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PubMed2002

Transient concentration of a gamma-tubulin-related protein with a pericentrin-related protein in the formation of basal bodies and flagella during the differentiation of Naegleria gruberi.

The distribution of two proteins in Naegleria gruberi, N-gammaTRP (Naegleria gamma-tubulin-related protein) and N-PRP (Naegleria pericentrin-related protein), was examined during the de novo formation of basal bodies and flagella that occurs during the differentiation of N. gruberi. After the initiation of differentiation, N-gammaTRP and N-PRP began to concentrate at the same site within cells. The percentage of cells with a concentrated region of N-gammaTRP and N-PRP was maximal (68%) at 40 min when the synthesis of tubulin had just started but no assembled microtubules were visible. When concentrated tubulin became visible (60 min), the region of concentrated N-gammaTRP and N-PRP was co-localized with the tubulin spot and then flagella began to elongate from the region of concentrated tubulin. When cells had elongated flagella, the concentrated N-gammaTRP and N-PRP were translocated to the opposite end of the flagellated cells and disappeared. The transient concentration of N-gammaTRP coincided with the transient formation of an F-actin spot at which N-gammaTRP and alpha-tubulin mRNA were co-localized. The concentration of N-gammaTRP and formation of the F-actin spot occurred without the formation of microtubules but were inhibited by cytochalasin D. These observations suggest that the regional concentration of N-gammaTRP and N-PRP is mediated by actin filaments and might provide a site of microtubule nucleation for the assembly of newly synthesized tubulins into basal bodies and flagella.

Actins↗

PubMed1987

Structure and mass analysis of 12S and 19S dynein obtained from bull sperm flagella.

The Brookhaven scanning transmission electron microscope (STEM) was used to elucidate the structures and masses of 12S and 19S dynein extracted from bull sperm flagella. The 12S particle was a single globular particle with an average mass of 311 +/- 10 kdaltons. The 19S dynein particles consisted of two globular heads joined to a common base. The average mass of the 19S particle was 1.6 +/- 0.04 X 10(6) daltons. Thus, with the exception of the larger mass, the bull sperm 19S dynein molecule resembles the two-headed 21S dynein obtained from sea urchin sperm flagella and the 18S dynein obtained from Chlamydomonas with the possibility of a third head giving rise to the 12S particle. The structure, mass and polypeptide composition of bull sperm flagella dynein is compared with outer arm dyneins previously obtained from Chlamydomonas, Tetrahymena, and sea urchin sperm flagella.

Adenosine Triphosphatases↗

PubMed1986

Reversible inhibition of flagella formation after specific inhibition of spermidine synthesis by dicyclohexylamine in Pseudomonas aeruginosa.

Dicyclohexylamine, which is an inhibitor of bacterial and mammalian spermidine synthase, greatly inhibited the synthesis of spermidine in Pseudomonas aeruginosa. The depletion of spermidine caused by dicyclohexylamine was accompanied by an inhibition of growth of bacteria. This inhibition was reversed by addition of 50 microM spermidine (but not putrescine or spermine) to growth medium. When its growth was inhibited Ps. aeruginosa also lost its motility. Electron microscopy showed a loss of flagella in spermidine-deficient bacteria: after 24 h 70% 85% of bacteria grown in the presence of dicyclohexylamine did not have flagella, whereas bacteria grown in the presence of dicyclohexylamine and spermidine had flagella. The loss of flagella was reversible, since after the inhibition of spermidine synthesis for 24 h, addition of 50 microM spermidine (but not putrescine or spermine) to the growth medium was able to restore the bacterial motility almost completely after a further 12 h growth period.

Cell Movement↗

PubMed1975

Effect of temperature, salts, pH, and other factors on the development of peritrichous flagella in Vibrio alginolyticus.

