Biology subjects

Sattler, M.

Publications and source records attributed to Sattler, M..

At least 19 recordsLinked to original sources

bioRxiv2026

Direct interaction between miR-210-5p and HIF-1α regulates HIF-dependent transcription

Hypoxia-inducible factors (HIFs) coordinate cellular adaptation to oxygen deprivation, yet whether hypoxia-induced microRNAs directly regulate HIF-dependent transcription remains unknown. Here, we identify miR-210-5p as a nuclear hypoxamiR that directly binds HIF-1 and enhances HIF-dependent transcription. Hypoxia induced rapid, HIF-dependent nuclear accumulation of mature miR-210-5p across multiple cell types. Biophysical analyses demonstrated direct interaction between miR-210-5p and the HIF-1 bHLH-PAS domain, while mutational studies identified a conserved 5'motif required for HIF-1 binding but dispensable for repression of the canonical cytoplasmic target ISCU. Functionally, transcriptional activity closely correlated with HIF-1-binding affinity, as binding-deficient variants failed to activate HIF reporters or endogenous target genes. Although AGO2 contributed to hypoxic transcriptional responses, miR-210-5p interacted directly with HIF-1 independently of AGO2. In ischemic myocardium, nuclear enrichment of miR-210-5p and its proximity to HIF-1 support the physiological relevance of this mechanism, revealing a previously unrecognized RNA-mediated layer of HIF transcriptional regulation.

molecular biology↗

bioRxiv2026

Molecular basis of SPRTN activation by DNA and ubiquitin

The promiscuous metalloprotease SPRTN is the key enzyme for proteolytic repair of DNA-protein crosslinks (DPCs). To prevent uncontrolled SPRTN activity, its activation must be tightly regulated. Using NMR spectroscopy and in vitro reconstitution, we elucidate the molecular basis of SPRTNs activation by DNA and ubiquitin. We identify an autoinhibitory mechanism governed by intramolecular electrostatic interactions between a negatively charged linker helix and SPRTNs positively charged DNA-binding domains. DNA relieves this autoinhibition by competitively displacing the linker from the DNA-binding domains, thereby inducing a conformational shift to an open, catalytically active state. This open state enables ubiquitin binding to SPRTNs protease domain, further stabilizing the active conformation. Disruption of the interaction between the autoinhibitory linker and the DNA-binding domains locks SPRTN in a constitutively open state, resulting in enhanced protease activity. Collectively, our data reveal how DNA and ubiquitin cooperate to convert SPRTN from an autoinhibited conformation into its active state.

biochemistry↗

bioRxiv2025

Evolutionary Remodelling of a Remnant GET-Pathway Factor into PEX38, a Novel and Essential Peroxin in Euglenozoa

PEX19 is a cytosolic receptor that directs membrane proteins post-translationally to peroxisomes, as well as to mitochondria, lipid droplets, and the endoplasmic reticulum. A comprehensive Trypanosoma PEX19 interactome analysis uncovered PEX38 as a novel and essential Euglenozoa-specific peroxin. PEX38 contains distinct domains that bind the co- chaperone Hip and the PEX3-binding motif of PEX19, suggesting a role in stabilizing membrane protein and preventing premature membrane docking. PEX38 illustrates functional repurposing in organelle biogenesis. It originated from a remnant of the GET/TRC pathway, typically responsible for the targeting of tail-anchored proteins to the endoplasmic reticulum. While most components of this machinery are absent in Euglenozoa, PEX38 has been retained and adapted to mediate peroxisomal membrane protein targeting. This evolutionary adaptation is unique to Euglenozoa. Because the PEX19-PEX38 interaction is essential for parasite viability and PEX38 has no human homologs, this complex is a promising therapeutic target against trypanosomatid parasites.

cell biology↗

bioRxiv2025

It is Not Always Crystal Clear: In-Pocket Analysis for Understanding Binding Site Microenvironment Properties

