thereby, contributes to furrowing in loaded embryos. Moreover, actomyosin regulators involved in RhoA regulation, cortical polarity and chirality are all required for rotational flow, and become essential for cytokinesis under mechanical stress. In sum, our findings extend the current framework of mechanical stress response during cell division and show scaling of orthogonal cortical flows to the amount of mechanical stress.

Conservation of structure, function and inhibitor binding in UNC-51-like kinase 1 and 2 (ULK1/2).

Autophagy is essential for cellular homeostasis and when deregulated this survival mechanism has been associated with disease development. Inhibition of autophagy initiation by inhibiting the kinase ULK1 (Unc-51-like autophagy activating kinase 1) has been proposed as a potential cancer therapy. While inhibitors and crystal structures of ULK1 have been reported, little is known about the other closely related kinase ULK2 (Unc-51-like autophagy activating kinase 2). Here, we present the crystal structure of ULK2 in complex with ATP competitive inhibitors. Surprisingly, the ULK2 structure revealed a dimeric assembly reminiscent of dimeric arrangements of auto-activating kinases suggesting a role for this association in ULK activation. Screening of a kinase focused library of pre-clinical and clinical compounds revealed several potent ULK1/2 inhibitors and good correlation of inhibitor-binding behavior with both ULK kinases. Aurora A was identified as a major off-target of currently used ULK1 inhibitors. Autophagic flux assays demonstrated that this off-target activity by strongly inducing autophagy in different cellular systems conferred an additional layer of complexity in the interpretation of cellular data. The data presented here provide structural models and chemical starting points for the development of ULK1/2 dual inhibitors with improved selectivity for future exploitation of autophagy inhibition.