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Abstract: The bacterial flagellum transforms its shape into several distinguishable helical shapes (polymorphs) under various environmental conditions. Polymorphs of each type of flagellum stay on a circle in the pitch-diameter (P versus πD) plot, indicating that they all belong to one family. Previously, we showed that the flagellar family of a marine bacterium Idiomarina loihiensis (Family II) differed from the conventional flagellar family of Salmonella typhimurium (Family I). The pitch and diameter of Family II flagella are half those of Family I flagella. We have suggested that Family I encompasses peritrichous flagella, while Family II forms a polar flagellum. In this study, we have surveyed the polymorphs of flagella from 18 other species and categorized their family types. Previous observations were confirmed; Family I form peritrichous flagella and Family II form polar flagella. Furthermore, we found that lateral flagella had helical parameters much smaller than those of the other two Families and thus belong to a new family (Family III). [Copyright &y& Elsevier]

The helical filaments of the bacterial flagella so far studied seem to be universal in the bacterial kingdom. Despite the variation in flagellin molecular masses, which range from 24kDa to 62kDa in different species, there are only two forms: either the so-called Normal (left-handed) or the Curly (right-handed). The Normal and Curly helical forms are asymmetric; the two characteristic helical parameters, which are the pitch and diameter, of Normal filaments are twice those of Curly filaments. Both the universality of these two helical forms and their asymmetry are biological puzzles. We found that the marine bacteria Idiomarina loihiensis have flagella with left-handed Curly-like filaments. Analysis of the polymorphic forms under different pH conditions showed that the Curly-like filaments are actually Normal filaments having a smaller pitch and diameter than those of Salmonella typhimurium. A minor modification of Calladine''s model for a filament lattice can explain the variant helical forms. Pseudomonas aeruginosa filaments also belong to the family of I.loihiensis filaments. Thus, there are at least two families of flagella filaments. [Copyright &y& Elsevier]

Abstract: To clarify the functions of flagella for pathogenesis of Erwinia carotovora subsp. carotovora EC1, non-polar in-frame deletion mutants of fliC and motA homologues were constructed. Both the ΔfliC and the ΔmotA mutants were confirmed to be non-motile on semisolid agar medium. The ΔmotA mutant was peritrichously flagellated like its parental strain under electron microscope, whereas the ΔfliC mutant was defective in flagellin production and aflagellated. We demonstrated that both the ΔfliC and the ΔmotA mutants reduced their ability to elicit the soft-rot symptoms on Chinese cabbage without any deleterious effect on the production of major extracellular enzymes. They retained their ability to induce hypersensitive reaction (HR) upon inoculation into tobacco leaves. These results suggest that motility but not flagellar component (s) is (are) involved in pathogenesis of E. carotovora subsp. carotovora. [Copyright &y& Elsevier]

The classic controversy of whether genotoxic chemicals induce cancers with or without a certain low-dose limit, i.e., the threshold, is revisited because of a number of current publications available addressing the plausibility of “practical” thresholds even for genotoxic carcinogens, the mechanism of which may be hypothesized to be due, in part, to a repair system composed of ordinarily available various defense mechanisms under the steady-state DNA damage. The question of whether an absolute nonthreshold or a relative nonthreshold, i.e., a “practical” threshold specifically in the low-dose level, is present may not be answered even with the use of a prohibitively large number of wild-type mice. Could the excessive incidence of tumorigenesis in p53-deficient mice contribute to our understanding of the threshold vs nonthreshold issue in genotoxic carcinogenesis? This is considered because an exaggeration of tumorigenesis in p53-deficient mice is hypothesized to reduce or eliminate the range of threshold due to the p53-deficiency-mediated reduction of DNA repair and apoptosis. The present study of chemical leukemogenesis in p53-deficient mice by transplantation assay was designed to answer this question. Briefly, 218 C3H/He mice were lethally irradiated and repopulated with bone marrow cells from wild-type, heterozygous p53-deficient, and homozygous p53-deficient C3H/He mice. This was followed by treatment with a single and graded dose of methyl nitrosourea at 6.6, 14.8, 33.3, 50.0, and 75.0 mg/kg body wt, with the vehicle-treated control groups treated with zero dose for each genotype. Whereas mice repopulated with p53-deficient bone marrow cells showed a marked reduction of the threshold for leukemogenicity, mice repopulated with wild-type bone marrow cells did not exhibit leukemia at a dose of 33.3 mg/kg body wt and showed a curve with a high probability for the linear regression model with a positive dose intercept, predicting a threshold by the likelihood ratio test. Thus, the failure of wild-type mice to show an increase in incidence of leukemogenesis at low doses of genotoxic carcinogens may be due not to a statistical rarity, but to various p53-related pharmacophysiological functions, possibly including DNA repair and apoptosis that may account for a threshold. [Copyright &y& Elsevier]