Structural biology enabled key advances in biochemistry, drug discovery, and AI-driven three-dimensional structure modelling. Despite its importance, reliable and general computational tools for chemical validation of structural models remain limited, especially when small-molecule ligands are involved. This challenge is exacerbated when experimental resolution is insufficient to define interaction chemistry, leaving critical interpretations ambiguous. We introduce In-Pocket, a quantum mechanical optimisation method biased by experimental data that evaluates the chemical nature of protein-ligand binding interfaces. In-Pocket improves model quality, corrects chemically inconsistent structures, and identifies the energetically most favourable ligand-conformers within biomacromolecular complexes. The approach is generally applicable to a wide range of systems, including those containing transition metals. Benchmarking demonstrates that In-Pocket enhances the reliability of legacy and newly solved structures. With automated, user-friendly routine chemical validation, In-Pocket offers unique opportunities to improve the integrity of structural databases and impact diverse domains: from molecular biology to rational drug design and application of generative AI.

bioinformatics↗

bioRxiv2025

YAP1 drives aggressive and therapy resistant state in melanoma through reprogramming the chromatin and regulating immune evasive programs

Despite promising initial results in targeting the RAF-MEK-ERK cascade, resistance to BRAF/MEK inhibitors remains a critical challenge in nearly 50% of melanoma patients. Our study demonstrates that robust YAP1 activation in metastatic melanoma correlates with poor survival and drives transcriptional programs linked to therapeutic resistance. Mechanistically, YAP1 predominantly remodels the chromatin landscape in resistant tumors by partnering with BRD4 and TEAD, creating a permissive transcriptional state that sustains oncogenic signaling. Clinical validation in biopsies from resistant melanoma confirms elevated expression of YAP1 target genes. Furthermore, pharmacological inhibition of BRD4 or TEAD reduces YAP1-driven transcription and reactivates antitumor immunity programs. TEAD specific inhibitors (and not verteporfin which is a highly non-specific inhibitor) synergize with immune checkpoint blockade in in vivo model system by promoting increased CD8 T cell infiltration and prolonged survival in the melanoma mouse model. Collectively, these findings reveal a chromatin-centric vulnerability in BRAF/MEK inhibitor-resistant melanoma and propose TEAD specific inhibitors as a promising dual strategy to overcome resistance and reinvigorate the immune response, offering a novel therapeutic avenue for patients. Resistance to BRAF and MEK inhibitors remains a major obstacle in the treatment of melanoma. Here, we show that elevated YAP1 activity in metastatic melanoma is associated with poor patient survival and drives transcriptional programs that promote therapeutic resistance. Mechanistically, YAP1 cooperates with BRD4 and TEAD to reprogram the chromatin landscape, establishing an oncogenic transcriptional state that sustains resistance. Analysis of patient-derived melanoma biopsies confirms increased expression of YAP1 target genes in resistant tumors. Targeting this epigenetic circuitry using pharmacological inhibitors of BRD4 or TEAD effectively suppresses YAP1-driven transcriptional output and restores antitumor immune activity, as evidenced by enhanced CD8 T cell infiltration and prolonged survival in resistant melanoma models. These effects are further supported by transcriptomic and chromatin accessibility analyses, which reveal reduced expression of immune-suppressive and proliferation-associated gene networks upon TEAD inhibition. Collectively, our findings identify a chromatin-centric mechanism underlying resistance to MAPK-targeted therapy and nominate TEAD inhibition as a promising dual-action strategy to both overcome resistance and re-engage antitumor immunity. This work offers a compelling therapeutic avenue for patients with drug-resistant melanoma.

cancer biology↗

bioRxiv2025

QKI ensures splicing fidelity during cardiogenesis by engaging the U6 tri-snRNP to activate splicing at weak 5' splice sites

