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Müller, F.

Publications and source records attributed to Müller, F..

12 recordsLinked to original sources
bioRxiv2025

Unified down-stream analysis of crosslinking mass spectrometry results with pyXLMS

Crosslinking mass spectrometry has become the method of choice for the identification of protein-protein interactions and for gaining insight into the structures of proteins in vivo. However, connecting crosslink search engine results with down-stream analysis tools, and therefore gaining biological insight from crosslink identifications, has remained a manual and cumbersome step in the analysis that often requires expert bioinformatics knowledge. Here we introduce pyXLMS, a python package and public web application which aims to simplify and streamline this intermediate step, enabling researchers even without bioinformatics knowledge to conduct in-depth crosslink analyses. In its current state pyXLMS supports input from seven different crosslink search engines, as well as the mzI-dentML format of the HUPO Proteomics Standards Initiative. Down-stream analysis is facilitated by functionality that is directly available within pyXLMS such as aggregation, validation, annotation, filtering, and visualization. In addition, the data can easily be exported to more than ten supported down-stream analysis tools and formats. We demonstrate the applicability and benefits of pyXLMS by re-analyzing a publicly available crosslink dataset with a variety of different search engines and show how the same data analysis workflow can be applied using pyXLMS. pyXLMs is available via https://github.com/hgb-bin-proteomics/pyXLMS.

bioinformatics↗
bioRxiv2025

A PLETHORA transcription factor shapes cucumber shoot architecture

O_LIPLETHORA transcription factors (PLTs) are master regulators of plant development. Loss of shoot meristematic PLTs leads to reduced phyllotactic regularity and robustness in Arabidopsis and increased inflorescence branching in tomato. Whether these factors have similar functions in other species is not known. C_LIO_LITo address this knowledge gap, we integrated phylogenetic, transcriptomic, and genome-wide in vitro binding strategies with quantitative shoot architecture phenotyping of a panel of cucumber TILLING mutants of each CsPLT homolog. C_LIO_LIWe determined that CsPLT3/7 and CsPLT are expressed in complementary domains in the cucumber shoot apex. DAP-seq data indicated that both transcription factors recognised an ANT-like consensus motif and bound to SAM organising genes. We identified strong phenotypic defects in multiple Csplt3/7 mutants. In mature seeds, Csplt3/7 mutants formed a flat apex instead of a shoot apical meristem, from which secondary meristems emerged during subsequent seedling development. Mature Csplt3/7 shoots exhibited defects in internode length regularity, stem architecture, flower morphology, and axillary organ initiation. Moreover, phyllotactic patterns were slightly shifted from a spiral towards a distichous orientation. C_LIO_LIWe present one of the first pieces of evidence that the PLT3/7 clade fulfils both conserved and diversified roles in shoot development across taxonomic family boundaries. C_LI

plant biology↗
bioRxiv2024

Proteome-wide non-cleavable crosslink identification with MS Annika 3.0 reveals the structure of the C. elegans Box C/D complex

AbstractThe field of crosslinking mass spectrometry has seen substantial advancements over the past decades, enabling the structural analysis of proteins and protein-complexes and serving as a powerful tool in protein-protein interaction studies. However, data analysis of large non-cleavable crosslink studies is still a mostly unsolved problem due to its n-squared complexity. We here introduce a novel algorithm for the identification of non-cleavable crosslinks implemented in our crosslinking search engine MS Annika that is based on sparse matrix multiplication and allows for proteome-wide searches on commodity hardware. Application of this new algorithm enabled us to employ a proteome-wide search of C. elegans nuclei samples, where we were able to uncover previously unknown protein interactions and conclude a comprehensive structural analysis that provides a detailed view of the Box C/D complex, enhancing our understanding of its assembly and functional dynamics. Our findings provide valuable insights into the intricate regulation of cellular homeostasis and immune responses, which are conserved across species, including humans. Moreover, our algorithm will enable researchers to conduct similar studies that were previously unfeasible.

bioinformatics↗
bioRxiv2024

Integration of ATAC and RNA-sequencing identifies chromatin and transcriptomic signatures in classical and non-classical zebrafish osteoblasts and indicates mechanisms of entpd5a regulation

