In situ structure of the Caulobacter crescentus flagellar motor and visualization of binding of a CheY-homolog

Bacterial flagellar motility is controlled by the binding of CheY proteins to the cytoplasmic switch complex of the flagellar motor, resulting in changes in swimming speed or direction. Despite its importance for motor function, structural information about the interaction between effector proteins and the motor are scarce. To address this gap in knowledge, we used electron cryotomography and subtomogram averaging to visualize such interactions inside Caulobacter crescentus cells. In C. crescentus, several CheY homologs regulate motor function for different aspects of the bacterial lifestyle. We used subtomogram averaging to image binding of the CheY family protein CleD to the cytoplasmic Cring switch complex, the control center of the flagellar motor. This unambiguously confirmed the orientation of the motor switch protein FliM and the binding of a member of the CheY protein family to the outside rim of the C ring. We also uncovered previously unknown structural elaborations of the alphaproteobacterial flagellar motor, including two novel periplasmic ring structures, and the stator ring harboring eleven stator units, adding to our growing catalog of bacterial flagellar diversity

Bacterial Flagellins: Does Size Matter?

The bacterial flagellum is the principal organelle of motility in bacteria. Here, we address the question of size when applied to the chief flagellar protein flagellin and the flagellar filament. Surprisingly, nature furnishes multiple examples of ‘giant flagellins’ greater than a thousand amino acids in length, with large surface-exposed hypervariable domains. We review the contexts in which these giant flagellins occur, speculate as to their functions, and highlight the potential for biotechnology to build on what nature provides

Benefit of second-line systemic chemotherapy for advanced biliary tract cancer: A propensity score analysis

Abstract Whether 2nd-line-chemotherapy (2LCTX) + best-supportive-care (BSC) benefits patients with advanced biliary tract cancer (aBTC) more than BSC alone is unclear. We therefore conducted a propensity-score-based comparative effectiveness analysis of overall survival (OS) outcomes in 80 patients with metastatic, recurrent, or inoperable aBTC, of whom 38 (48%) were treated with BSC + 2LCTX and 42 (52%) with BSC alone. After a median follow-up of 14.8 months and 49 deaths, the crude 6-, 12-, and 18-month Kaplan-Meier OS estimates were 77%, 53% and 23% in the BSC + 2LCTX group, and 29%, 21%, and 14% in patients in the BSC group (p = 0.0003; Hazard ratio (HR) = 0.36, 95%CI:0.20–0.64, p = 0.001). An inverse-probability-of-treatment-weighted (IPTW) analysis was conducted to rigorously account for the higher prevalence of favorable prognostic variables in the 2LCTX + BSC group. After IPTW-weighting, the favorable association between 2LCTX and OS prevailed (adjusted HR = 0.40, 95%CI: 0.17–0.95, p = 0.037). IPTW-weighted 6-, 12-, and 18-month OS estimates were 77%, 58% and 33% in the BSC + 2LCTX group, and 39%, 28% and 22% in the BSC group (p = 0.037). Moreover, the benefit of 2LCTX was consistent across several clinically-relevant subgroups. Within the limitations of an observational study, these findings support the concept that 2LCTX + BSC is associated with an OS benefit over BSC alone in aBTC

An ATP-dependent partner switch links flagellar C-ring assembly with gene expression

Bacterial flagella differ in their number and spatial arrangement. In many species, the MinD-type ATPase FlhG (also YlxH/FleN) is central to the numerical control of bacterial flagella, and its deletion in polarly flagellated bacteria typically leads to hyperflagellation. The molecular mechanism underlying this numerical control, however, remains enigmatic. Using the model species Shewanella putrefaciens, we show that FlhG links assembly of the flagellar C ring with the action of the master transcriptional regulator FlrA (named FleQ in other species). While FlrA and the flagellar C-ring protein FliM have an overlapping binding site on FlhG, their binding depends on the ATP-dependent dimerization state of FlhG. FliM interacts with FlhG independent of nucleotide binding, while FlrA exclusively interacts with the ATP- dependent FlhG dimer and stimulates FlhG ATPase activity. Our in vivo analysis of FlhG partner switching between FliM and FlrA reveals its mechanism in the numerical restriction of flagella, in which the transcriptional activity of FlrA is down-regulated through a negative feedback loop. Our study demonstrates another level of regulatory complexity underlying the spationumerical regulation of flagellar biogenesis and implies that flagellar assembly transcriptionally regulates the production of more initial building blocks