The development of peritrichous flagella and, consequently, swarming of Vibrio alginolyticus depend on a complex relationship between temperature, salt concentrations and pH. At temperatures above 28 degrees C V. alginolyticus did not develop peritrichous flagella unless certain minimal concentrations of NaCl are present: the higher the temperature, the higher the NaCl concentrations required for peritrichous flagella synthesis. This requirement for NaCl at high temperatures is much more pronounced at pH 9 than at pH 6. High temperatures and low concentrations of NaCl also inhibited swarming of cells already armed with peritrichous flagella. Other cations, such as Li+, K+ and Mg2+. replaced NaCl only at temperatures below 28 degrees C.

Cell Movement↗

PubMed1995

Unfolding of tertiary structure of Halobacterium halobium flagellins does not result in flagella destruction.

The structure of Halobacterium halobium R1M1 flagella is investigated by the methods of scanning microcalorimetry, circular dichroism, and electron microscopy. It is shown that melting curves of flagella in solutions with a different concentration of NaCl display only one peak of heat capacity that corresponds to one cooperatively melting domain. It is found that flagella do not dissociate after melting. The possible structural organization of archaebacterial flagella is discussed.

Amino Acid Sequence↗

PubMed1998

Isolation and characterization of flagella from Trichomonas vaginalis.

As a first step in the biochemical and biomedical analyses of flagella from Trichomonas vaginalis the flagella were isolated, purified, and analyzed. The flagella were detached by mechanical shearing and a crude flagellar preparation was obtained by low-speed differential centrifugation. The crude flagellar preparation was subjected to further purification by discontinuous sucrose density-gradient centrifugation. Electron micrographs (EM) of the purified flagella showed the typical 9 + 2 axonemal arrangement. The structural integrity and the flagellar membrane were not destroyed by the deflagellation method or the purification scheme employed. The flagellar preparations were resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The purified preparation contained many flagellar enriched proteins ranging from 20 to 120 kDa. Three major proteins of 65 kDa and a doublet of about 50-58 kDa were observed. The protein patterns and EM appearance of the fractions were highly reproducible.

Animals↗

PubMed1978

Purification and biochemical properties of complex flagella isolated from Rhizobium lupini H13-3.

1. The complex flagella of Rhizobium lupini H13-3 differ from plain bacterial flagella in the fine structure of their filaments dominated by conspicuous helical bands, in their fragility and their resistance against heat decomposition. To elucidate the basis of these differences, the composition of complex filaments and their subunits was analysed. 2. Isolated complex flagella containing the filament and hook protions were purified by differential centrifugation. Hooks were separated by ultracentrifugation after acid degradation of filaments at pH 2. The complex filaments consist of 43 000 dalton monomers (cx-flagellin), the hooks are composed of 41 000 dalton subunits. 3. Amino acid analysis of cx-flagellin indicated the presence of approx. 417 amino acid residues. These comprise 47% hydrophobic residues and 21% Asp and Glu (or amides), but no Cys, His, Pro and Trp. No carbohydrate, phosphate or lipid moieties have been detected. Fingerprint analysis after tryptic digestion yields approx. 36 peptides, about half of them clustered in the neutral region. A comparison with the composition of varous known flagellins from plain flagella indicates a 7% higher content of hydrophobic amino acid residues in complex filaments; this is largely compensated for by the higher content of Glu and Asp (presumably as Gln and Asn) in plain filaments. 4. Immunodiffusion and immunoelectrophoresis of cx-flagellin yield single precipitin bands indicating homogeneity. In contrast, isoelectric focusing lead to three close-running bands around pH4.7. When isolated, the two major bands again produced an "isoelectric spectrum" suggesting that it reflects an allomorphism of cx-flagellin. 5. Self-assembly experiments with cx-flagellin lead to coiled fibres including helical regions, but not to intact filaments. The products resemble heat-denatured complex filaments and may represent intermediates between monomers and complete polymers.

Amino Acids↗

PubMed1988

Swimming behaviour of the unicellular biflagellate Oxyrrhis marina: in vivo and in vitro movement of the two flagella.

