Biology subjects

Wierer, M.

Publications and source records attributed to Wierer, M..

11 recordsLinked to original sources

bioRxiv2025

A dietary pan-amino acid dropout screen in vivo reveals a critical role for histidine in T-ALL

Dietary interventions show therapeutic potential in cancer, but systematic comparisons are lacking. We performed a dietary pan-amino acid dropout screen in an orthotopic model of NOTCH1-driven T-cell acute lymphoblastic leukemia and identified histidine depletion as uniquely antileukemic. Histidine-restricted diets extended survival of leukemic mice in a dose-dependent manner, while remaining well-tolerated. Mechanistically, multiomic profiling revealed that histidine deprivation-induced ribosome stalling activates GCN2 to suppress cholesterol biosynthesis pathways critical for leukemic proliferation. Dietary cholesterol supplementation partially reverted the antileukemic effects of histidine restriction in vivo. These findings couple histidine levels and translational control to cholesterol metabolism, which can be therapeutically exploited for cancer treatment. Our results suggest that defined dietary amino acid restrictions may expose broader therapeutic opportunities in diseases beyond cancer.

cancer biology↗

bioRxiv2025

MODY mutations in transcription factors HNF1A and HNF1B affect the production of interferon signaling proteins: evidence at the proteome level

We investigated the proteomic consequences of MODY-associated mutations in the transcription factors HNF1A and HNF1B using two cell line models. Quantitative label-free mass spectrometry, employing both data-dependent and data-independent acquisition methods, revealed consistent suppression of energy metabolism and interferon signaling pathways. Pathway and protein interaction analyses confirmed these findings. In cells with the HNF1A frameshift mutation, several predicted transcriptional targets, including A1CF, were significantly downregulated. These results reinforce the link between HNF1A/HNF1B function and mitochondrial activity as well as innate immune signaling, providing a foundation for further mechanistic studies and causal inference research.

systems biology↗

bioRxiv2025

Rab2 and Arl8/BORC control retrograde axonal transport of dense core vesicles via Syd/dJIP3/4 and RUFY dynein adaptors

Neuropeptide-containing dense core vesicles (DCVs) are generated in the neuronal cell body and circulate throughout the axonal arbor to supply distal release sites. This circulation depends on the anterograde kinesin-1 and kinesin-3 motors and the retrograde dynein-dynactin motor. While kinesin-3 is recruited to DCVs with the aid of the small GTPase Arl8, it is unclear how dynein and kinesin-1 are recruited and regulated. Here we show that DCV motility in Drosophila (fruit flies) depends on the dynein and kinesin-1 adaptor Sunday Driver/dJIP3/4 (Syd) and the novel dynein adaptor RUFY. Syd and RUFY bind each other; moreover, Syd binds the DCV-located GTPase Rab2 that controls retrograde DCV transport, and RUFY binds Arl8. Disruption of Rab2, Syd, RUFY, dynein, and the Arl8 activator BORC all produce a similar DCV axonal transport phenotype characterized by axonal accumulation of immobile DCVs and a selective reduction in retrograde DCV flux. Our data suggest a model where dynein is recruited and activated by a complex of Syd and RUFY, which is anchored to DCVs by a Rab2- and Arl8-dependent mechanism. Lastly, we show that loss of Rab2 results in missorting of the DCV membrane proteins VMAT and Synaptotagmin-, similar to the reported effect of Rab2 deletion on the sorting of synaptic vesicle and active zone proteins. However, disruption of Syd, RUFY or dynein does not phenocopy the Rab2-specific VMAT sorting defect, suggesting that Rab2 employs separate effectors in DCV biogenesis and motility.

neuroscience↗

bioRxiv2025

New methods on the block: Taxonomic identification of archaeological bones in resin-embedded sediments through palaeoproteomics