Two types of osteoblasts are required to assemble the zebrafish embryonic skeleton: classical osteoblasts homologous to the mammalian cell, and notochord sheath cells, which serve as non-classical osteoblasts. The gene entpd5a is critically required for ossification via both types of osteoblasts. Despite the indispensability of zebrafish models in vertebrate research, the genetic regulation of bone formation, as well as mechanisms of transcriptional control of entpd5a, remain largely unknown. Here, using a newly generated transgenic line, we isolate classical and non-classical osteoblasts from zebrafish embryos and performed both ATAC-seq and RNA-seq. We analysed results independently and integratively to understand those chromatin dynamics and accompanying transcriptomic changes that occur in different skeletal cell types. We show that although Dlx family factors are playing important roles in classical osteoblast regulation, Hox family factors are involved in governing spinal ossification via non-classical osteoblasts. We further present a resource-driven analysis of the entpd5a promoter, experimentally validating the ATAC-seq dataset and proposing mechanisms of regulating the complex entpd5a expression pattern during zebrafish osteogenesis. Our results thus provide a necessary comprehensive resource for the field of bone development and indicate spatio-temporally regulated promoter/enhancer interactions taking place in the entpd5a locus.

developmental biology↗
bioRxiv2022

The effect of lysergic acid diethylamide (LSD) on whole-brain functional and effective connectivity

Psychedelics have emerged as promising candidate treatments for various psychiatric conditions, and given their clinical potential, there is a need to identify biomarkers that underlie their effects. Here, we investigate the neural mechanisms of lysergic acid diethylamide (LSD) using regression dynamic causal modelling (rDCM), a novel technique that assesses whole-brain effective connectivity (EC) during resting-state functional magnetic resonance imaging (fMRI). We modelled data from two randomized, placebo-controlled, double-blind, cross-over trials, in which 45 participants were administered 100g LSD and placebo in two resting-state fMRI sessions. We compared EC against whole-brain functional connectivity (FC) using classical statistics and machine learning methods. Multivariate analyses of EC parameters revealed widespread increases in interregional connectivity and reduced self-inhibition under LSD compared to placebo, with the notable exception of primarily decreased interregional connectivity and increased self-inhibition in occipital brain regions. This finding suggests that LSD perturbs the Excitation/Inhibition balance of the brain. Moreover, random forests classified LSD vs. placebo states based on FC and EC with comparably high accuracy (FC: 85.56%, EC: 91.11%) suggesting that both EC and FC are promising candidates for clinically-relevant biomarkers of LSD effects.

neuroscience↗
bioRxiv2022

Multi-excitation Raman Spectroscopy Complements Whole Genome Sequencing for Rapid Detection of Bacterial Infection and Resistance in WHO Priority Pathogens

Current methods for diagnosing acute and complex infections mostly rely on culture-based methods and, for biofilms, fluorescence in-situ hybridization. These techniques are labor-intensive and can take 2-4 days to return a test result, especially considering an extra culturing step required for the antibiotic susceptibility testing (AST). This places a significant burden on healthcare providers, delaying treatment and leading to adverse patient outcomes. Here, we report the complementary use of our newly developed multi-excitation Raman spectroscopy (ME-RS) method with whole-genome sequencing (WGS). Four WHO priority pathogens are AST phenotyped and their antimicrobial resistance (AMR) profile determined by WGS. On application of ME-RS method we find high correlation with the WGS characterization. Highly accurate classification based on the species (98.93%), wild-type/non-wild type (99.45%), and presence or absence of thick peptidoglycan layers in cell walls (100%), as well as at the individual strain level (99.29%). These results clearly demonstrate the potential of ME-RS as a rapid and first-stage tool for species, resistance and strain-level classification which can be followed up by WGS for confirmation. Such a workflow can facilitate efficient antimicrobial stewardship to handle and prevent the spread of AMR.

microbiology↗
bioRxiv2020

Genome wide efficiency profiling reveals modulation of maintenance and de novo methylation by Tets

