Biology subjects

Becker, R.

Publications and source records attributed to Becker, R..

16 recordsLinked to original sources

bioRxiv2026

Chromosome-scale genome assembly of the North American invasive Asteraceae, Tanacetum vulgare (common tansy)

Common tansy (Tanacetum vulgare), a significant agricultural invasive species, threatens forage productivity, livestock carrying capacity, and plant biodiversity across North American agroecosystems. Despite being a close relative to the ornamental Asteraceae Chrysanthemum, T. vulgare genomic studies are severely lacking, but remain critical to the development of novel weed management tools and the investigation of plant invasion evolution. In this study, we performed the assembly and annotation of the diploid genome (2n=2x=18) of an invasive North American T. vulgare genotype using long-read PacBio HiFi and Iso-Seq sequencing technology. The assembled haplotype genome comprised 9 pseudomolecules with a total size of ~4.8 Gb and a scaffold N50 of 546 Mb. In addition, both the plastid and mitochondrial genome sequences were assembled. BUSCO analysis revealed a high overall genome completeness with 69,343 predicted protein-coding genes (AED<0.6: 98%). A high-quality North American invasive reference T. vulgare genome, complementing a recent one assembled for a native European accession, is critical for the investigation of plant invasion biology, genome evolution, and the future development of novel genetic biocontrol weed management technologies.

genomics↗

bioRxiv2025

STARR-CRAAVT: A platform to design cell type-specific regulatory elements for next-generation gene therapy

Precise control of transgene expression through novel enhancer-promoter combinations is a promising strategy for advancing gene therapy mediated by adeno-associated virus (AAV). We present STARR-CRAAVT, a novel STARR-Seq-derived platform to screen for enhancers in the AAV context. Using HepG2 and HaCaT cells as screening models, we applied STARR-CRAAVT for the identification of cell type-specific enhancers. We integrated epigenetic datasets into an in-silico library of putative HepG2 enhancers and captured corresponding fragments from genomic DNA. The fragments were processed into AAV libraries and applied to HepG2 and HaCaT cells. STARR-CRAAVT analysis revealed a selective activity of the libraries confirming the HepG2-directed in-silico design and the specificity of single enhancer-promoter combinations could be validated using luciferase reporter assays. In addition, we scrutinized the impact of key experimental parameters on enhancer identification and found that the used promoter type had significant influence on the ability of candidates to act as enhancers. Furthermore, switching the location of identified enhancers in reporter assays revealed that the level of enhancer activity is highly dependent on the position in the AAV genome. Taken together, our study yields novel insights into enhancer function and demonstrates that STARR-CRAAVT can be employed to identify cell type-specific enhancers, highlighting promoter preference and enhancer positioning as key considerations for enhancer screening campaigns.

genomics↗

bioRxiv2023

N-cadherin mechanosensing in ovarian follicles controls oocyte maturation and ovulation

The cell adhesion molecule N-cadherin (CDH2) is a membrane component of adherens junctions which regulates tissue morphogenesis and architecture. In the follicles of mammalian ovaries, N-cadherin adherens junctions are present between granulosa cells, cumulus cells and at the interface of cumulus cell transzonal projections and the oocyte. We demonstrate a mechanosensory role of N-cadherin integrating tissue structure and hormonal regulation of follicular morphogenic events including expansion of the cumulus oocyte complex (COC) matrix, oocyte maturation and ovulation. Two small molecule N-cadherin antagonists inhibited COC maturation in vitro. Transcriptome profiling revealed that targets of {beta}-catenin and YAP1 pathways were dysregulated by N-cadherin antagonists. In vivo, N-cadherin antagonist significantly reduced ovulation in mice compared to controls (11 vs 26 oocytes/ovary; p=5.8x10-6). Ovarian follicles exhibited structural dysgenesis with granulosa and cumulus cell layers becoming disorganised and the connection between cumulus cells and the oocyte disrupted and the transcriptome again indicated altered mechanical sensing causing dysregulation of the Hippo/YAP and {beta}-catenin pathways and ECM reorganization. Granulosa specific N-cadherin depletion in Cdh2Fl/FL;Amhr2-Cre also showed significantly altered mechanosensitive gene expression and reduced ovulation. Our findings demonstrate a critical role for N-cadherin in ovarian follicular development and ovulation, and the potential to inhibit ovulation through targeting this signalling mechanism.

