Project description:Vibrio cholerae uses multiple strategies to resist predation by heterotrophic protozoa. For example, V. cholerae releases toxic compounds such as ammonium and pyomelanin, that can kill protists such as Tetrahymena pyriformis. V. cholerae also survives intracellularly and escapes as viable cells inside protozoan expelled food vacuoles (EFVs). We previously reported that V. cholerae encased in EFVs are hyperinfectious, establishing an important link between anti-protozoal strategies and bacterial virulence. Although the intracellular resistance and escape of V. cholerae in EFVs has been reported, the molecular mechanisms behind this remain poorly understood. Here, we used single cell transcriptomics of V. cholerae exposed to T. pyriformis and captured a total of 5,344 bacterial cells with heterogeneous gene expression. Cells with the same pattern of gene expression were grouped, resulting in eleven clusters of cells with a unique gene expression profile. Genes encoding outer membrane proteins, F1F0-Na+/H+ ATPase, metabolites and toxins showed differential expression among the clusters. Furthermore, the motility-associated killing factor (Mak) toxins (makA, makB and makC) were differentially expressed. Individual V. cholerae ΔmakA, ΔmakB, and ΔmakE strains were not capable of killing T. pyriformis and ΔmakA and ΔmakE showed reduced survival inside EFVs compared to the wild type. Our findings reveal new insights into the grazing resistance mechanisms of V. cholerae, identify factors associated with the survival of V. cholerae within EFVs and more broadly, highlight the connection between antiprotozoal and virulence factors displayed by pathogenic bacteria.
Project description:Three-dimensional genome architecture shapes how regulatory elements controls gene expression, yet how compact unicellular genomes are folded is poorly understood. Ciliates such as Tetrahymena thermophila carry a fragmented, gene-dense somatic macronucleus (MAC) and a silent germline micronucleus (MIC), imposing distinct constraints on chromosome folding. Here we combine nucleosome-resolution Micro-C, ATAC-seq and RNA-seq to map 3D chromatin organization across Tetrahymena life cycle stages and in the related ciliate Tetrahymena pyriformis. We find that MAC chromosomes lack strong mammalian-style compartments and TADs but behave as short interaction units whose telomere-capped ends form accessible, strongly interacting hubs. Within these units, chromatin loops concentrate at promoter-proximal open chromatin and are closely associated with transcription level. During conjugation, long-range internal loops and promoter–promoter contacts are transiently weakened and then rebuilt, while end–end interactions persist. Similar promoter- and end-anchored folding in T. pyriformis points to a conserved ciliate strategy for organizing gene-rich genomes in three dimensions.