Characterization of the cytoplasmic fibrils of Treponema refringens (Nichols)

Eipert, Susan R.; Black, S. H.

doi:10.1007/bf00423067

Cited by 17 publications

(25 citation statements)

References 29 publications

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“…1), the ribbon shape and apparent lack of a lumen in the usual sense indicate that these structures are more properly referred to as filaments or fibrils rather than tubules. The cytoplasmic filaments are difficult to visualize by electron microscopy of ultrathin sections (5,19,29,41), but they appear to run as a bundle, with the wide cross-sectional dimension of each filament in close apposition with that of the neighboring filament (41). This array is closely associated with the inner membrane (perhaps indicating a specific interaction with inner membrane lipids or proteins), and is always located just underneath the cluster of periplasmic flagella.…”

Section: Discussionmentioning

confidence: 99%

“…The function of the cytoplasmic filaments is unknown. Their location indicates that they may be involved in cell motility, although other functions related to helical morphology or cell division are also possible (5). Masuda and Kawata (25) purified the cytoplasmic filaments of Treponema denticola TD-2, Treponema phagedenis biotypes Reiter and Kazan, three Treponema sp.…”

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Characterization of the cytoplasmic filament protein gene (cfpA) of Treponema pallidum subsp. pallidum

You¹,

Elmore²,

Colton³

et al. 1996

J Bacteriol

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Treponema pallidum and other members of the genera Treponema, Spirochaeta, and Leptonema contain multiple cytoplasmic filaments that run the length of the organism just underneath the cytoplasmic membrane. These cytoplasmic filaments have a ribbon-like profile and consist of a major cytoplasmic filament protein subunit (CfpA, formerly called TpN83) with a relative molecular weight of ϳ80,000. Degenerate DNA primers based on N-terminal and CNBr cleavage fragment amino acid sequences of T. pallidum subsp. pallidum (Nichols) CfpA were utilized to amplify a fragment of the encoding gene (cfpA). A 6.8-kb EcoRI fragment containing all but the 5 end of cfpA was identified by hybridization with the resulting PCR product and cloned into Lambda ZAP II. The 5 region was obtained by inverse PCR, and the complete gene sequence was determined. The cfpA sequence contained a 2,034-nucleotide coding region, a putative promoter with consensus sequences (5-TTTACA-3 for ؊35 and 5-TACAAT-3 for ؊10) similar to the 70 recognition sequence of Escherichia coli and other organisms, and a putative ribosome-binding site (5-AGGAG-3). The deduced amino acid sequence of CfpA indicated a protein of 678 residues with a calculated molecular mass of 78.5 kDa and an estimated pI of 6.15. No significant homology to known proteins or structural motifs was found among known prokaryotic or eukaryotic sequences. Expression of a LacZ-CfpA fusion protein in E. coli was detrimental to survival and growth of the host strain and resulted in the formation of short, irregular filaments suggestive of partial self-assembly of CfpA. The cytoplasmic filaments of T. pallidum and other spirochetes appear to represent a unique form of prokaryotic intracytoplasmic inclusions.Spirochetes possess a number of unusual structural features, including helical or flat plane-wave shape and periplasmic flagella that tightly entwine the cell body and are encased by the outer membrane (13-16, 19, 31, 41). T. pallidum differs from other spirochetes in that it contains relatively few intramembranous protein particles in its outer membrane, as determined by freeze-fracture electron microscopy (33, 40). Seven of eight Treponema species examined, some Spirochaeta species, and Leptonema illini have also been shown to contain cytoplasmic filaments (also called cytoplasmic tubules and cytoplasmic fibrils) (14). Cytoplasmic filaments are ribbon-like structures 7.0 to 7.5 nm wide that run the length of the organism (5, 13-16, 19, 25, 31, 41). An array of 4 to 6 filaments lie in close apposition to the inner membrane and are always localized directly underneath the corresponding group of periplasmic flagella. The function of the cytoplasmic filaments is unknown. Their location indicates that they may be involved in cell motility, although other functions related to helical morphology or cell division are also possible (5).Masuda and Kawata (25) purified the cytoplasmic filaments of Treponema denticola TD-2, Treponema phagedenis biotypes Reiter and Kazan, three Treponema sp. strains (E-21,...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Characterization of the cytoplasmic filament protein gene (cfpA) of Treponema pallidum subsp. pallidum