A precise understanding of DNA methylation dynamics on a genome wide scale is of great importance for the comprehensive investigation of a variety of biological processes such as reprogramming of somatic cells to iPSCs, cell differentiation and also cancer development. To date, a complex integration of multiple and distinct genome wide data sets is required to derive the global activity of DNA modifying enzymes. We present GwEEP - Genome-wide Epigenetic Efficiency Profiling as a versatile approach to infer dynamic efficiency changes of DNA modifying enzymes at base pair resolution on a genome wide scale. GwEEP relies on genome wide oxidative Hairpin Bisulfite sequencing (HPoxBS) data sets, which are translated by a sophisticated hidden Markov model into quantitative enzyme efficiencies with reported confidence around the estimates. GwEEP in its present form predicts de novo and maintenance methylation efficiencies of Dnmts, as well as the hydroxylation efficiency of Tets but its purposefully flexible design allows to capture further oxidation processes such as formylation and carboxylation given available data in the future. Applied to a well characterized ES cell model, GwEEP precisely predicts the complex epigenetic changes following a Serum-to-2i shift i.e., (i) instant reduction in maintenance efficiency (ii) gradually decreasing de novo methylation efficiency and (iii) increasing Tet efficiencies. In addition, a complementary analysis of Tet triple knock-out ES cells confirms the previous hypothesized mutual interference of Dnmts and Tets. GwEEP is applicable to a wide range of biological samples including cell lines, but also tissues and primary cell types. MOTIVATIONDynamic changes of DNA methylation patterns are a common phenomenon in epigenetics. Although a stable DNA methylation profile is essential for cell identity, developmental processes require the rearrangement of 5-methylcytosine in the genome. Stable methylation patterns are the result of balanced Dnmts and Tets activities, while methylome transformation results from a coordinated change in Dnmt and Tet efficiencies. Such transformations occur on a global scale, for example during the reprogramming of maternal and paternal methylation patterns and the establishment of novel cell type specific methylomes during embryonic development in vivo, but also in vitro during (re)programming of induced pluripotent stem cells, as well as somatic cells. In addition, local (de)methylation events are key for gene regulation during cell differentiation. A detailed understanding of Dnmt and Tet cooperation is essential for understanding natural epigenetic adaptation as well as optimization of in vitro (re)programming protocols. For this purpose, we developed a pipeline for quantitative and precise estimation of Dnmt and Tet activity. Using only double strand methylation information, GwEEP infers accurate maintenance and de novo methylation efficiency of Dnmts, as well as hydroxylation efficiency of Tets at single base resolution. Thus, we believe GwEEP provides a powerful tool for the investigation of methylome rearrangements in various systems.

genomics↗
bioRxiv2020

Cdk1-dependent destabilization of long astral microtubules is required for spindle orientation

The precise execution of mitotic spindle orientation in response to cell shape cues is important for tissue organization and development. The presence of astral microtubules extending from the centrosome towards the cell cortex is essential for this process, but little is understood about the contribution of astral microtubule dynamics to spindle positioning, or how astral microtubule dynamics are regulated spatiotemporally. The mitotic regulator Cdk1-CyclinB promotes destabilization of centrosomal microtubules and increased microtubule dynamics as cells transition from interphase to mitosis, but how Cdk1 activity specifically modulates astral microtubule stability, and whether it impacts spindle positioning, is unknown. Here we uncover a mechanism revealing that Cdk1 destabilizes astral microtubules to ensure spindle reorientation in response to cell shape. Phosphorylation of the EB1-dependent microtubule plus-end tracking protein GTSE1 by Cdk1 in early mitosis abolishes its interaction with EB1 and recruitment to microtubule plus-ends. Loss of Cdk1 activity, or mutation of phosphorylation sites in GTSE1, induces recruitment of GTSE1 to growing microtubule plus-ends in mitosis. This decreases the catastrophe frequency of astral microtubules, and causes an increase in the number of long astral microtubules reaching the cell cortex, which restrains the ability of cells to reorient spindles along the long cellular axis in early mitosis. Astral microtubules must thus not only be present, but also dynamic to allow the spindle to reorient in response to cell shape, a state achieved by selective destabilization of long astral microtubules via Cdk1.

cell biology↗
bioRxiv2020

A localization screen reveals translation factories and widespread co-translational RNA targeting

Local translation allows a spatial control of gene expression. Here, we performed a dual protein/mRNA localization screen, using smFISH on 523 human cell lines expressing GFP-tagged genes. A total of 32 mRNAs displayed specific cytoplasmic localizations, and we observed local translation at unexpected locations, including cytoplasmic protrusions, cell edges, endosomes, Golgi, the nuclear envelope and centrosomes, the latter being cell cycle dependent. Quantitation of mRNA distribution and automatic pattern classification revealed a high degree of localization heterogeneity between cells. Surprisingly, mRNA localization frequently required ongoing translation, indicating widespread co-translational RNA targeting. Interestingly, while P-body accumulation was frequent (15 mRNAs), four mRNAs accumulated in foci that were distinct structures. These foci lacked the mature protein, but nascent polypeptide imaging showed that they were specialized translation factories. For {beta}-catenin, foci formation was regulated by Wnt, relied on APC-dependent polysome aggregation, and led to nascent protein degradation. Thus, translation factories uniquely regulate nascent protein metabolism and create a fine granular compartmentalization of translation.