The bacterial flagellar hook is a short and uniquely curved tube that connects the basal body to the filament. Hook length is controlled at 55 nm on average by a soluble protein FliK in Salmonella enterica serovar Typhimurium. The N-terminal segment of FliK responsible for measuring the hook length is considered to be intrinsically disordered. Here, we show by high-speed atomic force microscopy that a FliK molecule in solution takes on a shape of two balls linked by a flexible string; the larger ball corresponds to the N-terminal region and the smaller one corresponds to the C-terminal region. The N-terminal domain is stable but the C-terminal domain fluctuates in shape. Based on these and other features of FliK, we propose that the folding of the N-terminal segment at the tip of the growing hook plays a major role in determining the minimal length of the hook. [ABSTRACT FROM AUTHOR]

Abstract: The flagellar cytoplasmic protein FliK controls hook elongation by two successive events: by determining hook length and by stopping the supply of hook protein. These two distinct roles are assigned to different parts of FliK: the N-terminal half (FliKN) determines length and the C-terminal half (FliKC) switches secretion from the hook protein to the filament protein. The interaction of FliKC with FlhB, the switchable secretion gate, triggers the switch. By NMR spectroscopy, we demonstrated that FliK is largely unstructured and determined the structure of a compact domain in FliKC. The compact domain, denoted the FliKC core domain, consists of two α-helices, a β-sheet with two parallel and two antiparallel strands, and several exposed loops. Based on the functional data obtained by a series of deletion mutants of the FliKC core domain, we constructed a model of the complex between the FliKC core domain and FlhBC. The model suggested that one of the FliKC loops has a high probability of interacting with the C-terminal domain of FlhB (FlhBC) as the FliK molecule enters the secretion gate. We suggest that the autocleaved NPTH sequence in FlhB contacts loop 2 of FliKC to trigger the switching event. This contact is sterically prevented when NPTH is not cleaved. Thus, the structure of FliK provides insight into the mechanism by which this bifunctional protein triggers a switch in the export of substrates. [Copyright &y& Elsevier]

Abstract: Bdellovibrio bacteriovorus cells have a single polar flagellum whose helical pitch and diameter characteristically change near the midpoint, resulting in a tapered wave. There are six flagellin genes in the genome: fliC1 to fliC6. Accordingly, the flagellar filament is composed of several similar flagellin species. We have used knockout mutants of each gene and analyzed the mutational effects on the filament length and on the composition and localization of each flagellin species in the filament by electron microscopy and one- and two-dimensional polyacrylamide gel electrophoresis. The location and amounts of flagellins in a filament were determined to be as follows: a small amount of FliC3 at the proximal end, followed by a large amount of FliC5, a large amount of FliC1, a small amount of FliC2 in this order, and a large amount of FliC6 at the distal end. FliC4 was present at a low level, but the location was not determined. Filament lengths of newly born progeny cells increased during prolonged incubation in nutrient-deficient buffer. The newly formed part of the elongated filament was composed of mainly FliC6. Reverse transcription PCR analysis of flagellar gene expression over 5 days in buffer showed that fliC gene expression tailed off over 5 days in the wild-type cells, but in the fliC5 mutant, expression of the fliC2, fliC4, and fliC6 genes was elevated on day 5, suggesting that they may be expressed to compensate for the absence of a major component, FliC5. [Copyright &y& Elsevier]

Abstract: A bacterial cell tethered through a flagellum on a glass slide rotates its cell body in either counter-clockwise or clockwise at around 10 Hz. To analyze the detailed manner of rotation, we have constructed and expressed yellow fluorescent protein (YFP)-FliN fusion protein in a fliN deletion mutant, resulting in the recovery of motility of the Fla- mutant cells. The tethered cells that incorporated the fusion protein in the flagellar motor rotate around one of the fluorescent spots. Tracing the center spot of a rotating cell, we have found that the rotating circles of the tethered cells were often distorted, and that the cell has seldom rotated smoothly but gyrated around the center point. The radii of the gyrating circles were 100–200 nm for the wild-type cells, and 50 nm for the cells carrying short hooks, suggesting that the flexibility of the hook is responsible for asymmetrical rotation. These observations indicate that tethered cells almost always interact with the glass surface in one cycle of rotation, where the length and flexibility of the hook have an important role. [Copyright &y& Elsevier]