cell biology↗

bioRxiv2022

Simulated tempering-enhanced umbrella sampling improves convergence of free energy calculations of drug membrane permeation

Molecular dynamics simulations have been widely used to study solute permeation across biological membranes. The potential of mean force (PMF) for solute permeation is typically computed using enhanced sampling techniques such as umbrella sampling (US). For bulky drug-like permeants, however, obtaining converged PMFs remains challenging and often requires long simulation times, resulting in an unacceptable computational cost. Here, we augmented US with simulated tempering, introducing Simulated Tempering-enhanced US (STeUS), to improve the convergence of PMF calculations for the permeation of methanol and three common drug molecules. Simulate tempering helps to enhance sampling by varying the temperature of the system along a pre-defined temperature ladder. To obtain sufficient sampling of the umbrella histograms, which were computed only from the ground temperature, we modified the simulation time fraction spent at the ground temperature between 1/K and 50%, where K is the number of ST temperature states. We found that STeUS accelerates convergence compared to standard US, and the benefit of STeUS is system-dependent. For bulky molecules, for which standard US poorly converged, the application of ST was highly successful, leading to a more than five-fold accelerated convergence of the PMFs. For the small methanol solute, for which conventional US converges moderately, the application of ST is only beneficial if 50% of the STeUS simulation time is spent at the ground temperature. This study establishes STeUS as an efficient and simple method for PMF calculations, thereby strongly reducing the computational cost of routine high-throughput studies of drug permeability.

biophysics↗

bioRxiv2022

Plastid DNA sequences and oospore characters of some European species of Tolypella section Tolypella (Obtusifolia, Characeae) indicate a new cryptic Tolypella species from the Mediterranean island Sardinia

In Europe, the genus Tolypella (Characeae) comprises four to eight Tolypella species in sections Rothia and Tolypella that have been distinguished by vegetative morphology and gametangial characters such as antheridial size and oospore cell wall ornamentation. However, morphological species differentiation is difficult in some cases due to overlapping and variable vegetative features, which in many cases are difficult to observe clearly. To clarify the taxonomic status of the five European species of Tolypella in section Tolypella, sequence data of the plastid genes atpB, rbcL and psbC for Tolypella glomerata (Desv.) Leonh., Tolypella hispanica Allen, Tolypella nidifica (O.F. Mull.) A. Braun, Tolypella normaniana (Nordst.) Nordst. and Tolypella salina Cor. were combined with data on oospore morphology, including oospore wall ornamentation. Gene sequence data identified five distinct clusters, but they differed from the morphologically identified five species. T. glomerata consisted of some of the samples morphologically identified as T. glomerata and seven samples of T. normaniana, while the remaining T. glomerata samples clustered with specimens of unclear affiliation ("Tolypella. sp."). T. hispanica I consisted of samples from various locations, whereas "T. hispanica II" consisted of samples of T. hispanica from the Mediterranean island, Sardinia. The remaining cluster consisted of all the specimens that had been determined as T. salina or T. nidifica in addition to two specimens of T. normaniana. Oospore morphology was most clearly distinguishable for T. glomerata. Oospore characteristics for all other taxa were not as informative but showed some geographical and/or environmentally influenced differences, especially for T. nidifica and T. salina. Our results suggest a significantly different taxonomy of Tolypella sect. Tolypella in which specimens normally identified as T. glomerata might be two different species, T. glomerata and an unidentified species; T. nidifica and T. salina are not separate species; T. normaniana is a diminutive variant of T. nidifica or T. salina; and T. hispanica comprises two different species, one from the Mediterranean island Sardnia.

plant biology↗

bioRxiv2021

High-throughput identification of RNA localization elements reveals a regulatory role for A/G rich sequences

