Abstract
Salmonella enterica (e.g., serovars Typhi and Typhimurium) relies on translocation of effectors via type III secretion systems (T3SS). Specialization of typhoidal serovars is thought to be mediated via pseudogenesis. Here, we show that the Salmonella Typhi STY1076/t1865 protein, named StoD, a homologue of the enteropathogenic Escherichia coli/enterohemorrhagic E. coli/Citrobacter rodentium NleG, is a T3SS effector. The StoD C terminus (StoD-C) is a U-box E3 ubiquitin ligase, capable of autoubiquitination in the presence of multiple E2s. The crystal structure of the StoD N terminus (StoD-N) at 2.5 Å resolution revealed a ubiquitin-like fold. In HeLa cells expressing StoD, ubiquitin is redistributed into puncta that colocalize with StoD. Binding assays showed that StoD-N and StoD-C bind the same exposed surface of the β-sheet of ubiquitin, suggesting that StoD could simultaneously interact with two ubiquitin molecules. Consistently, StoD interacted with both K63- (KD = 5.6 ± 1 μM) and K48-linked diubiquitin (KD = 15 ± 4 μM). Accordingly, we report the first S. Typhi-specific T3SS effector. We suggest that StoD recognizes and ubiquitinates pre-ubiquitinated targets, thus subverting intracellular signaling by functioning as an E4 enzyme.
Highlights
ObjectivesThe aim of this study was to determine whether STY1076 is a T3SS effector and to elucidate its structure and function
Salmonella enterica subspecies enterica is divided into typhoidal
Since first identified as T3SS effectors in the mouse pathogen Citrobacter rodentium [14], NleG proteins have been found in enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli (EHEC) [15], as well as Salmonella bongori, where it is named SboD (S. bongori contains two truncated NleG family members named SboE and SboF) [16]
Summary
The aim of this study was to determine whether STY1076 is a T3SS effector and to elucidate its structure and function. Considering that S. bongori only expresses SPI-1, we aimed to determine if StoD is an SPI-1 S. To gain further insights into the cellular function of StoD, we aimed to localize it during infection of cultured cells
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