During organogenesis, precise pre-mRNA splicing is essential to assemble tissue architecture. Many developmentally essential exons bear weak 5' splice sites (5'SS) yet are spliced with high precision, implying unknown yet active splicing fidelity mechanisms. By combining transcriptome and alternative splicing profiling with temporal eCLIP mapping of RNA interactions across development, we identify the RNA-binding protein QKI as an essential direct regulator of splicing fidelity in key cardiac transcripts. Although QKI is dispensable for cardiac specification, its loss disrupts sarcomere assembly despite intact expression of sarcomere mRNAs through exon skipping and nuclear retention of mis-spliced RNAs. QKI-dependent exons in essential cardiac genes have weak 5'SS and frequently show poor complementarity with U6 snRNA. We show that QKI directly interacts with U6 snRNA using an overlapping interface to its traditional intronic binding activity, securing U4/U6{middle dot}U5 tri-snRNP to ensure splicing fidelity. Thus, QKI exemplifies how context-aware RBPs enforce splicing fidelity at structurally vulnerable splice sites during organogenesis.

cell biology↗

bioRxiv2025

The essential co-chaperone Sgt1 regulates client dwell time in the Hsp90 chaperone cycle

The Hsp90 machinery is the most complex chaperone system in the eukaryotic cell. It is characterized by numerous co-chaperones that modulate the function of Hsp90. In S. cerevisiae, most of these cofactors can be deleted without affecting viability. Of the three essential ones, only the function of Sgt1 remained enigmatic. Our in vivo and in vitro experiments define key structural elements and determine the essential function of Sgt1 in the chaperoning of client proteins. We show that yeast Sgt1 exhibits a unique binding mode to Hsp90. The simultaneous interaction of Sgt1 with Hsp90 and client proteins enhances client maturation efficiency. Specifically, Sgt1 stabilizes Hsp90-client complexes and prevents their dissociation by the co-chaperone Aha1. Together, our findings reveal a distinct regulatory mechanism of the Hsp90 function, highlighting Sgt1 as a critical modulator of chaperone cycle progression.

biochemistry↗

bioRxiv2024

Allosteric activation of the SPRTN protease by ubiquitin maintains genome stability

The DNA-dependent protease SPRTN maintains genome stability by degrading toxic DNA-protein crosslinks (DPCs). To understand how SPRTNs promiscuous protease activity is confined to the cleavage of crosslinked proteins, we reconstitute the repair of DPCs including their modification with SUMO and ubiquitin chains, using recombinant human proteins. We discover that DPC ubiquitylation strongly activates SPRTN independently of SPRTNs known ubiquitin-binding domains. Using protein structure prediction, MD simulations and NMR spectroscopy we reveal that ubiquitin binds to an interface at the back of SPRTNs protease domain, promoting an active conformation. Replacing key interfacial residues prevents ubiquitin-dependent activation of SPRTN, which leads to genomic instability and cell cycle defects in cells expressing hypomorphic SPRTN variants that cause premature aging and liver cancer in Ruijs-Aalfs syndrome patients. Collectively, our results demonstrate that SPRTN activation is coupled to the modification of the crosslinked protein, explaining how specificity is achieved during DPC repair.

biochemistry↗

bioRxiv2024

Deciphering the RNA recognition by Musashi-1 to design protein and RNA mutants for in vitro and in vivo applications

RNA Recognition Motifs (RRMs) are essential post-transcriptional regulators of gene expression in eukaryotic cells. The Human Musashi-1 (MSI-1) is an RNA-binding protein that recognizes (G/A)U1-3AGU and UAG sequences in diverse RNAs through two RRMs and regulates the fate of target RNA. Here, we combined structural biology and computational approaches to analyse the binding of the RRM domains of human MSI-1 with single-stranded and structured RNAs ligands. We used our recently developed computational tool RRMScorer to design a set of mutants of the MSI-1 protein to bind novel RNA sequences to alter the binding selectivity. The in-silico predictions of the designed protein-RNA interactions are assessed by NMR and SPR. These experiments also are used to study the competition of the two RRM domains of MSI-1 for the same binding site within linear and harpin RNA. Our experimental results confirm the in-silico designed interactions, thus opening the way for the development of new biomolecules for in vitro and in vivo studies and downstream applications.

