Biology subjects

Bonn, F.

Publications and source records attributed to Bonn, F..

4 recordsLinked to original sources

bioRxiv2026

High Resolution Multi-Pass Astral Analyzer Quantification Enables Highly Multiplexed 35-Plex Tandem Mass Tag Proteomics

Tandem mass tags (TMT) allow highly multiplexed and thus high-throughput, precisely quantitative proteomic analysis. Incorporation of additional deuterated reporter channels has near-doubled the multiplexation achieved with Thermo Scientific TMTpro reagents from 18 to 35-plex but requires extremely high [~]100k analyzer resolving power at m/z 128 to differentiate and quantify reporter ion channels, far beyond any single reflection time-of-flight analyzer, and exceeding the multi-reflection Thermo Scientific Astral analyzer in its standard operation. A multi-pass mode of Astral operation has been developed for the Thermo Scientific Orbitrap Astral Zoom mass spectrometer that triples the ion path to 90 m, more than doubling resolving power for a narrow m/z range. This "TMT HR mode" has been integrated into a new method of TMT proteomic analysis that splits regular MS2 analysis of labeled peptides into paired measurements comprising wide mass range scans for peptide identification, and TMT HR mode scans for reporter ion quantification. The method has been shown to accurately quantify 32-plex labeled HeLa protein lysate and provide far greater depth of analysis as state-of-the-art Orbitrap-only methods, while analysis of 11-plex labeled yeast showed no analytical depth sacrificed vs regular Orbitrap Astral TMT analysis. Further comparative measurements of a 2-cell line 35-plex sample demonstrated greater analytical depth, and similar quantitative precision, to "gold standard" Orbitrap MS3 methods.

biochemistry↗

bioRxiv2020

Serine-ubiquitination regulates Golgi morphology and the secretory pathway upon Legionella infection

SidE family of Legionella effectors catalyze non-canonical phosphoribosyl-linked ubiquitination (PR-ubiquitination) of host proteins during bacterial infection. SdeA localizes predominantly to ER and partially to the Golgi apparatus, and mediates serine ubiquitination of multiple ER and Golgi proteins. Here we show that SdeA causes disruption of Golgi integrity due to its ubiquitin ligase activity. The Golgi linking proteins GRASP55 and GRASP65 are PR-ubiquitinated on multiple serine residues, thus preventing their ability to cluster and form oligomeric structures. In addition, we found that the functional consequence of Golgi disruption is not linked to the recruitment of Golgi membranes to the growing Legionella-containing vacuoles. Instead, it affects the secretory pathway, including cytokine release in cells. Taken together, our study sheds light on the Golgi manipulation strategy by which Legionella hijacks the secretory pathway and promotes bacterial infection.

cell biology↗

bioRxiv2020

Metabolic differences between symbiont subpopulations in the deep-sea tubeworm Riftia pachyptila

The hydrothermal vent tube worm Riftia pachyptila lives in intimate symbiosis with intracellular sulfur-oxidizing gammaproteobacteria. Although the symbiont population consists of a single 16S rRNA phylotype, bacteria in the same host animal exhibit a remarkable degree of metabolic diversity: They simultaneously utilize two carbon fixation pathways and various energy sources and electron acceptors. Whether these multiple metabolic routes are employed in the same symbiont cells, or rather in distinct symbiont subpopulations, was unclear. As Riftia symbionts vary considerably in cell size and shape, we enriched individual symbiont cell sizes by density gradient centrifugation in order to test whether symbiont cells of different sizes show different metabolic profiles. Metaproteomic analysis and statistical evaluation using clustering and random forests, supported by microscopy and flow cytometry, strongly suggest that Riftia symbiont cells of different sizes represent metabolically dissimilar stages of a physiological differentiation process: Small symbionts actively divide and may establish cellular symbiont-host interaction, as indicated by highest abundance of the cell division key protein FtsZ and highly abundant chaperones and porins in this initial phase. Large symbionts, on the other hand, apparently do not divide, but still replicate DNA, leading to DNA endoreduplication. Highest abundance of enzymes for CO2 fixation, carbon storage and biosynthesis in large symbionts indicates that in this late differentiation stage the symbionts metabolism is efficiently geared towards the production of organic material. We propose that this division of labor between smaller and larger symbionts benefits the productivity of the symbiosis as a whole.

microbiology↗

bioRxiv2019

Wss1 promotes replication stress tolerance by degrading histones.

Timely completion of DNA replication is central to accurate cell division and to the maintenance of genomic stability. However, certain DNA-protein interactions can physically impede DNA replication fork progression. Cells remove or bypass these physical impediments by different mechanisms to preserve DNA macromolecule integrity and genome stability. In Saccharomyces cerevisiae, Wss1, the DNA-protein crosslink repair protease, allows cells to tolerate hydroxyurea-induced replication stress but the underlying mechanism by which Wss1 promotes this function has remained unknown. Here we report that Wss1 provides cells tolerance to replication stress by directly degrading core histone subunits that non-specifically and non-covalently bind to single-stranded DNA. Unlike Wss1-dependent proteolysis of covalent DNA-protein crosslinks, proteolysis of histones does not require Cdc48 nor SUMO-binding activities. Wss1 thus acts as a multi-functional protease capable of targeting a broad range of covalent and non-covalent DNA-binding proteins to preserve genome stability during adverse conditions.

cell biology↗