Biology subjects

Sedzicki, J.

Publications and source records attributed to Sedzicki, J..

5 recordsLinked to original sources

bioRxiv2024

The putative Type 4 secretion system effector BspD is involved in maintaining envelope integrity of the pathogen Brucella

Brucellosis is a debilitating disease caused by the Gram-negative, facultative intracellular zoonotic pathogen Brucella. En route to its intracellular replicative niche, Brucella encounters various stressful environments that may compromise envelop integrity. Here we show, that the proposed Type 4 secretion system (T4SS) effector BspD is a conserved protein of the Rhizobiales, which does not show signs of co-evolution with the presence of a T4SS or a certain lifestyle. We further present data indicating that BspD is critical for Brucella abortus envelope integrity in the stationary phase and in the presence of EDTA, a compound known to destabilize the outer membrane. Deletion of bspD resulted in abnormal bacterial morphologies, indicating its involvement in maintaining envelope integrity. Additionally, the absence of BspD led to the formation of fewer and smaller intracellular microcolonies in a macrophage infection model. From our observations, we propose that BspD of B. abortus is critical for preserving the integrity of the bacterial envelope, particularly under stressful conditions, which may enhance Brucellas ability to survive within host cells. ImportanceBrucellosis, caused by the intracellular pathogen Brucella, poses a significant health threat. Understanding how Brucella adapts to stressful environments is crucial. This study unveils BspD, a conserved protein within the Rhizobiales order, as a key player in maintaining Brucellas envelope integrity. Remarkably, BspDs function appears independent of the presence of a Type 4 Secretion System (T4SS) or specific lifestyle. Deletion of bspD resulted in compromised envelope integrity, abnormal bacterial morphologies, and reduced intracellular microcolony formation. These findings underscore BspDs critical role, particularly in stressful conditions like the stationary phase and EDTA exposure, and highlight its significance in Brucellas survival within host cells. This elucidation deepens our understanding of Brucella pathogenesis and may inform future therapeutic strategies against brucellosis.

microbiology↗

bioRxiv2023

Structure-function analysis of the cyclic beta-1,2-glucan synthase

The synthesis of complex sugars is a key aspect of microbial biology. Cyclic {beta}-1,2-glucan (C{beta}G) is a circular polysaccharide critical for host interactions of many bacteria, including major pathogens of humans (Brucella) and plants (Agrobacterium). C{beta}G is produced by the cyclic glucan synthase (Cgs), a massive multi-domain membrane protein. So far, its structure as well as the mechanisms underlining the synthesis have not been clarified. Here we use cryo-electron microscopy (cryo-EM) and functional approaches to study Cgs from A. tumefaciens. We were able to determine the structure of this complex protein machinery and clarify key aspects of C{beta}G synthesis. Our research opens new possibilities for combating pathogens that rely on polysaccharide virulence factors and can lead to new synthetic biology approaches for producing complex cyclic sugars.

molecular biology↗

bioRxiv2022

BNIP3L-mediated mitophagy triggered by Brucella in host cells is required for bacterial egress

The facultative intracellular pathogen Brucella abortus interacts with several organelles of the host cell to reach its replicative niche inside the endoplasmic reticulum. However, little is known about the interplay between the bacteria and the host cell mitochondria. Here, we showed that B. abortus triggers a strong mitochondrial network fragmentation accompanied by mitophagy and the formation of mitochondrial Brucella-containing vacuoles in the late steps of cellular infection. The expression of the mitophagy receptor BNIP3L induced by B. abortus is essential for these events and relies on the iron-dependent stabilization of the hypoxia-inducible factor 1 alpha. Functionally, BNIP3L-mediated mitophagy appears to be advantageous for bacterial exit of the host cell as BNIP3L depletion drastically reduced the number of reinfection events. Altogether, these findings highlight the intricate link between Brucella trafficking and the mitochondria during host cell infection.

microbiology↗

bioRxiv2021

The Bartonella autotransporter CFA is a protective antigen and hypervariable target of neutralizing antibodies blocking erythrocyte infection

Antibodies are key to the clearance of Bartonella bacteremia, but the mechanisms and targets of protective antibodies are unknown and bacterial evasion strategies remain elusive. We studied experimental Bartonella taylorii infection of mice, its natural host, and investigated protective immune responses. Clearance of bacteremia depended on specific antibodies that interfere with bacterial attachment to erythrocytes. Accordingly, antibodies were effective in the absence of complement and Fc-receptors. Moreover, they formed independently of B-cell hypermutation and isotype class switch. The cloning of neutralizing monoclonal antibodies (mAbs) led to the identification of the bacterial autotransporter CFA as a protective antibody target, and vaccination against CFA protected against Bartonella bacteremia. MAb binding mapped to a region of CFA that is hypervariable in both human- and mouse-pathogenic Bartonella strains, suggesting mutational antibody evasion. These insights further our understanding of Bartonella immunity and immune evasion and elucidate mechanisms driving high Bartonella prevalence in the wild.

microbiology↗

bioRxiv2018

A role for VPS35 in Brucella intracellular replication revealed by genome-wide siRNA screening

Brucella, the causing agent of brucellosis, is a major zoonotic pathogen with worldwide distribution. Brucella resides and replicates inside infected host cells in membrane-bound compartments called BCVs (Brucella-containing vacuoles). Following uptake, Brucella resides in eBCVs (endosomal BCVs) that gradually mature from early to late endosomal features. Through a poorly understood process that is key to the intracellular lifestyle of Brucella, the eBCV escapes fusion with lysosomes by transitioning to the rBCV (replicative BCV), a replicative niche directly connected to the endoplasmic reticulum (ER). Despite the notion that this complex intracellular lifestyle must depend on a multitude of host factors, a holistic view on which of these components control Brucella cell entry, trafficking and replication is still missing. Here we used a systematic cell-based siRNA knockdown screen in HeLa cells infected with Brucella abortus and identified 425 components of the human infectome for Brucella infection. These include multiple components of pathways involved in central processes such as cell cycle, actin cytoskeleton dynamics or vesicular trafficking. Using assays for pathogen entry, knockdown complementation and co-localization at single-cell resolution, we identified the requirement of the VPS retromer for Brucella to escape the lysosomal degradative pathway and to establish its intracellular replicative niche. We thus validated a component of the VPS retromer as novel host factor critical for Brucella intracellular trafficking. Further, our genome-wide data shed light on the interplay between central host processes and the biogenesis of the Brucella replicative niche.\n\nImportanceWith >300,000 new cases of human brucellosis annually, Brucella is regarded as one of the most important zoonotic bacterial pathogen worldwide. The causing agent of brucellosis resides inside host cells within vacuoles termed Brucella containing vacuoles (BCVs). Although few host components required to escape the degradative lysosomal pathway and to establish the ER-derived replicative BCV (rBCV) have already been identified, the global understanding of this highly coordinated process is still partial and many factors remain unknown. To gain a deeper insight into these fundamental questions we performed a genome-wide RNA interference (RNAi) screen aiming at discovering novel host factors involved in the Brucella intracellular cycle. We identified 425 host proteins that contribute to Brucella cellular entry, intracellular trafficking, and replication. Together, this study sheds light on previously unknown host pathways required for the Brucella infection cycle and highlights the VPS retromer components as critical factors for the establishment of the Brucella intracellular replicative niche.

microbiology↗