biophysics↗

bioRxiv2024

Intrinsically disordered RNA-binding motifs cooperate to catalyze RNA folding and drive phase separation

RNA-binding proteins are essential for gene regulation and the spatial organization of cells. Here, we report that the yeast ribosome biogenesis factor Loc1p is an intrinsically disordered RNA-binding protein with eight repeating positively charged, unstructured nucleic acid binding (PUN) motifs. While a single of these previously undefined motifs stabilizes folded RNAs, multiple copies strongly cooperate to catalyze RNA folding. In the presence of RNA, these multivalent PUN motifs drive phase separation. Proteome-wide searches in pro-and eukaryotes for proteins with similar arrays of PUN motifs reveal a strong enrichment in RNA-mediated processes and DNA remodeling. Thus, PUN motifs are potentially involved in a large variety of RNA-and DNA-related processes by concentrating them in membrane-less organelles. The general function and wide distribution of PUN motifs across species suggests that in an ancient "RNA world" PUN-like motifs may have supported the correct folding of early ribozymes.

biochemistry↗

bioRxiv2023

The prion-like domain of the chloroplast RNA binding protein CP29A is required for cold-induced phase separation next to nucleoids and supports RNA splicing and translation during cold acclimation

Arabidopsis thaliana is capable of producing photosynthetic tissue with active chloroplasts at temperatures as low as 4{degrees}C, and this process depends on the presence of the nuclear-encoded, chloroplast-localized RNA-binding protein CP29A. In this study, we demonstrate that CP29A undergoes phase separation in vitro and in vivo in a temperature-dependent manner, which is mediated by a prion-like domain (PLD) located between the two RNA recognition motif (RRM) domains of CP29A. The resulting droplets display liquid-like properties and are found in close proximity to chloroplast nucleoids. The PLD is required to support chloroplast RNA splicing and translation in cold-treated tissue. Together, our findings suggest that plant chloroplast gene expression is compartmentalized by inducible condensation of CP29A at low temperatures, a mechanism that could play a crucial role for plant cold resistance.

plant biology↗

bioRxiv2023

PURA Syndrome-causing mutations impair PUR-domain integrity and affect P-body association

Mutations in the human PURA gene cause the neuro-developmental PURA syndrome. In contrast to several other mono-genetic disorders, almost all reported mutations in this nucleic acid binding protein result in the full disease penetrance. In this study, we observed that patient mutations across PURA impair its previously reported co-localization with processing bodies. These mutations either destroyed the folding integrity, RNA binding or dimerization of PURA. We also solved the crystal structures of the N- and C-terminal PUR domains of human PURA and combined them with molecular dynamics simulations and NMR measurements. The observed unusually high dynamics and structural promiscuity of PURA indicated that this protein is particularly susceptible to mutations impairing its structural integrity. It offers an explanation why even conservative mutations across PURA result in the full penetrance of symptoms in patients with PURA syndrome.

molecular biology↗

bioRxiv2023

A novel PEX14/PEX5 interface links peroxisomal protein import and receptor recycling

Newly synthesized peroxisomal proteins are recognized in the cytosol by the cycling receptor PEX5 and directed to a docking complex comprising PEX14 and PEX13 at the peroxisomal membrane. After cargo translocation, the unloaded PEX5 is recycled in an ATP-dependent manner. Receptor docking involves the WxxxF-motifs in the N-terminal domain (NTD) of PEX5 that are recognized by the N-terminal domain of PEX14. Here, we combine biochemical methods and NMR spectroscopy to identify a novel binding interface between human PEX5 and PEX14. The interaction involves the PEX5 C-terminal cargo-binding TPR domain and a conserved IPSWQI peptide motif in the C-terminal intrinsically disordered region of PEX14. The three-dimensional structure of the PEX14 IPSWQI peptide bound the PEX5 TPR domain, shows the PEX14 interaction is non-overlapping with PTS1 binding to the TPR domain. Notably, PEX14 IPSWQI motif binding to a hinge region in the TPR domain shows a more open supercoil of the TPR fold that resembles the apo conformation in the absence of PTS1 peptide. Mutation of binding site residues in PEX5 or PEX14 leads to a partial protein import defect and decrease of the steady-state-concentration of PEX5. This resembles the mutant phenotype of cells affected in receptor recycling, suggesting a role in this process.