The movement of the 2 flagella of Oxyrrhis marina was examined with respect to their individual waveforms and the swimming behavior of the organism. The longitudinal flagella propagated helicoidal waves whose amplitude decreased toward the tip of th flagellum. Their beat frequencies were 50-60 Hz. The transverse flagella beat helicoidally within a furrow. Sudden changes in the direction of the cell trajectories were generated by transient arrests of the longitudinal flagellum beat, which were accompanied by a switch from the backward orientation to a forward one. This sweeping motion generated the rotation of the cell body. Ca2+ ions highly stimulated the frequencies of this arrest response, which compared to the "walking-stick" behavior of sea urchin spermatozoa. Isolated flagella were ATA-reactivated after detergent treatment. They exhibited 2 types of motion within the same experimental conditions. A progressive helicoidal motion was generated upon longitudinal flagellum reactivation, whereas a rolling motion with little progression characterized transverse flagellum reactivation. The differences in motile behavior reflect regulations of flagellar movement which were not destroyed by the isolation procedure and may be indicative of regulation by accessory structures.

Adenosine Triphosphate↗

PubMedDate not supplied

Electron microscopy of frozen hydrated eukaryotic flagella.

Resting and active sea urchin sperm flagella have been examined by low-dose electron microscopy of frozen hydrated specimens. The flagella are unfixed, unstained, completely intact, and able to swim vigorously after going through the entire preparative procedure. The most prominent features of the image arise from the edges of the axonemal doublets and central-pair microtubules seen in projection. By comparison with these longitudinal markings, transverse features are less easy to discern, being camouflaged by superposition. However, Fourier transforms of digitized micrographs reveal a remarkable degree of crystalline order in quiescent flagella. Filtered images derived from these Fourier transforms show clearly features arising from the central-pair complex and radial spokes that were obscured in the original data. Potentially complicating effects of specimen thickness are shown to be quantitatively insignificant in the formation of images of unstained frozen hydrated flagella. Determination of native flagellar structure by 3-D reconstruction from multiple-tilted views appears to be feasible.

Animals↗

PubMed2005

Rapid decay of Salmonella flagella antibodies during human gastroenteritis: a follow up study.

An indirect enzyme-linked immunosorbent assay (ELISA) based on Salmonella re-polymerized flagella was employed to measure levels of immunoglobulin (Ig) G, IgM and IgA antibodies in sera from 303 Danish patients diagnosed with either Salmonella enteritidis or Salmonella typhimurium. The antibody-levels were assessed at one, three and six months after onset of salmonellosis, and sera from a control-group of 170 healthy blood donors were additionally analysed in order to establish cut-off values for the analysis. Cross-reactions to other Salmonella serotypes, as well as to Escherichia coli, Yersinia enterocolitica, Campylobacter jejuni, Campylobacter coli and Helicobacter pylori were observed. At one month after onset of symptoms, 70% of the patients recovering from a S. enteritidis infection carried detectable levels of anti-flagella antibodies, as did 77% of the patients recovering from S. typhimurium infection. Three months after onset of symptoms these detection rates had decreased to 46% and 40%; and six months after onset of symptoms the detection rates were 34% and 38%. This rapid decrease in the serum levels of flagella antibodies is in conflict with the "common knowledge" statement of a long-lasting anti-flagella immunoresponse. The present study suggests that such a tenacious statement is (or may be) inaccurate.

Adolescent↗

PubMed1971

Contractile mechanisms in flagella.

The elastic theory of flexural waves in thin rods accurately predicts the velocity of flagellar bending waves over a wide range of viscosities. This shows that flagella behave as a purely mechanical system for the transmission of these waves. An evaluation of the total bending moment reveals that this moment occurs in phase over the entire length of a flagellum. From this it is concluded that each contractile fiber in the flagella is activated simultaneously over its whole length. The magnitude of the bending moment decreases linearly along the flagellum. This is most easily explained by a sliding filament hypothesis in flagella with the elementary 9 + 2 fibers. The expression found for the bending moment explains logically that the wave velocity in flagella is determined by their mechanical properties and the outside viscosity only.

Animals↗

PubMed2005

A coherent feed-forward loop with a SUM input function prolongs flagella expression in Escherichia coli.

