Biology subjects

Reinhardt, M.

Publications and source records attributed to Reinhardt, M..

6 recordsLinked to original sources

bioRxiv2026

A structured RNA balances DEAD-box RNA helicase function in plant alternative splicing control

Eukaryotic gene expression is a multi-layered process influenced by multiple factors. One of them is the secondary structure of precursor mRNAs that can impact various aspects of their processing including alternative splicing (AS). Here, we report the functional characterization of the conserved RNA structural element DEAD that is located in DEAD-box RNA helicase (DRH) genes from land plants and serves as a sensor for RNA helicase activity by controlling AS. In Arabidopsis thaliana, it is found in DRH1 and its closest paralog, regulating usage of an alternative splice site as part of a negative feedback loop. Accordingly, opening of the structure shifts splicing towards non-coding variants, thereby balancing transcript and protein levels. Interestingly, the system is specific to DRH1 and its paralog and does not react to related helicases, which is at least partially conferred by the disordered and RGG/RG motif-containing C-terminus of DRH1. The importance of DEAD is underlined by the observation that releasing this attenuation mechanism causes massive changes in AS - mainly intron retention and exon skipping - and gene expression and results in a severe stress phenotype. Thus, DEAD provides a critical buffering mechanism to fine-tune helicase levels and their global impact on RNA structure-responsive gene expression.

plant biology↗

bioRxiv2025

Loss of noradrenergic Fkbp5 disrupts social behavior and norepinephrine dynamics in the basolateral amygdala

Social dysfunction is common in depression and varies with stress exposure and genetic risk. The current study identifies a cell-type specific role for Fkbp5, a glucocorticoid receptor co-chaperone, in noradrenergic neurons engaged during social stress. Acute social stress upregulated Fkbp5 in the locus coeruleus (LC), whereas repeated exposure attenuated this effect. Noradrenergic Fkbp5 deletion (Fkbp5Nat) increased pro-social behavior exclusively in male mice. In the basolateral amygdala (BLA), social interaction reduced norepinephrine (NE) turnover in wild-type but not Fkbp5Nat mice. Consistently, proteomics revealed mitochondrial/energy and synapse-related remodeling in BLA neurons. Miniscope imaging showed that behavior-locked NE transients in BLA were selectively blunted in Fkbp5Nat mice during interaction with outbred CD1 conspecifics, while same-strain C57BL/6N encounters preserved NE dynamics. Together, this study indicates that Fkbp5 tunes LC-BLA output to social salience in a sex- and context-dependent manner, suggesting a circuit-specific route to normalize social salience without broadly suppressing noradrenergic function.

neuroscience↗

bioRxiv2025

The structured mRNA element 45ABC mediates auto- and cross-regulation of RBP45 genes via alternative splicing

Alternative splicing (AS) is a common gene regulatory mechanism involving distinct interactions between trans-acting factors and cis-regulatory elements on the precursor mRNA (pre-mRNA). In this study, we have functionally characterized the structured motif 45ABC, which is located in the pre-mRNAs of RNA-binding protein (RBP) 45 genes in many plant species. Our data revealed that this element mediates a negative auto- and cross-regulatory feedback loop via AS of the three 45ABC-containing RBP45 genes in Arabidopsis thaliana. We identified a G-rich stretch within the first stem as a potential RBP45 binding site and observed increased RBP45-dependent AS upon structural weakening of this pairing element. The second stem includes the alternative 5 splice site being activated in the presence of RBP45. Based on the known interaction between RBP45 homologs and U1 snRNP components required for 5 splice site recognition, we propose that RBP45 binding to stem I of 45ABC induces usage of the alternative 5 splice site in stem II. The resulting splicing variant is unproductive, thereby diminishing RBP45 expression. Analysing the splicing regulatory impact of the three At-RBP45 genes in auto- and cross-regulation and a transcriptome-wide manner revealed unequal genetic redundance with a major role of RBP45B. Furthermore, phenotypical analysis of single and higher order rbp45 mutants pointed at these genes functions in controlling primary root growth and flowering time. Taken together, we demonstrated that both sequence and structural features of 45ABC are critical for proper splicing control, balancing RBP45 expression and functions in plants via a conserved mRNA motif. Significance statementFunctional characterisation of a structured mRNA motif present in plant RBP45 genes identified sequence and pairing elements underlying a negative auto- and cross-regulatory expression circuit on the level of alternative splicing. Our study provides a rationale for the evolutionary conservation of this RNA element, which allows balancing levels and functions of RBP45 proteins as a requirement for proper plant development.

plant biology↗

bioRxiv2025

Within-host evolution of drug tolerance in Mycobacterium tuberculosis.