The integration of biomolecular studies of past organisms with geoarchaeological studies can significantly improve our understanding of the relative chronology and context of archaeologically (in)visible behaviours. However, the complexity and sedimentological heterogeneity of archaeological deposits at a microscopic scale is often not taken into consideration in biomolecular studies. Here, we investigate the preservation and retrieval of palaeoproteomic data from bone fragments embedded in Pleistocene resin-impregnated sediment blocks. We show that resin impregnation has minimal effect on skeletal protein taxonomic identifications in modern skeletal material, but observe an increase in oxidation-related post-translational modifications. We then successfully retrieve proteins from resin-impregnated blocks from the Palaeolithic sites of Bacho Kiro Cave, La Ferrassie and Quincay. The taxonomic identifications of minute bones encased in resin are in line with previous analyses of the faunal communities of these sites, with a diversity of taxa (Bos sp./Bison sp., Equus sp., Ursus sp., and Caprinae) observed at a microscale in Bacho Kiro. This differs from results from La Ferrassie where most of the samples are identified as a single taxon (Bos sp./Bison sp.) across different areas of the site. The block from Quincay only provided taxonomic identification of two out of eleven bone-derived samples, likely due to diagenesis. Our work indicates that palaeoproteomes can be retrieved from bone fragments at a microstratigraphic resolution, enabling the detailed study of faunal community composition at a scale that more closely matches that of past human occupations. Significance StatementResin-embedded sediment blocks are widely used in archaeology and soil sciences to reconstruct past environments and human behavior, but their potential for biomolecular analysis is underexplored. Here, we demonstrate that ancient proteins can be successfully retrieved from bone fragments embedded in resin-impregnated sediment blocks from Pleistocene archaeological sites. Our findings show that resin impregnation has a minimal impact on protein recovery and that palaeoproteomics enables taxonomic identification of Pleistocene bone fragments at a microstratigraphic scale. This approach allows for reconstructing past faunal communities with unprecedented detail, improving our understanding of ancient ecosystems and the environmental contexts of early hominin occupations.

evolutionary biology↗

bioRxiv2024

Reprogramming neuroblastoma by diet-enhanced polyamine depletion

Neuroblastoma is a highly lethal childhood tumor derived from differentiation-arrested neural crest cells1,2. Like all cancers, its growth is fueled by metabolites obtained from either circulation or local biosynthesis3,4. Neuroblastomas depend on local polyamine biosynthesis, with the inhibitor difluoromethylornithine showing clinical activity5. Here we show that such inhibition can be augmented by dietary restriction of upstream amino acid substrates, leading to disruption of oncogenic protein translation, tumor differentiation, and profound survival gains in the TH-MYCN mouse model. Specifically, an arginine/proline-free diet decreases the polyamine precursor ornithine and augments tumor polyamine depletion by difluoromethylornithine. This polyamine depletion causes ribosome stalling, unexpectedly specifically at adenosine-ending codons. Such codons are selectively enriched in cell cycle genes and low in neuronal differentiation genes. Thus, impaired translation of these codons, induced by the diet-drug combination, favors a pro-differentiation proteome. These results suggest that the genes of specific cellular programs have evolved hallmark codon usage preferences that enable coherent translational rewiring in response to metabolic stresses, and that this process can be targeted to activate differentiation of pediatric cancers. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=192 SRC="FIGDIR/small/573662v1_ufig1.gif" ALT="Figure 1"> View larger version (64K): org.highwire.dtl.DTLVardef@1fdbaecorg.highwire.dtl.DTLVardef@fb03f4org.highwire.dtl.DTLVardef@1c0afa7org.highwire.dtl.DTLVardef@14c384f_HPS_FORMAT_FIGEXP M_FIG C_FIG Highlights- Extra-tumoral conversion of arginine feeds tumor ornithine via uptake from circulation in MYCN-neuroblastoma. - A proline and arginine free diet enhances pharmacological polyamine depletion via reduced ornithine substrate availability. - Polyamine depletion disrupts oncogenic translation to induce a pro-differentiation proteome causing neuroblast differentiation and prolonged survival in the TH-MYCN mouse model. - Genes of specific cellular programs have evolved codon usage preferences that enable coherent translational rewiring in response to metabolic stress, such as polyamine depletion.

cancer biology↗

bioRxiv2023

The novel coronary artery disease risk factor ADAMTS-7 modulates atherosclerotic plaque formation by degradation of TIMP-1