Complex gene-regulation networks are made of simple recurring gene circuits called network motifs. The functions of several network motifs have recently been studied experimentally, including the coherent feed-forward loop (FFL) with an AND input function that acts as a sign-sensitive delay element. Here, we study the function of the coherent FFL with a sum input function (SUM-FFL). We analyze the dynamics of this motif by means of high-resolution expression measurements in the flagella gene-regulation network, the system that allows Escherichia coli to swim. In this system, the master regulator FlhDC activates a second regulator, FliA, and both activate in an additive fashion the operons that produce the flagella motor. We find that this motif prolongs flagella expression following deactivation of the master regulator, protecting flagella production from transient loss of input signal. Thus, in contrast to the AND-FFL that shows a delay following signal activation, the SUM-FFL shows delays after signal deactivation. The SUM-FFL in this system works as theoretically predicted despite being embedded in at least two additional feedback loops. The present function might be carried out by the SUM-FFL in systems found across organisms.

Arabinose↗

PubMed2003

Perspectives on the origin of microfilaments, microtubules, the relevant chaperonin system and cytoskeletal motors--a commentary on the spirochaete origin of flagella.

The origin of cytoskeleton and the origin of relevant intracellular transportation system are big problems for understanding the emergence of eukaryotic cells. The present article summarized relevant information of evidences and molecular traces on the origin of actin, tubulin, the chaperonin system for folding them, myosins, kinesins, axonemal dyneins and cytoplasmic dyneins. On this basis the authors proposed a series of works, which should be done in the future, and indicated the ways for reaching the targets. These targets are mainly: 1) the reconstruction of evolutionary path from MreB protein of archaeal ancestor of eukaryotic cells to typical actin; 2) the finding of the MreB or MreB-related proteins in crenarchaea and using them to examine J. A. Lake's hypothesis on the origin of eukaryote from "eocytes" (crenarchaea); 3) the examinations of the existence and distribution of cytoskeleton made of MreB-related protein within coccoid archaea, especially in amoeboid archaeon Thermoplasm acidophilum; 4) using Thermoplasma as a model of archaeal ancestor of eukaryotic cells; 5) the searching for the homolog of ancestral dynein in present-day living archaea. During the writing of this article, Margulis' famous spirochaete hypothesis on the origin of flagella and cilia was unexpectedly involved and analyzed from aspects of tubulins, dyneins and spirochaetes. Actually, spirochaete cannot be reasonably assumed as the ectosymbiotic ancestor of eukaryotic flagella and cilia, since their swing depends upon large amount of bacterial flagella beneath the flexible outer wall, but not depends upon their intracellular tubules and the assumed dyneins. In this case, if they had "evolved" into cilia and lost their bacterial flagella, they would immediately become immobile! In fact, tubulin and dynein-like proteins have not been found in any spirochaete.

Actin Cytoskeleton↗

PubMed1998

Genetic analysis of flagellar length control in Chlamydomonas reinhardtii: a new long-flagella locus and extragenic suppressor mutations.

Flagellar length in the biflagellate alga Chlamydomonas reinhardtii is under constant and tight regulation. A number of mutants with defects in flagellar length control have been previously identified. Mutations in the three long-flagella (lf) loci result in flagella that are up to three times longer than wild-type length. In this article, we describe the isolation of long-flagellar mutants caused by mutations in a new LF locus, LF4. lf4 mutations were shown to be epistatic to lf1, while lf2 was found to be epistatic to lf4 with regard to the flagellar regeneration defect. Mutations in lf4 were able to suppress the synthetic flagella-less phenotype of the lf1, lf2 double mutant. In addition, we have isolated four extragenic suppressor mutations that suppress the long-flagella phenotype of lf1, lf2, or lf3 double mutants.

Animals↗

PubMed2003

Effects of mupirocin at subinhibitory concentrations on flagella formation in Pseudomonas aeruginosa and Proteus mirabilis.