SynopsisO_ST_ABSBackgroundC_ST_ABSMycobacterium tuberculosis (Mtb) causes tuberculosis (TB) in humans. Poor treatment responses are a threat to global TB control, as such, understanding contributing factors to poor responses is important. We hypothesized that antibiotic tolerance could contribute to delayed culture conversion (recalcitrant TB), and resistance amplification in patients during TB treatment. ObjectivesTo investigate the role of drug tolerance in delayed culture conversion and resistance amplification in TB patients. MethodsWe collected serial Mtb isolates from i) patients with drug-susceptible TB who remained culture positive for up to 6 years (i.e. recalcitrant TB), and ii) patients with multidrug-resistant TB (MDR-TB) where resistance amplified during treatment. We measured tolerance to rifampicin (RIF) in drug-susceptible TB strains and tolerance to moxifloxacin (MFX) in MDR-TB strains using a real-time time-kill assay. ResultsRIF tolerance evolved within-host, increasing up to and ~1.5-fold, however, there was no apparent contribution of RIF tolerance to delayed culture conversion. Tolerance to Mfx in MDR-TB patients appeared negatively associated with resistance amplification and consistently decreased over time in patients. ConclusionOur findings confirm that antibiotic tolerance evolves in Mtb within patients over time during treatment. However, there was no evidence that this tolerance influences treatment responses, calling for further investigation of contributors to adverse treatment responses and their mitigation.

microbiology↗

bioRxiv2025

Variability in intrinsic drug tolerance in Mycobacterium tuberculosis corresponds with phylogenetic lineage

Drug tolerance allows bacteria to survive extended exposure to bactericidal drugs and is thought to play a role in drug resistance evolution. In Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB), multidrug resistant TB (MDR-TB) outbreaks are frequently caused by strains belonging to two phylogenetic lineages of the human-adapted strains of the Mtb Complex, namely lineages (L) 2 and L4. We hypothesized that members of L2 and L4 are more intrinsically drug tolerant, and as such, more readily evolve drug resistance. To explore this, we devised a high throughput in vitro assay to measure drug tolerance in Mtb. We selected a cohort of strains representative of the globally most frequent lineages L1 - L4. We measured tolerance to rifampicin and bedaquiline and found L3 and L4 strains to have higher tolerance compared to L1 and L2 strains. In addition, phylogenetically closely related strains exhibited similar levels of tolerance, suggesting that tolerance is heritable. Finally, we explored genes previously reported to be associated with tolerance in Mtb and found significant enrichment in mutations in genes involved in cell wall and cell processes, intermediary metabolism and respiration, as well as lipid metabolism in high-tolerance strains.

microbiology↗

bioRxiv2022

Automatically annotated motion tracking identifies a distinct social behavioral profile following chronic social defeat stress

Severe stress exposure is a global problem with long-lasting negative behavioral and physiological consequences, increasing the risk of stress-related disorders such as major depressive disorder (MDD). An essential characteristic of MDD is the impairment of social functioning and lack of social motivation. Chronic social defeat stress is an established animal model for MDD research, which induces a cascade of physiological and social behavioral changes. The current developments of markerless pose estimation tools allow for more complex and socially relevant behavioral tests, but the application of these tools to social behavior remains to be explored. Here, we introduce the open-source tool "DeepOF" to investigate the individual and social behavioral profile in mice by providing supervised and unsupervised pipelines using DeepLabCut annotated pose estimation data. The supervised pipeline relies on pre-trained classifiers to detect defined traits for both single and dyadic animal behavior. Subsequently, the unsupervised pipeline explores the behavioral repertoire of the animals without label priming, which has the potential of pointing towards previously unrecognized motion motifs that are systematically different across conditions. We here provide evidence that the DeepOF supervised and unsupervised pipelines detect a distinct stress-induced social behavioral pattern, which was particularly observed at the beginning of a novel social encounter. The stress-induced social behavior shows a state of arousal that fades with time due to habituation. In addition, while the classical social avoidance task does identify the stress-induced social behavioral differences, both DeepOF behavioral pipelines provide a clearer and more detailed profile. DeepOF aims to facilitate reproducibility and unification of behavioral classification of social behavior by providing an open-source tool, which can significantly advance the study of rodent individual and social behavior, thereby enabling novel biological insights as well as drug development for psychiatric disorders.

neuroscience↗