BackgroundThe ADAMTS7 locus was genome-wide significantly associated with coronary artery disease (CAD). Lack of the extracellular matrix (ECM) protease ADAMTS-7 was shown to reduce atherosclerotic plaque formation. ObjectiveTo identify molecular mechanisms and downstream targets of ADAMTS-7 mediating risk of atherosclerosis. MethodsTargets of ADAMTS-7 were identified by high-resolution mass spectrometry of atherosclerotic plaques from Apoe-/- and Apoe-/-Adamts7-/- mice. ECM proteins were identified using solubility profiling. Putative targets were validated using immunofluorescence, in vitro degradation assays, co-immunoprecipitation, and Forster resonance energy transfer (FRET)-based protein-protein interaction assays. ADAMTS7 expression was measured in fibrous caps of human carotid artery plaques. ResultsIn humans, ADAMTS7 expression was higher in caps of unstable as compared to stable carotid plaques. Compared to Apoe-/- mice, atherosclerotic aortas of Apoe-/- mice lacking Adamts-7 (Apoe-/-Adamts7-/-) contained higher protein levels of tissue inhibitor of metalloproteases 1 (Timp-1). In co-immunoprecipitation experiments, the catalytic domain of ADAMTS-7 bound to TIMP-1, which was degraded in the presence of ADAMTS-7 in vitro. ADAMTS-7 reduced the inhibitory capacity of TIMP-1 at its canonical target matrix metalloprotease 9 (MMP-9) As a downstream mechanism, we investigated collagen content in plaques of Apoe-/- and Apoe-/-Adamts7-/- mice after Western diet. Picrosirius red staining of the aortic root revealed less collagen as a readout of higher MMP-9 activity in Apoe-/- as compared to Apoe-/- Adamts7-/- mice. In order to facilitate high-throughput screening for ADAMTS-7 inhibitors with the aim to decrease TIMP-1 degradation, we designed a FRET-based assay targeting the ADAMTS-7 catalytic site. ConclusionADAMTS-7, which is induced in unstable atherosclerotic plaques, decreases TIMP-1 stability reducing its inhibitory effect on MMP-9, which is known to promote collagen degradation and is likewise genome-wide significantly associated with CAD. Disrupting the interaction of ADAMTS-7 and TIMP-1 might be a strategy to increase collagen content and plaque stability for reduction of atherosclerosis-related events.

molecular biology↗

bioRxiv2022

FK506-binding protein 2 participates in proinsulin folding

Apart from chaperoning, disulphide bond formation and downstream processing, the molecular sequence of proinsulin folding is not completely understood. Proinsulin requires proline isomerization for correct folding. Since FK506-binding protein 2 (FKBP2) is an ER-resident proline isomerase, we hypothesized that FKBP2 contributes to proinsulin folding. We found that FKBP2 co-immunoprecipitated with proinsulin and its chaperone GRP94, and that inhibition of FKBP2 expression increased proinsulin turnover with reduced intracellular proinsulin and insulin levels. This phenotype was accompanied by an increased proinsulin secretion and the formation of proinsulin high molecular weight complexes, a sign of proinsulin misfolding. FKBP2 knockout in pancreatic {beta}-cells increased apoptosis without detectable upregulation of ER-stress response genes. Interestingly, FKBP2 mRNA was overexpressed in {beta}-cells from pancreatic islets of T2D patients. Based on molecular modelling and an in vitro enzymatic assay, we suggest that proline at position 28 of the proinsulin B chain (P28) is the substrate of FKBP2s isomerization activity. We propose that this isomerization step catalyzed by FKBP2 is an essential sequence required for correct proinsulin folding.

molecular biology↗

bioRxiv2022

Comprehensive chromatin proteomics resolves functional phases of pluripotency

The establishment of cellular identity is driven by transcriptional and epigenetic regulation exerted by the components of the chromatin proteome - the chromatome. However, chromatome composition and its dynamics in functional phases of pluripotency have not been comprehensively analyzed thus limiting our understanding of these processes. To address this problem, we developed an accurate mass spectrometry (MS)-based proteomic method called Chromatin Aggregation Capture (ChAC) followed by Data-Independent Acquisition (DIA) to analyze chromatome reorganizations during the transition from ground to formative and primed pluripotency states. This allowed us to generate a comprehensive atlas of proteomes, chromatomes, and chromatin affinities for the three pluripotency phases, revealing the specific binding and rearrangement of regulatory complexes. The technical advances, the comprehensive chromatome atlas, and the extensive analysis reported here provide a foundation for an in-depth understanding of mechanisms that govern the phased progression of pluripotency and changes of cellular identities in development and disease.