biochemistry↗

bioRxiv2023

MISATO - Machine learning dataset for structure-based drug discovery

Large language models (LLMs) have greatly enhanced our ability to understand biology and chemistry. Yet, relatively few robust methods have been reported for structure-based drug discovery. Highly precise biomolecule-ligand interaction datasets are urgently needed in particular for LLMs, that require extensive training data. We present MISATO, the first dataset that combines quantum mechanics properties of small molecules and associated molecular dynamics simulations of about 20000 experimental protein-ligand complexes. Starting from the PDBbind dataset, semi-empirical quantum mechanics was used to systematically refine these structures. The largest collection to date of molecular dynamics traces of protein-ligand complexes in explicit water are included, accumulating to 170 s. We give ML baseline models and simple Python data loaders, and aim to foster a thriving community around MISATO (https://github.com/t7morgen/misato-dataset). An easy entry point for ML experts is provided without the need of deep domain expertise to enable the next generation of drug discovery AI models.

bioinformatics↗

bioRxiv2022

The RBM17/SPF45-SAP30BP interaction is essential for splicing in a subset of human short introns

Human pre-mRNA splicing requires the removal of introns with highly variable lengths, from tens to over a million nucleotides. Therefore, mechanisms of intron recognition and splicing are likely not universal. Recently, we reported that splicing in a subset of human short introns with truncated polypyrimidine tracts depends on RBM17 (SPF45), instead of the canonical splicing factor U2AF heterodimer. Here, we demonstrate that SAP30BP, a factor previously implicated in transcriptional control, is an essential splicing cofactor for RBM17. In vitro binding and NMR analyses demonstrate that a U2AF-homology motif (UHM) in RBM17 binds directly to a newly identified UHM-ligand motif (ULM) in SAP30BP. We show that this RBM17-SAP30BP interaction is required to specifically recruit RBM17 to phosphorylated SF3B1 (SF3b155), a U2 snRNP component in active spliceosomes. We propose a unique mechanism for splicing in a subset of short introns, in which SAP30BP guides RBM17 in the assembly of active spliceosomes. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=193 SRC="FIGDIR/small/522300v4_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@12da1c3org.highwire.dtl.DTLVardef@d71e53org.highwire.dtl.DTLVardef@108160corg.highwire.dtl.DTLVardef@1df7275_HPS_FORMAT_FIGEXP M_FIG C_FIG In briefFukumura et al. discover a general splicing mechanism in a subset of human short introns with truncated polypyrimidine tracts. This splicing reaction is mediated by intermediary RBM17-SAP30BP complex, instead of the known U2AF heterodimer. SAP30BP binding to RBM17 may support RBM17 association with active phosphorylated SF3B1 in U2 snRNP. HighlightsO_LIRBM17 (SPF45) is a splicing factor required for a subset of human short introns C_LIO_LISAP30BP is an essential cofactor, which interacts with RBM17 via UHM-ULM binding C_LIO_LIRBM17 forms a weak complex with SAP30BP before its binding with SF3B1 in U2 snRNP C_LIO_LIRBM17-SAP30BP complex supports RBM17 to be recruited to active phosphorylated SF3B1 C_LI