Hundreds of RNAs are enriched in the projections of neuronal cells. For the vast majority of them, though, the sequence elements within them that regulate their localization are unknown. To identify RNA elements capable of directing transcripts to neurites, we designed and deployed a massively parallel reporter assay that tested the localization regulatory ability of thousands of sequence fragments drawn from endogenous mouse 3' UTRs. We identified peaks of regulatory activity within several 3' UTRs and found that sequences derived from these peaks were both necessary and sufficient for RNA localization to neurites in mouse cells as well as active in human neurons. The active localization elements were enriched in adenosine and guanosine residues and were tens to hundreds of nucleotides long as shortening of the elements led to significantly reduced activity. Using RNA affinity and mass spectrometry, we found that the RNA-binding protein Unk was associated with the active localization elements. Depletion of Unk in cells reduced the neurite-enrichment of the elements, indicating a functional requirement for Unk in their trafficking. These results provide a framework for the unbiased, high-throughput identification of RNA elements and mechanisms that govern transcript localization.

molecular biology↗

bioRxiv2021

Exploration and analysis of molecularly annotated, 3D models of breast cancer at single-cell resolution using virtual reality

A set of increasingly powerful approaches are enabling spatially resolved measurements of growing numbers of molecular features in biological samples. While important insights can be derived from the two-dimensional data that many of these technologies generate, it is clear that extending these approaches into the third and fourth dimensions will magnify their impact. Realizing biological insights from datasets where thousands to millions of cells are annotated with tens to hundreds of parameters in space will require the development of new computational and visualization strategies. Here, we describe Theia, a virtual reality-based platform, which enables exploration and analysis of either volumetric or segmented, molecularly-annotated, three-dimensional datasets, with the option to extend the analysis to time-series data. We also describe our pipeline for generating annotated 3D models of breast cancer and supply several datasets to enable users to explore the utility of Theia for understanding cancer biology in three dimensions.

cancer biology↗

bioRxiv2021

Alternative splicing of pericentrin contributes to cell cycle control in cardiomyocytes

Induction of cardiomyocyte proliferation is a promising option to regenerate the heart. Thus, it is important to elucidate mechanisms that contribute to the cell cycle arrest of mammalian cardiomyocytes. Here, we assessed the contribution of the pericentrin (Pcnt) S isoform to the cell cycle arrest in postnatal cardiomyocytes. Immunofluorescence staining of Pcnt isoforms combined with siRNA-mediated depletion indicates that Pcnt S preferentially localizes to the nuclear envelope, while the Pcnt B isoform is enriched at centrosomes. This is further supported by the localization of ectopically expressed FLAG-tagged Pcnt S and Pcnt B in postnatal cardiomyocytes. Analysis of centriole configuration upon Pcnt depletion revealed that Pcnt B but not Pcnt S is required for centriole cohesion. Importantly, ectopic expression of Pcnt S induced centriole splitting in a heterologous system, ARPE-19 cells, and was sufficient to impair DNA synthesis in C2C12 myoblasts. Moreover, Pcnt S depletion enhanced serum-induced cell cycle re-entry in postnatal cardiomyocytes. Analysis of mitosis, binucleation rate, and cell number suggests that Pcnt S depletion promotes progression of postnatal cardiomyocytes through the cell cycle resulting in cell division. Collectively, our data indicate that alternative splicing of Pcnt contributes to the establishment of cardiomyocyte cell cycle arrest shortly after birth.

cell biology↗

bioRxiv2021

Generation and first characterization of TRDC-Knockout pigs lacking γδ T cells

The TRDC-Locus encodes the T cell receptor delta constant region, one component of the {gamma}{delta} T cell receptor which is essential for development of {gamma}{delta} T cells. In contrast to peptide recognition by {beta} T cells, antigens activating {gamma}{delta} T cells are mostly MHC independent and not well characterized. Therefore, the function of {gamma}{delta} T cells and their contribution to protection against infections is still unclear. Higher numbers of circulating {gamma}{delta} T cells compared to mice, render the pig a suitable animal model to study {gamma}{delta} T cells. Knocking-out the porcine TRDC-locus by intracytoplasmic microinjection and somatic cell nuclear transfer resulted in healthy living {gamma}{delta} T cell deficient offspring. Flow cytometric analysis revealed that TRDC-KO pigs lack {gamma}{delta} T cells in peripheral blood mononuclear cells (PBMC) and spleen cells. The composition of the remaining leucocyte subpopulations was not affected by the depletion of {gamma}{delta} T cells. Genome-wide transcriptome analyses in PBMC revealed a pattern of changes reflecting the impairment of known or expected {gamma}{delta} T cell dependent pathways. Histopathology did not reveal developmental abnormalities of secondary lymphoid tissues. However, in a vaccination experiment the KO pigs stayed healthy but had a significantly lower neutralizing antibody titer as the syngenic controls.

