Biology subjects

Weidemann, D. E.

Publications and source records attributed to Weidemann, D. E..

5 recordsLinked to original sources

bioRxiv2026

Establishing MS2-MCP-based single-molecule RNA visualization in Schizosaccharomyces pombe

Single-molecule RNA imaging using the MS2-MCP system has transformed the study of RNA biology across model organisms. However, this technology has remained unavailable for fission yeast (Schizosaccharomyces pombe), even though fission yeast is a central model for eukaryotic gene expression. Achieving single-molecule sensitivity requires identifying a narrow optimum where RNA labels are sufficiently bright while background fluorescence remains minimal. We have now accomplished this for S. pombe by systematically optimizing MCP expression and localization--screening a panel of constitutive S. pombe promoters and evaluating combinations of nuclear localization and export signals (NLSs and NESs). The resulting, successful constructs use tandem StayGold as the MCP fluorescent tag, taking advantage of its superior photostability. Together with optimized vectors for MS2 stem-loop tagging of endogenous transcripts, these tools enable single-molecule RNA imaging in fission yeast, opening the door to quantitative analyses of RNA dynamics in this core genetic model. Summary statementWe establish single-molecule RNA visualization with MS2/MCP for fission yeast, S. pombe, an important model organism to study RNA biology.

molecular biology↗

bioRxiv2023

A comparative study of algorithms detecting differential rhythmicity in transcriptomic data

Rhythmic transcripts play pivotal roles in driving the daily oscillations of various biological processes. Genetic or environmental disruptions can lead to alterations in the rhythmicity of transcripts, ultimately impacting downstream circadian outputs, including metabolic processes and even behavior. To statistically compare the differences in transcript rhythms between two or more conditions, several algorithms have been developed to analyze circadian transcriptomic data, each with distinct features. In this study, we compared the performance of seven algorithms that were specifically designed to detect differential rhythmicity. We found that even when applying the same statistical threshold, these algorithms yielded varying numbers of differentially rhythmic transcripts. Nevertheless, the set of transcripts commonly identified as differentially rhythmic exhibited substantial overlap among algorithms. Furthermore, the phase and amplitude differences calculated by these algorithms displayed significant correlations. In summary, our study highlights a high degree of similarity in the results produced by these algorithms. Furthermore, when selecting an algorithm for analysis, it is crucial to ensure the compatibility of input data with the specific requirements of the chosen algorithm and to assess whether the algorithms output fits the needs of the user.

bioinformatics↗

bioRxiv2023

Coordination of rhythmic RNA synthesis and degradation orchestrates 24-hour and 12-hour RNA expression patterns in mouse fibroblasts

Circadian RNA expression is essential to ultimately regulate a plethora of downstream rhythmic biochemical, physiological, and behavioral processes. Both transcriptional and post transcriptional mechanisms are considered important to drive rhythmic RNA expression, however, the extent to which each regulatory process contributes to the rhythmic RNA expression remains controversial. To systematically address this, we monitored RNA dynamics using metabolic RNA labeling technology during a circadian cycle in mouse fibroblasts. We find that rhythmic RNA synthesis is the primary contributor of 24 hr RNA rhythms, while rhythmic degradation is more important for 12 hr RNA rhythms. These rhythms were predominantly regulated by Bmal1 and/or the core clock mechanism, and interplay between rhythmic synthesis and degradation has a significant impact in shaping rhythmic RNA expression patterns. Interestingly, core clock RNAs are regulated by multiple rhythmic processes and have the highest amplitude of synthesis and degradation, presumably critical to sustain robust rhythmicity of cell-autonomous circadian rhythms. Our study yields invaluable insights into the temporal dynamics of both 24 hr and 12 hr RNA rhythms in mouse fibroblasts.

molecular biology↗

bioRxiv2023

The minimal intrinsic stochasticity of constitutively expressed eukaryotic genes is sub-Poissonian

Stochastic variation in gene products ("noise") is an inescapable by-product of gene expression. Noise must be minimized to allow for the reliable execution of cellular functions. However, noise cannot be suppressed beyond an intrinsic lower limit. For constitutively expressed genes, this limit is believed to be Poissonian, meaning that the variance in mRNA numbers cannot be lower than their mean. Here, we show that several cell division genes in fission yeast have mRNA variances significantly below this limit, which cannot be explained by the classical gene expression model for low-noise genes. Our analysis reveals that multiple steps in both transcription and mRNA degradation are essential to explain this sub-Poissonian variance. The sub-Poissonian regime differs qualitatively from previously characterized noise regimes, a hallmark being that cytoplasmic noise is reduced when the mRNA export rate increases. Our study re-defines the lower limit of eukaryotic gene expression noise and identifies molecular requirements for ultra-low noise which are expected to support essential cell functions.

molecular biology↗

bioRxiv2021

Mitotic checkpoint gene expression is tuned by coding sequences

The mitotic checkpoint (also called spindle assembly checkpoint, SAC) is a signaling pathway that safeguards proper chromosome segregation. Proper functioning of the SAC depends on adequate protein concentrations and appropriate stoichiometries between SAC proteins. Yet very little is known about SAC gene expression. Here, we show in fission yeast (S. pombe) that a combination of short mRNA half-lives and long protein half-lives supports stable SAC protein levels. For the SAC genes mad2+ and mad3+, their short mRNA half-lives are supported by a high frequency of non-optimal codons. In contrast, mad1+ mRNA has a short half-life despite a low frequency of non-optimal codons and despite the lack of known destabilizing motifs. Hence, different SAC genes employ different strategies of expression. We further show that Mad1 homodimers form co-translationally, which may necessitate a certain codon usage pattern. Taken together, we propose that the codon usage of SAC genes is fine-tuned for proper SAC function. Our work shines light on gene expression features that promote spindle assembly checkpoint function and suggests that synonymous mutations may weaken the checkpoint.

cell biology↗