molecular biology↗

bioRxiv2022

Intramolecular autoinhibition of human PEX13 modulates peroxisomal import

Targeting and import of peroxisomal proteins depends on PEX5, PEX14 and PEX13. We present a biochemical and structural characterization of the PEX13 C-terminal region. By combining NMR spectroscopy, X-ray crystallography and biochemical methods, we show that the PEX13 SH3 domain mediates intramolecular interactions with a newly identified proximal FxxxF motif and also binds to WxxxF peptide motifs from the PEX5 NTD, demonstrating evolutionary conservation of this interaction from yeast to human. Strikingly, the C-terminal FxxxF motif autoinhibits the WxxxF/Y binding surface on the PEX13 SH3 domain. This is supported by high-resolution crystal structures, which show FxxxF or WxxxF/Y binding to the same, non-canonical surface on the SH3 domain. The FxxxF motif also binds the PEX14 NTD with micromolar affinity. Surprisingly, the canonical binding surface for PxxP motifs on the human PEX13 SH3 fold does not recognize PxxP motifs in PEX14, distinct from the yeast ortholog. The dynamic network of PEX13, PEX14 and PEX5 interactions mediated by diaromatic peptide motifs fine-tunes and modulates peroxisomal matrix import in cells.

biochemistry↗

bioRxiv2022

Structural basis for specific RNA recognition by the alternative splicing factor RBM5

The RNA-binding motif protein RBM5 belongs to a family of multi-domain RNA binding proteins that are implicated in cancer and regulate alternative splicing of genes important for apoptosis and cell proliferation and have been implicated in cancer. RBM5 harbors structural modules for RNA recognition, such as RRM domains and a Zn finger, and protein-protein interactions such as an OCRE domain. Here, we characterize binding of the RBM5 RRM1-ZnF1-RRM2 domains to cis-regulatory RNA elements. A structure of the RRM1-ZnF1 region in complex with RNA shows how the tandem domains cooperate to sandwich target RNA and specifically recognize a GG dinucleotide in a non-canonical fashion. While the RRM1-ZnF1 domains act as a single structural module, RRM2 is connected by a flexible linker and tumbles independently. However, all three domains participate in RNA binding and adopt a closed architecture upon RNA binding. Our data highlight how cooperativity and conformational modularity of multiple RNA binding domains enable the recognition of distinct RNA motifs, thereby contributing to the regulation of alternative splicing. Remarkably, we observe surprising differences in coupling of the RNA binding domains between the closely related homologs RBM5 and RBM10. HighlightsO_LIMultiple RNA binding domains enable differential recognition of distinct RNA motifs to regulate alternative splicing C_LIO_LIThe RRM1-ZnF1 domains of RBM5 mediate specific recognition of cis regulatory RNA motifs to modulate alternative splicing C_LIO_LIRRM1-ZnF1 sandwich the target RNA for non-canonical recognition of a GG dinucleotide C_LIO_LIRRM1-ZnF1-RRM2 bind cooperatively to contiguous cis-regulatory motifs in caspase-2 and NUMB pre-mRNAs C_LI

biochemistry↗

bioRxiv2022

Reliability and accuracy of single-molecule FRET studies for characterization of structural dynamics and distances in proteins

Single-molecule FRET (smFRET) has become an established tool to study biomolecular structure and dynamics in vitro and in live cells. We performed a worldwide blind study involving 19 labs to assess the uncertainty of FRET experiments for proteins with respect to the measured FRET efficiency histograms, determination of distances, and the detection and quantification of structural dynamics. Using two protein systems that undergo distinct conformational changes, we obtained an uncertainty of the FRET efficiency of less than {+/-} 0.06, corresponding to an interdye distance precision of [≤] 0.2 nm and accuracy of [≤] 0.5 nm. We further discuss the limits for detecting distance fluctuations with sensitivity down to [lsim] 10% of the Forster distance and provide guidelines on how to detect potential dye perturbations. The ability of smFRET experiments to simultaneously measure distances and avoid averaging of conformational dynamics slower than the fluorescence lifetime is unique for dynamic structural biology.

biophysics↗