immunology↗

bioRxiv2021

Hierarchical cross-scale analysis identifies parallel ventral striatal networks coding for dynamic and stabilized olfactory reward predictions

The unbiased identification of brain circuits responsible for behavior and their local cellular computations is a challenge for neuroscience. We establish here a hierarchical cross-scale approach from behavioral modeling and fMRI in task-performing mice to cellular network dynamics to identify how reward predictions are represented in the forebrain upon olfactory conditioning. fMRI identified functional segregation in reward prediction and error computations among olfactory cortices and subcortical circuits. Among them, the olfactory tubercle contributed both to dynamic reward predictions and prediction error. In this region, cellular recordings revealed two parallel neuronal populations for prediction coding. One population produced stabilized predictions as distributed stimulus-bound transient network activity; the other evolved during anticipatory waiting and fully reflected predicted value in single-units, dynamically integrating the recent cue-specific history of uncertain outcomes. Thus, the cross-scale approach revealed regional functional differentiation among the distributed forebrain circuits with a limbic hotspot for multiple non-redundant reward prediction coding.

neuroscience↗

bioRxiv2020

Myogenin controls via AKAP6 non-centrosomal microtubule organizing center formation at the nuclear envelope

Non-centrosomal microtubule organizing centers (MTOC) are pivotal for the function of multiple cell types, but the processes initiating their formation are unknown. Here, we find that the transcription factor myogenin is required in myoblasts for the localization of MTOC proteins to the nuclear envelope. Moreover, myogenin is sufficient in fibroblasts for nuclear envelope MTOC (NE-MTOC) formation and centrosome attenuation. Bioinformatics combined with loss- and gain-of-function experiments identified induction of AKAP6 expression as one central mechanism for myogenin-mediated NE-MTOC formation. Promoter studies indicate that myogenin preferentially induces the transcription of muscle- and NE-MTOC-specific isoforms of Akap6 and Syne1, which encodes nesprin-1, the NE-MTOC anchor protein in muscle cells. Overexpression of AKAP6{beta} and nesprin-1 was sufficient to recruit endogenous MTOC proteins to the nuclear envelope of myoblasts in the absence of myogenin. Taken together, our results illuminate how mammals transcriptionally control the switch from a centrosomal MTOC to an NE-MTOC and identify AKAP6 as a novel NE-MTOC component in muscle cells.

cell biology↗

bioRxiv2019

The Noise-Resilient Brain: Resting-State Oscillatory Activity Predicts Words-In-Noise Recognition

The role of neuronal oscillations in the processing of speech has recently come to prominence. Since resting-state (RS) brain activity has been shown to predict both task-related brain activation and behavioural performance, we set out to establish whether inter-individual differences in spectrally-resolved RS-MEG power are associated with variations in words-in-noise recognition in a sample of 88 participants made available by the Human Connectome Project. Positive associations with resilience to noise were observed with power in the range 21 and 29Hz in a number of areas along the left temporal gyrus and temporo-parietal association areas peaking in left posterior superior temporal gyrus (pSTG). Significant associations were also found in the right posterior superior temporal gyrus in the frequency range 30 to 40Hz. We propose that individual differences in words-in-noise performance are related to baseline excitability levels of the neural substrates of phonological processing. HighlightsO_LIPower of resting MEG activity predicts Words-In-Noise recognition performance C_LIO_LISignificant associations in higher beta and lower gamma frequency band C_LIO_LIStrongest in left-lateralised perisylvian cluster peaking in posterior STG C_LIO_LIEffects are spectrally and spatially consistent with phoneme-level processing C_LI

neuroscience↗

bioRxiv2019

Resolving the connectome - Spectrally-specific functional connectivity networks and their distinct contributions to behaviour

