Biology subjects

Winkler, M.

Publications and source records attributed to Winkler, M..

24 records · Page 2Linked to original sources

bioRxiv2020

Sustained nitrogen loss in a symbiotic association of Comammox Nitrospira and Anammox bacteria

The discovery of complete aerobic and anaerobic ammonia-oxidizing bacteria (Comammox and Anammox) significantly altered our understanding of the global nitrogen cycle. A high affinity for ammonia (Km(app),NH3 {approx} 63nM) and oxygen place the first described isolate, Comammox Nitrospira inopinata in the same trophic category as organisms such as some ammonia-oxidizing archaea. However, N. inopinata has a relatively low affinity for nitrite (Km,NO2 {approx} 449.2M) suggesting it would be less competitive for nitrite than other nitrite-consuming aerobes and anaerobes. We examined the ecological relevance of the disparate substrate affinities by coupling it with Anammox (Nitrospira inopinata and Brocadia anammoxidans, respectively). Synthetic communities were established in hydrogel granules in which Comammox grew in the aerobic outer layer to provide Anammox with nitrite in the inner anoxic core to form dinitrogen gas. This spatial organization was confirmed with FISH imaging, supporting a mutualistic or commensal relationship. Successful co-habitation of Comammox N. inopinata and Anammox in synthetic granules broadens our limited understanding of the interplay between these two species and offers potential biotechnological applications to study any type of bacterial pairings in a systematic and reproducible manner.

ecology↗

bioRxiv2020

Camostat mesylate inhibits SARS-CoV-2 activation by TMPRSS2-related proteases and its metabolite GBPA exerts antiviral activity

Antiviral therapy is urgently needed to combat the coronavirus disease 2019 (COVID-19) pandemic, which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The protease inhibitor camostat mesylate inhibits SARS-CoV-2 infection of lung cells by blocking the virus-activating host cell protease TMPRSS2. Camostat mesylate has been approved for treatment of pancreatitis in Japan and is currently being repurposed for COVID-19 treatment. However, potential mechanisms of viral resistance as well as camostat mesylate metabolization and antiviral activity of metabolites are unclear. Here, we show that SARS-CoV-2 can employ TMPRSS2-related host cell proteases for activation and that several of them are expressed in viral target cells. However, entry mediated by these proteases was blocked by camostat mesylate. The camostat metabolite GBPA inhibited the activity of recombinant TMPRSS2 with reduced efficiency as compared to camostat mesylate and was rapidly generated in the presence of serum. Importantly, the infection experiments in which camostat mesylate was identified as a SARS-CoV-2 inhibitor involved preincubation of target cells with camostat mesylate in the presence of serum for 2 h and thus allowed conversion of camostat mesylate into GBPA. Indeed, when the antiviral activities of GBPA and camostat mesylate were compared in this setting, no major differences were identified. Our results indicate that use of TMPRSS2-related proteases for entry into target cells will not render SARS-CoV-2 camostat mesylate resistant. Moreover, the present and previous findings suggest that the peak concentrations of GBPA established after the clinically approved camostat mesylate dose (600 mg/day) will result in antiviral activity.

molecular biology↗

bioRxiv2020

Neuropilin-1 facilitates SARS-CoV-2 cell entry and provides a possible pathway into the central nervous system

The causative agent of the current pandemic and coronavirus disease 2019 (COVID-19) is the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)1. Understanding how SARS-CoV-2 enters and spreads within human organs is crucial for developing strategies to prevent viral dissemination. For many viruses, tissue tropism is determined by the availability of virus receptors on the surface of host cells2. Both SARS-CoV and SARS-CoV-2 use angiotensin-converting enzyme 2 (ACE2) as a host receptor, yet, their tropisms differ3-5. Here, we found that the cellular receptor neuropilin-1 (NRP1), known to bind furin-cleaved substrates, significantly potentiates SARS-CoV-2 infectivity, which was inhibited by a monoclonal blocking antibody against the extracellular b1b2 domain of NRP1. NRP1 is abundantly expressed in the respiratory and olfactory epithelium, with highest expression in endothelial cells and in the epithelial cells facing the nasal cavity. Neuropathological analysis of human COVID-19 autopsies revealed SARS-CoV-2 infected NRP1-positive cells in the olfactory epithelium and bulb. In the olfactory bulb infection was detected particularly within NRP1-positive endothelial cells of small capillaries and medium-sized vessels. Studies in mice demonstrated, after intranasal application, NRP1-mediated transport of virus-sized particles into the central nervous system. Thus, NRP1 could explain the enhanced tropism and spreading of SARS-CoV-2.

microbiology↗

bioRxiv2018

The cell cycle regulator GpsB functions as cytosolic adaptor for multiple cell wall enzymes

Bacterial growth and cell division requires precise spatiotemporal regulation of the synthesis and remodelling of the peptidoglycan layer that surrounds the cytoplasmic membrane. GpsB is a cytosolic protein that affects cell wall synthesis by binding to the cytoplasmic mini-domains of peptidoglycan synthases to ensure their correct subcellular localisation. Here we have discovered critical structural features for the interaction of GpsB with peptidoglycan synthases from three different bacteria and demonstrated their importance for cell wall growth and viability. We have used these structural motifs to predict and confirm novel partners of GpsB in Bacillus subtilis, illuminating the role of this key regulator of peptidoglycan synthesis. GpsB thus functions as an adaptor, to mediate the interaction between membrane proteins, scaffolding proteins, signalling proteins and enzymes to generate larger protein complexes at specific sites in a bacterial cell cycle-dependent manner. Given the importance of GpsB in pathogenic bacteria, this study has not only revealed mechanistic details of how cell wall synthesis is co-ordinated with the bacterial cell cycle but could also represent a starting point for the design of much needed new antibiotics.

microbiology↗

bioRxiv2018

Resource limitation modulates the fate of dissimilated nitrogen in a dual-pathway Actinobacterium

Respiratory ammonification and denitrification are two evolutionarily unrelated dissimilatory nitrogen (N) processes central to the global N cycle, the activity of which is thought to be controlled by carbon (C) to nitrate (NO3-) ratio. Here we find that Intrasporangium calvum C5, a novel menaquinone-based dual-pathway denitrifier/respiratory ammonifier, disproportionately utilizes ammonification rather than denitrification when grown under carbon or nitrate limitation, not C:NO3- ratio. Higher growth rates are promoted by ammonification and metabolite and transcriptional profiles during growth show that the bacterium produces its own formate from a fermentable carbon source (lactate) to further generate a proton motive force for the ammonification pathway. Transcript abundances encoding for nitrite reducing enzymes, NrfAH and NirK, also significantly increase in response to nitrite production. Mechanistically, our results suggest that pathway selection is driven by intracellular redox potential (redox poise), which may be lowered during resource limitation, thereby decreasing catalytic activity of upstream electron transport steps needed for denitrification enzymes. Our work advances our understanding of the biogeochemical flexibility of N-cycling organisms and pathway evolution.

microbiology↗

bioRxiv2018

FASTGenomics: An analytical ecosystem for single-cell RNA sequencing data

Recent technological advances enable genomics of individual cells, the building blocks of all living organisms. Single cell data characteristics differ from those of bulk data, which led to a plethora of new analytical strategies. However, solutions are only useful for experts and currently, there are no widely accepted gold standards for single cell data analysis. To meet the requirements of analytical flexibility, ease of use and data security, we developed FASTGenomics (https://fastgenomics.org) as a powerful, efficient, versatile, robust, safe and intuitive analytical ecosystem for single-cell transcriptomics.

bioinformatics↗