You¹,

Elmore²,

Colton³

et al. 1996

J Bacteriol

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“…They are located underneath the periplasmic flagellar bundle in close apposition to the cytoplasmic membrane (8,18). The filaments are composed of one major protein (33) that is well conserved among species (21), and the gene encoding them, cfpA, is preceded by a sigma 70 promoter (21,53).…”

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Cytoplasmic Filament-Deficient Mutant of Treponema denticola Has Pleiotropic Defects

2001

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In Treponema denticola, a ribbon-like structure of cytoplasmic filaments spans the cytoplasm at all stages of the cell division process. Insertional inactivation was used as a first step to determine the function of the cytoplasmic filaments. A suicide plasmid was constructed that contained part of cfpA and a nonpolar erythromycin resistance cassette (ermF and ermAM) inserted near the beginning of the gene. The plasmid was electroporated into T. denticola, and double-crossover recombinants which had the chromosomal copy of cfpA insertionally inactivated were selected. Immunoblotting and electron microscopy confirmed the lack of cytoplasmic filaments. The mutant was further analyzed by dark-field microscopy to determine cell morphology and by the binding of two fluorescent dyes to DNA to assess the distribution of cellular nucleic acids. The cytoplasmic filament protein-deficient mutant exhibited pleiotropic defects, including highly condensed chromosomal DNA, compared to the homogeneous distribution of the DNA throughout the cytoplasm in a wild-type cell. Moreover, chains of cells are formed by the cytoplasmic filament-deficient mutant, and those cells show reduced spreading in agarose, which may be due to the abnormal cell length. The chains of cells and the highly condensed chromosomal DNA suggest that the cytoplasmic filaments may be involved in chromosome structure, segregation, or the cell division process in Treponema.

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“…However, only one electron microscopic study using the isolated preparation showed that the CF from T. refringens was a complex arrangement of thinner strands, but not a tubular structure; each CF was constructed from four strands arranged at each corner of a rectangle in cross-sectional view (5). We demonstrated here by electron and immunoelectron microscopic observations that the CF of Treponema sp.…”

Section: Discussionmentioning

confidence: 71%

“…The CFs helically wind as a bundle in the protoplasmic cylinder, and have been considered to be a tubular structure, because negative stains have penetrated the lumen-like region of the CFs (8,13,25). The CFs have been hitherto isolated only from one treponeme, T. refringens, and each fibril was shown to be a complex arrangement of strands rather than a tubule by electron microscopy of the negatively stained isolated CFs (5).…”

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Isolation and Characterization of Cytoplasmic Fibrils from Treponemes

Masuda¹,

Kawata²

1989

Microbiology and Immunology

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Electron microscopy of Triton X-100-treated whole cells of an oral treponeme, Treponema sp. strain E-21, revealed that six cytoplasmic fibrils (CFs) helically wound as a bundle in the cytoplasm. The CFs were isolated and purified by disruption and solubilization of the cells followed by CsCl density gradient centrifugation. The purified CF preparation contained mostly fibrils of about 9 nm in width and very small amounts of thinner strands of about 3 nm in diameter. The CFs were apparently seen to be a tubular structure, but the isolated CFs had narrowed sites of about 4-5 nm in width lacking lumen-like images, possibly representing twisted sites. Thus, the CF did not seem to be a tubular structure. The purified CFs were composed of one major 82 kDa protein and a few minor proteins. The CFs were destructed by treatment with proteases, 8 M urea or 4 M guanidine hydrochloride. Very low tyrosine content (0.76 mol %) and lack of methionine were characteristic features for the 82 kDa protein. The CF preparations from the other five treponemes including Treponema phagedenis and T. denticola also had 82 kDa proteins as a major component, and the 82 kDa proteins of all of the treponemes had a common antigen when examined by using antiserum against the 82 kDa protein from Treponema sp. strain E-21. Furthermore, the 82 kDa protein was demonstrated to be a principal component of the CFs of all the treponemes by immunoelectron microscopy.

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Characterization of the cytoplasmic fibrils of Treponema refringens (Nichols)

Cited by 17 publications

References 29 publications

Characterization of the cytoplasmic filament protein gene (cfpA) of Treponema pallidum subsp. pallidum

Characterization of the cytoplasmic filament protein gene (cfpA) of Treponema pallidum subsp. pallidum

Cytoplasmic Filament-Deficient Mutant of Treponema denticola Has Pleiotropic Defects

Isolation and Characterization of Cytoplasmic Fibrils from Treponemes

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