11-12 November 2021
Digital (Zoom)
Europe/Berlin timezone

Super-resolution microscopy reveals the dynamic subcellular localization of pneumococcal proteins during an immunological challenge

12 Nov 2021, 13:30
30m
Digital (Zoom)

Digital (Zoom)

Speakers

Dr Claudia Vilhena (Infection Biology, Leibniz Institute for Natural Product Research and Infection Biology – Hans Knöll Institute (HKI), Jena, Germany) Dr Zoltan Cseresnyes (Applied Systems Biology, Leibniz Institute for Natural Product Research and Infection Biology – Hans Knöll Institute (HKI), Jena, Germany)

Description

Streptococcus pneumoniae is a gram-positive human pathogen that colonizes the upper respiratory tract and acts as the causal agent of otitis media, bronchitis and community-acquired pneumonia.
On the bacterial surface, S. pneumoniae harbours several proteins with diverse functions (transport channels, carbohydrate metabolic enzymes, iron receptors) that protrude to the extracellular space. A subset of these proteins is the so-called Choline-Binding Protein (CBP) group.
To allow a precise subcellular localization, single-cell super-resolution Structured Illumination Microscopy (SIM) coupled with batch processing image analysis was used under various conditions.
The super-resolution 3D image stacks were processed using HuygensPro and Imaris. The 2D images were processed using custom-developed workflows written in JIPipe (www.jipipe.org). The protein-specific signals were reconstructed by fitting uniformly small spheres over the bacterial volume. This analysis indicated that the protein distribution is not homogenous along the bacterial surface, but rather concentrated along an equatorial plane of the bacterium. Average distances to the five nearest neighbours showed a similar, equatorial distribution, and revealed a linear relationship between the fluorescence intensity of the clusters and their average distance to their 5-nearest neighbours. The redistribution of peptide clusters was similarly characterized during cell growth using various strains and growth conditions. End-point experiments of peptide distributions were examined using template matching algorithms to reveal the various stages of development during cell division.
The localization of the analysed CBP was not homogeneous along the cell wall which contrasts with the current knowledge presented in the literature. This finding is suggestive of a protective effect exerted by this CBP on designated positions on the cell wall that might correlate with immune attack. The linear relationship between protein cluster intensity and neighbour distance suggests a dynamic and fine-tuned allocation of proteins to the cell wall to avoid areas with simultaneously high protein density and intensity.
Our work presents a novel view and an optimized analytical tool to provide insights into the dynamic spatial positioning of pneumococcal proteins, thus opening new ways to reassess how these pathogens exploit their protein machineries to evade the immune system.

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