Biology subjects

Schuhmann, A.

Publications and source records attributed to Schuhmann, A..

4 recordsLinked to original sources

bioRxiv2025

Absolute Quantification of Fluorescent Protein Fusions by Proteomics

Fusions with fluorescence proteins (FPs) play a pivotal role in experimental biology because of their sensitive and spatially precise visualization by spectroscopy. However, observed fluorescence is not always proportional to their molar abundance. Only a fraction of the fusion protein containing the mature fluorescence chromophore is detectable by spectroscopy and there is no generic method for estimating its molar abundance. We developed a fluorescence-independent mass-spectrometry based approach for accurate absolute sub-femtomole quantification of FP-fusions that also estimates the protein faction having fully matured chromophore. The method exploits an isotopically labelled 68 kDa recombinant protein standard containing peptide proxies for 6 prototypical FPs (mCherry, mScarlet-I, mKate2, EGFP, mNeonGreen and Dendra2) and two self-labelling (Halo- and SNAP-) tags. It enables quantification of proteins fused to any of more than 70 FPs or self-labelling tags. It is versatile, robust, precise and can be used broadly for the absolute quantification of fluorescent fusions in in-vivo and in-vitro cellular systems. As case study we combined mass spectrometry with fluorescence spectroscopy to study expression kinetics of FP fusions in cell-free systems. Molar concentrations of the expressed fusion protein, its fraction containing the mature fluorescent chromophore and of the fluorescent protein were integrated into a mathematical model to obtain kinetic rates of translation, chromophore maturation and folding.

biochemistry↗

bioRxiv2024

A comparative analysis of planarian regeneration specificity reveals tissue polarity contributions of the axial cWnt signalling gradient.

Planarians exhibit remarkable whole-body regeneration abilities. The formation of heads at forward-facing wounds and tails at rearward-facing wounds suggests an intrinsic tissue polarity guiding regeneration. While the underlying mechanisms remain unclear, reports of double-headed regenerates from increasingly narrow tissue fragments have long been hypothesised to reflect gradient-based polarity specification. Here, we systematically re-examine this hypothesis in the modern model species Schmidtea mediterranea and a representative of the genus likely used in the original studies, Girardia sinensis. While we never observed double-heads in S. mediterranea, G. sinensis readily regenerated double-heads in a manner dependent on piece length, anatomical position and body size. We found that the reduced regeneration robustness of G. sinensis was partially explained by wound site-symmetric expression of the head determinant notum, which is highly anterior-specific in S. mediterranea. Exploring what else might mediate head/tail regeneration specificity in G. sinensis, we examined the role of the conserved tail-to-head cWnt signalling gradient. By developing a time-resolved pharmacological approach to reduce the cWnt gradient slope without affecting wound-induced cWnt signalling dynamics, we observed an increased incidence of double-headed regenerates. In addition, the body size-dependence of double-head formation correlated with the decreasing steepness of the cWnt gradient due to scaling. Taken together, our results indicate that the slope of the cWnt gradient may contribute to planarian head/tail regeneration specificity. Furthermore, they suggest that planarian tissue polarity is composed of multiple parallely-acting polarity cues, the differential reliance on which contributes to the observed interspecies variation in regeneration specificity.

developmental biology↗

bioRxiv2024

A multi-year longitudinal harmonization of quality controls in mass spectrometry proteomics core facilities

Quality control procedures play a pivotal role in ensuring the reliability and consistency of data generated in mass spectrometry-based proteomics laboratories. However, the lack of standardized quality control practices across laboratories poses challenges for data comparability and reproducibility. In response, we conducted a harmonization study within proteomics laboratories of the Core for Life alliance with the aim to establish a common quality control framework, which facilitates comprehensive quality assessment and identification of potential sources of performance drift. Through collaborative efforts, we developed a consensus quality control standard for longitudinal assessment and we adopted a common processing software. We generated a 4-year longitudinal dataset from multiple instruments and laboratories, which enabled us to assess intra- and inter-laboratory variability, to identify causes of performance drift, and to establish community reference values for several quality control parameters. Our study enhances data comparability and reliability and fosters a culture of collaboration and continuous improvement within the proteomics community to ensure the integrity of proteomics data.

biochemistry↗

bioRxiv2018

Fine Endmesolithic Fish Caviar Meal Discovered by Proteomics of Foodcrusts

The role of aquatic resources in ancient economies and paleodiet is important for understanding the evolution of prehistorical societies. However, conventional archaeometric approaches lack organismal specificity, are affected by abundant environmental contaminants and do not elucidate food processing recipes. We performed proteomics analysis of charred organic deposits adhered on early ceramics from Mesolithic-Neolithic inland site Friesack 4 (Brandenburg, Germany). Proteomics of foodcrust from a vessel attributed to Endmesolithic pottery identified fine carp roe meal and revealed details of a prehistorical culinary recipe. We propose that Endmesolithic occupants of Friesack at the end of the 5th millennium BC utilized fish as a food reserve and adopted delayed-return subsistence strategy. These data contribute to better understanding of the dietary context of Neolithic transition in European inland.

paleontology↗