Graduate Students Mini Symposium II-2024

01:15 PM Katherine Pintor (RG Diepold)
The role of type III secretion chaperones in effector export

The type III secretion system (T3SS) is used by many bacteria, including important pathogens, to inject effector proteins into eukaryotic cells. Critical steps of the secretion process such as the selection, recruitment, and export path of the effector proteins are still poorly understood. Most effectors are bound to specific T3SS chaperones that are often co-expressed in the cytosol, while some cargo are secreted in a chaperone-independent manner. In order to elucidate how chaperones enable or enhance effector export and translocation to the target host cell, we measured and visualized the lifecycle of the chaperones SycH and SycE in real time and at a single molecule level in Yersinia enterocolitica. Using single particle tracking photoactivated localization microscopy (sptPALM) and fluorescence correlation spectroscopy (FCS), we could show that SycH and SycE diffuse faster in the absence of effectors, which verifies the interaction of effectors and chaperones. Upon initiation of secretion, SycH became more mobile, which could imply a transition from cargo recognition to cargo handover mode during export. In contrast, SycE exhibited slower mobility under secreting conditions, based on our FCS data. This suggests that although both chaperone proteins share a similar biological function on the secretion of substrates, they might adapt different regulatory mechanisms during export. Moreover, we also explored how chaperones behave in the absence of the mobile T3SS sorting platform (ΔSctQ) and the T3SS membrane rings (ΔSctD). While effector-bound chaperones diffuse slightly faster in the absence of SctQ, mobility of the chaperones in the absence of effectors was not significantly altered, implying that chaperones only bind to the sorting platform transiently and in the presence of their respective effector proteins. Moreover, deletion of SctD has no impact on the diffusion of chaperones, which means the interaction of chaperones and effectors is independent of the injectisome assembly. These findings suggest that chaperones recognize and bind to effectors in the bacterial cytosol, and hand over their cargo to mobile T3SS sorting platform components for delivery into the T3SS upon activation of secretion.

01:45 PM Joana Kästle (RG Brune)
Reductive genome evolution in Ca. Ancillulaceae (Bifidobacteriales), a novel lineage of endosymbionts in termite gut flagellates

Cellulolytic gut flagellates of lower termites are essential for the symbiotic digestion of lignocellulose. They are associated with diverse but host-specific consortia of ecto- and endosymbiotic bacteria, whose functional roles in the multilayered symbiotic system are largely unknown. Here, we investigated a termite-specific clade of Bifidobacteriales (Actinobacteria) using 16S rRNA gene libraries of capillary-picked flagellates, fluorescence in situ hybridization (FISH), and comparative genome analysis of metagenome-assembled genomes (MAGs). We found that Ca. Ancillulaceae comprises two phylogenetically and functionally distinct groups that experienced substantial genome reduction. Ca. Ancillula (average genome size 1.48 Mbp) are endosymbionts of Trichonympha spp. and retained the capacity to synthesize amino acids, vitamins, and co-factors. Ca. Opitulatrix (average genome size 1.2 Mbp), a hitherto unknown lineage of putative flagellate symbionts, has lost most biosynthetic capacities and depends on the uptake of numerous metabolites from its flagellate hosts. Both groups retained an amino acid export system and acquired novel gene functions in response to their intracellular lifestyle.

02:15 PM Juliana Effert (AG Bode)
Biosynthesis of bacterial pyrrolizidine alkaloids and production of new derivatives

Pyrrolizidine alkaloids (PA) represent a class of widespread natural products from plants and bacteria, where they potentially have a biological role in host-bacteria interactions and virulence. PA compounds are recognized as privileged structural motifs in small molecule drug discovery and show a broad pharmaceutical activity. Therefore, it is of high interest to identify or generate novel PAs with beneficial bioactivities. Here, we identify a novel PA dimer, pyrrolizwilline, and unravel its biosynthesis pathway. Furthermore, we exploit the biosynthetic pathway of pyrrolizixenamide and successfully apply the NRPS engineering concepts developed in our lab, to broaden the structural diversity of PAs.