FliK regulates flagellar hook length as an internal ruler

Abstract

SummaryThe mechanism of length control of the flagellar hook is under debate between two theories. One claims that the FliK directly measures the hook length as a molecular ruler, while the other claims that the cytoplasmic substructure measures the amount of hook subunits to determine the hook length. Both agree that the FliK C-terminal domain catalyses the substrate-specificity switch to terminate hook elongation. In this study, we systematically created fliK mutants with deletions and insertions at various sites within the FliK N-terminal domain and analysed their effects on the final hook length. Insertions of peptide fragments from the Yersinia YscP into FliK gave rise to hooks with defined lengths, which was proportional to the molecular size of the FliK-YscP chimeras. Among fliK deletion mutants, only those with small truncations in three specific sites of FliK produced hooks of a defined, shortened length. For the majority of deletion mutants, FliK was secreted, but hook length was not controlled. On the other hand, for some deletion mutants FliK was not secreted, but the hook length was controlled, indicating that FliK secretion is not necessary for hooklength control. We conclude that FliK regulates hook length as an internal molecular ruler.

Protein Export According to Schedule: Architecture, Assembly, and Regulation of Type III Secretion Systems from Plant- and Animal-Pathogenic Bacteria

“…In agreement with this model, substrate specificity switching by FliK ⌬1-99 was increased in the absence of Fluke (385). Notably, deletions in the central part (amino acids 208 to 278) of FliK, outside the T3S4 domain (amino acids 265 to 405), did not abolish filament formation and hook length control (502). Since these FliK derivatives were initially not detected in the culture supernatant, FliK was proposed to act as an internal ruler (502).…”

“…Since insertions and deletions outside the C-terminal T3S4 domain of FliK lead to increased and reduced hook lengths, respectively, FliK most likely acts as a molecular ruler, as proposed for YscP from Yersinia spp. (502). Interestingly, experimental evidence suggests that FliK is involved not only in hook length control but also in length control of the inner rod of the flagellar T3S system (538).…”

“…Notably, deletions in the central part (amino acids 208 to 278) of FliK, outside the T3S4 domain (amino acids 265 to 405), did not abolish filament formation and hook length control (502). Since these FliK derivatives were initially not detected in the culture supernatant, FliK was proposed to act as an internal ruler (502). However, secretion of FliK derivatives with deletions in the central protein region was shown in a later study by the use of a more sensitive FliK-specific antibody (159).…”

Type III secretion systems: the bacterial flagellum and the injectisome

“…However, once the core T3SS is assembled, it starts to independently export early substrates, the flagellar hook FlgE and the hook-capping protein FlgD in the case of the flagellum and the needle subunit in the case of the injectisome. In both cases, the correct length of the filament or needle is measured, in a so-far incompletely understood way, by a ruler protein (FliK/SctP) [136][137][138]. In interaction with an export apparatus component, the type III secretion substrate specificity switch (T3S4) protein FlhB/ SctU, it induces a change in substrate specificity, allowing secretion and assembly of the flagellin subunits and the needle tip proteins, respectively (figure 3).…”

Role of EscP (Orf16) in Injectisome Biogenesis and Regulation of Type III Protein Secretion in Enteropathogenic Escherichia coli

“…Many different mechanisms have been proposed to explain needle/hook length control (12,27,29,45,59,60,76,93,106,109). Interestingly, extragenic suppressor mutations of fliK and yscP are localized within flhB and yscU (25,41,53,110).…”