The resting brain exhibits spontaneous patterns of activity that reflect features of the underlying neural substrate. Examination of inter-areal coupling of resting state oscillatory activity has revealed that the brains resting activity is composed of functional networks, whose topographies differ depending upon oscillatory frequency, suggesting a role for carrier frequency as a means of creating multiplexed, or functionally segregated, communication channels between brain areas. Using canonical correlation analysis, we examined spectrally resolved resting-state connectivity patterns derived from MEG recordings to determine the relationship between connectivity intrinsic to different frequency channels and a battery of over a hundred behavioural and demographic indicators, in a group of 89 young healthy participants. We demonstrate that each of the classical frequency bands in the range 1-40Hz (delta, theta, alpha, beta and gamma) delineates a subnetwork that is behaviourally relevant, spatially distinct, and whose expression is either negatively or positively predictive of individual traits, with the strongest link in the alpha band being negative and networks oscillating at different frequencies, such as theta, beta and gamma carrying positive function.

neuroscience↗

bioRxiv2019

The knockout of the HMG-Domain of the porcine SRY-gene by CRISPR/Cas RNP microinjection causes sex reversal in gene-edited pigs

1The sex-determining region on the Y chromosome (SRY) is thought to be the central genetic element of male sex development. Mutations within the SRY gene are associated with a male-to-female sex reversal syndrome in humans and other mammalian species such as mice and rabbits. However, the underlying mechanisms are largely unknown. To understand the biological function of the SRY gene, a site-directed mutational analysis is required to investigate associated phenotypic changes at the molecular, cellular and morphological level. In our study, we successfully generated a knockout of the porcine SRY gene by microinjection of two clustered regularly interspaced short palindromic repeats (CRISPR) - associated protein - 9 nuclease (Cas9) ribonucleoprotein (RNP) complexes targeting the centrally located "high mobility group" (HMG) domain of the SRY gene. Mutations within this region resulted in the development of complete external and internal female genitalia in genetically male pigs. The internal female genitalia including uteri, ovaries, and oviducts, revealed substantial size differences in 9-months old SRY-knockout pigs compared to age-matched female wild type controls. In contrast, a deletion within the 5 flanking region of the HMG domain was not associated with sex reversal. Results of this study demonstrates for the first time the central role of the HMG domain of the SRY gene in male sex determination in pigs. Moreover, quantitative analysis by digital PCR revealed evidence for a duplication of the porcine SRY locus. Our results pave the way towards the generation of boars exclusively producing phenotypically female offspring to avoid surgical castration without anesthesia in piglets. Moreover, the study establishes a large animal model that is much more similar to humans in regard of physiology and anatomy and pivotal for longitudinal studies.

developmental biology↗

bioRxiv2018

Transient spectral events in resting state MEG predict individual time-frequency task responses

Even in response to apparently simple tasks such as hand moving, human brain activity shows remarkable inter-subject variability. Presumably, this variability reflects genuine behavioural or functional variability. Recently, spatial variability of resting-state features in fMRI - specifically connectivity - has been shown to explain (spatial) task-response variability. Such a link, however, is still missing for M/EEG data and its spectrally rich structure. At the same time, it has recently been shown that task responses in M/EEG can be well represented using transient spectral events bursting at fast time scales. Here, we show that individual differences in the spatio-spectral structure of M/EEG task responses, can, to a reasonable degree, be predicted from individual differences in transient spectral events identified at rest. In a MEG dataset of diverse task conditions (including motor responses, working memory and language comprehension tasks) and resting-state sessions for each subject (n = 89), we used Hidden-Markov-Modelling to identify transient spectral events as a feature set to learn the mapping of space-time-frequency content from rest to task. Resulting trial-averaged, subject-specific task-response predictions were then compared with the actual task responses in left-out subjects. All task conditions were predicted significantly above chance. Furthermore, we observed a systematic relationship between genetic similarity (e.g. unrelated subjects vs. twins) and predictability. These findings support the idea that subject-specific transient spectral events in resting-state neural activity are linked to, and predictive of, subject-specific trial-averaged task responses in a wide range of experimental conditions.

neuroscience↗

bioRxiv2018

Motor learning shapes temporal activity in human sensorimotor cortex

Although neuroimaging techniques have provided vital insights into the anatomical regions involved in motor learning, the underlying changes in temporal dynamics are not well understood. Using magnetoencephalography and Hidden Markov Modelling to model the dynamics of neural oscillations on data-adaptive time-scales, we detected specific changes in movement-related sensorimotor {beta}-activity during practice of a self-paced sequential visuo-motor task. The behaviourally-relevant neural signature generalised to another motor task, emphasising the centrality of {beta}-activity in motor plasticity.

neuroscience↗