biochemistry↗

bioRxiv2021

Unscheduled DNA replication in G1 causes genome instability through head-to-tail replication fork collisions

DNA replicates once per cell cycle. Interfering with the regulation of DNA replication initiation generates genome instability through over-replication and has been linked to early stages of cancer development. Here, we engineered genetic systems in budding yeast to induce unscheduled replication in the G1-phase of the cell cycle. Unscheduled G1 replication initiated at canonical S-phase origins across the genome. We quantified differences in replisomes in G1- and S-phase and identified firing factors, polymerase , and histone supply as factors that limit replication outside S-phase. G1 replication per se did not trigger cellular checkpoints. Subsequent replication during S-phase, however, resulted in over-replication and led to chromosome breaks via head-to-tail replication fork collisions that are marked by chromosome-wide, strand-biased occurrence of RPA-bound single-stranded DNA. Low-level, sporadic induction of G1 replication induced an identical response, indicating findings from synthetic systems are applicable to naturally occurring scenarios of unscheduled replication initiation by G1/S deregulation.

molecular biology↗

bioRxiv2020

Defining the RBPome of T helper cells to study higher order post-transcriptional gene regulation

Post-transcriptional gene regulation is complex, dynamic and ensures proper T cell function. The targeted transcripts can simultaneously respond to various factors as evident for Icos, an mRNA regulated by several RNA binding proteins (RBPs), including Roquin. However, fundamental information about the entire RBPome involved in post-transcriptional gene regulation in T cells is lacking. Here, we applied global RNA interactome capture (RNA-IC) and orthogonal organic phase separation (OOPS) to human and mouse primary T cells and identified the core T cell RBPome. This defined 798 mouse and 801 human proteins as RBPs, unexpectedly containing signaling proteins like Stat1, Stat4 and Vav1. Based on the vicinity to Roquin-1 in proximity labeling experiments, we selected [~]50 RBPs for testing coregulation of Roquin targets. Induced expression of these candidate RBPs in wildtype and Roquin-deficient T cells unraveled several Roquin-independent contributions, but also revealed Celf1 as a new Roquin-1-dependent and target-specific coregulator of Icos. One sentence statementWe provide an atlas of RNA-binding proteins in human and mouse T helper cells as a resource for studying higher order post-transcriptional gene regulation.

immunology↗

bioRxiv2020

Drosophila SWR1 and NuA4 complexes are defined by DOMINO isoforms

Histone acetylation and deposition of H2A.Z variant are integral aspects of active transcription. In Drosophila, the single DOMINO chromatin regulator complex is thought to combine both activities via an unknown mechanism. Here we show that alternative isoforms of the DOMINO nucleosome remodeling ATPase, DOM-A and DOM-B, directly specify two distinct multi-subunit complexes. Both complexes are necessary for transcriptional regulation but through different mechanisms. The DOM-B complex incorporates H2A.V (the fly ortholog of H2A.Z) genome-wide in an ATP-dependent manner, like the yeast SWR1 complex. The DOM-A complex, instead, functions as an ATP-independent histone acetyltransferase complex similar to the yeast NuA4, targeting lysine 12 of histone H4. Our work provides an instructive example of how different evolutionary strategies lead to similar functional separation. In yeast and humans, nucleosome remodeling and histone acetyltransferase complexes originate from gene duplication and paralog specification. Drosophila generates the same diversity by alternative splicing of a single gene. HighlightsO_LIIsoforms of DOMINO dictate the formation of distinct complexes C_LIO_LIDOM-B complex is the Drosophila SWR1 and incorporates H2A.V genome-wide C_LIO_LIDOM-A complex acetylates H4K12 and is the Drosophila NuA4 C_LI

molecular biology↗