cell biology↗
bioRxiv2019

Unique and assay specific features of NOMe-, ATAC- and DNase I-seq data

Chromatin accessibility maps are important for the functional interpretation of the genome. Here, we systematically analysed assay specific differences between DNase I-Seq, ATAC-Seq and NOMe-Seq in a side by side experimental and bioinformatic setup. We observe that most prominent nucleosome depleted regions (NDRs, e.g. in promoters) are roboustly called by all three or at least two assays. However we also find a high proportion of assay specific NDRs that are often "called" by only one of the assays. We show evidence that these assay specific NDRs are indeed genuine open chromatin sites and contribute important information for accurate gene expression prediction. While technically ATAC-Seq and DNAse I-Seq provide a high NDR calling rate for relatively low sequencing costs in comparison to NOMe-Seq, NOMe-Seq singles out as it provides a multitude of information: it allows to not only detect NDRs but also endogenous DNA methylation, genome wide segmentation into heterochromatic A/B domains and local phasing of nucleosomes outside of NDRs. In summary our comparison strongly suggest to consider assay specific differences for the experimental desgin and for generalized and comparative functional interpretations.

genomics↗
bioRxiv2018

SLIC-CAGE: high-resolution transcription start site mapping using nanogram-levels of total RNA

Cap analysis of gene expression (CAGE) is a methodology for genome-wide quantitative mapping of mRNA 5ends to precisely capture transcription start sites at a single nucleotide resolution. In combination with high-throughput sequencing, CAGE has revolutionized our understanding of rules of transcription initiation, led to discovery of new core promoter sequence features and discovered transcription initiation at enhancers genome-wide. The biggest limitation of CAGE is that even the most recently improved version (nAnT-iCAGE) still requires large amounts of total cellular RNA (5 micrograms), preventing its application to scarce biological samples such as those from early embryonic development or rare cell types. Here, we present SLIC-CAGE, a Super-Low Input Carrier-CAGE approach to capture 5ends of RNA polymerase II transcripts from as little as 5-10 ng of total RNA. The dramatic increase in sensitivity is achieved by specially designed, selectively degradable carrier RNA. We demonstrate the ability of SLIC-CAGE to generate data for genome-wide promoterome with 1000-fold less material than required by existing CAGE methods by generating a complex, high quality library from mouse embryonic day (E) 11.5 primordial germ cells.

genomics↗
bioRxiv2018

Partially methylated domains are hallmarks of a cell specific epigenome topology

BackgroundPartially methylated domains, PMDs, are extended regions in the genome exhibiting a reduced average DNA-methylation level. PMDs cover gene-poor and transcriptionally inactive regions and tend to be heterochromatic. Here, we present a first comprehensive comparative analysis of PMDs across more than 190 WGBS methylomes of human and mouse cells providing a deep insight into structural and functional features associated with PMDs.\n\nResultsPMDs are ubiquitous signatures covering up to 75% of the genome in human and mouse cells irrespective of their tissue or cell origin. Additionally, each cell type comes with a distinct set of specific PMDs, and genes expressed in such PMDs show a strong cell type effect. Demethylation strength varies in PMDs with a tendency towards a more pronounced effect in differentiating and replicating cells. The strongest demethylation is observed in highly proliferating and immortal cancer cell lines. A decrease of DNA-methylation within PMDs tends to be linked to an increase in heterochromatic histone marks and a decrease of gene expressions. Characteristic combinations of heterochromatic signatures in PMDs are linked to domains of early, middle and late DNA-replication.\n\nConclusionPMDs are prominent signatures of long-range epigenomic organization. Integrative analysis identifies PMDs as important general, lineage- and cell-type specific topological features. PMD changes are hallmarks of cell differentiation. Demethylation of PMDs combined with increased heterochromatic marks is a feature linked to enhanced cell proliferation. In combination with broad histone marks PMDs demarcate distinct domains of late DNA-replication.

bioinformatics↗