Colonization of Pseudomonas aeruginosa at trachea, nares and oropharynx can cause ventilator-acquired pneumonia. To identify beneficial effects of antibiotics on expression of virulence factors related to colonization by such pathogens, we evaluated the effect of mupirocin on flagella formation in P. aeruginosa and on motility and flagella formation in Proteus mirabilis. In P. aeruginosa, subinhibitory concentrations of mupirocin inhibited flagella formation, which was associated with reduced flagellin expression. In P. mirabilis, subinhibitory concentrations of mupirocin dose-dependently suppressed bacterial motility and flagella formation, again with reduced flagellin expression. Our results indicate that subinhibitory concentrations of mupirocin can suppress expression of virulence factors in P. aeruginosa and P. mirabilis.

Anti-Bacterial Agents↗

PubMed1986

Experimental studies of the pathogenesis of infections due to Pseudomonas aeruginosa: immunization using divalent flagella preparations.

Flagella preparations of the two antigenic types that make up all the flagella antigens in Pseudomonas aeruginosa were used, individually, to immunize mice. Mice thus immunized were protected when burned and infected with P. aeruginosa. Protection was specific for the flagella antigen of the challenge strain but independent of its somatic antigen. When both antigens were used, together, for immunization, protection was independent of both the flagella and somatic antigen of the challenge strain. These results suggest that a divalent P. aeruginosa 'vaccine' would be of considerable value for immunoprophylaxis in burn patients.

Animals↗

PubMed2000

The role of fimbriae and flagella in the adherence of avian strains of Escherichia coli O78:K80 to tissue culture cells and tracheal and gut explants.

To investigate the role of fimbriae and flagella in the pathogenesis of avian colibacillosis, isogenic insertionally inactivated mutant strains of Escherichia coli O78:K80 strain EC34195 defective in the elaboration of type-1 and curli fimbriae and flagella were constructed by allelic exchange. Single and multiple non-fimbriate and non-flagellate mutant strains were compared to the wild-type in vitro in adherence assays with a HEp-2 cell line, a mucus-secreting cell line HT2916E, a non-mucus-secreting cell line HT2919A, tracheal explant and proximal gut explant. Mutant strains defective in the elaboration of type-1 fimbriae were significantly less adherent--in the order of 90% reduction--than the wild-type strain in all assays. Mutant strains defective in the elaboration of flagella were generally as adherent as the wild-type strain except when assayed with the mucus-secreting cell line HT2916E, for which a significant reduction of adherence--of the order of 90%--compared with the wild-type strain was observed. Mutant strains defective for the elaboration of curli fimbriae adhered as well as the wild-type strain in all assays, except when assayed in tests with gut explant tissue for which a significant reduction of adherence--of the order of 80%--compared with the wild-type strain was observed. Adherence to explants was to epithelial, not serous, surfaces and was 10-fold greater to tracheal than to gut explants. Together, these data support the hypothesis that type-1 fimbriae are significant factors in adherence, aided by flagella for penetration of mucus and curli fimbriae for adherence to the gut.

Animals↗

PubMed2002

A novel paralogous gene family involved in phase-variable flagella-mediated motility in Campylobacter jejuni.

Flagella-mediated motility is recognized to be one of the major factors contributing to virulence in Campylobacter jejuni. Motility of this bacterium is known to be phase variable, although the mechanism of such variation remains unknown. C. jejuni genome sequencing revealed a number of genes prone to phase variation via a slipped-strand mispairing mechanism. Many of these genes are hypothetical and are clustered in the regions involved in formation of three major cell surface structures: capsular polysaccharide, lipooligosaccharide and flagella. Among the genes of unknown function, the flagellar biosynthesis and modification region contains seven hypothetical paralogous genes designated as the motility accessory factor (maf) family. Remarkably, two of these genes (maf1 and maf4) were found to be identical and both contain homopolymeric G tracts. Using insertional mutagenesis it was demonstrated that one of the genes, maf5, is involved in formation of flagella. Phase variation of the maf1 gene via slipped-strand mispairing partially restored motility of the maf5 mutant. The maf family represents a new class of bacterial genes related to flagellar biosynthesis and phase variation. Reversible expression of flagella may be advantageous for the adaptation of C. jejunito the varied in vivo and ex vivo environments encountered during its life cycle, as well in evasion of the host immune response.

Amino Acid Sequence↗