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The supramolecular organization of fibrillin-rich microfibrils.

Baldock C, Koster AJ, Ziese U, Rock MJ, Sherratt MJ, Kadler KE, Shuttleworth CA, Kielty CM - J. Cell Biol. (2001)

Bottom Line: Mass mapping shows that, in solution, microfibrils with periodicities of <70 and >140 nm are stable, but periodicities of approximately 100 nm are rare.Microfibrils comprise two in-register filaments with a longitudinal symmetry axis, with eight fibrillin molecules in cross section.We present a model of fibrillin alignment that fits all the data and indicates that microfibril extensibility follows conformation-dependent maturation from an initial head-to-tail alignment to a stable approximately one-third staggered arrangement.

View Article: PubMed Central - PubMed

Affiliation: Wellcome Trust Centre for Cell-Matrix Research, Schools of Biological Sciences and Medicine, University of Manchester, Manchester, M13 9PT, United Kingdom. clair.baldock@man.ac.uk

ABSTRACT
We propose a new model for the alignment of fibrillin molecules within fibrillin microfibrils. Automated electron tomography was used to generate three-dimensional microfibril reconstructions to 18.6-A resolution, which revealed many new organizational details of untensioned microfibrils, including heart-shaped beads from which two arms emerge, and interbead diameter variation. Antibody epitope mapping of untensioned microfibrils revealed the juxtaposition of epitopes at the COOH terminus and near the proline-rich region, and of two internal epitopes that would be 42-nm apart in unfolded molecules, which infers intramolecular folding. Colloidal gold binds microfibrils in the absence of antibody. Comparison of colloidal gold and antibody binding sites in untensioned microfibrils and those extended in vitro, and immunofluorescence studies of fibrillin deposition in cell layers, indicate conformation changes and intramolecular folding. Mass mapping shows that, in solution, microfibrils with periodicities of <70 and >140 nm are stable, but periodicities of approximately 100 nm are rare. Microfibrils comprise two in-register filaments with a longitudinal symmetry axis, with eight fibrillin molecules in cross section. We present a model of fibrillin alignment that fits all the data and indicates that microfibril extensibility follows conformation-dependent maturation from an initial head-to-tail alignment to a stable approximately one-third staggered arrangement.

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Automated electron tomography images of negatively stained untensioned bovine zonular microfibrils. (A) A zero degree tilt image of a negatively stained microfibril. (B) XY slices at six different Z heights through a reconstruction showing a region of the same microfibril. Stain-penetrable space between the arms is seen in all Z slices and is therefore a 3-D feature. (C) Three Z slices shown in black and white using IMOD (Kremer et al.. 1996) compared with the same region of microfibril 3-D volume rendered (green) using AVS Express (Advanced Visual Systems Inc.). (D) Schematic diagram of a microfibril repeating unit with the mean measurements taken from a representative data set. For standard deviations, see Table . (E) Colloidal gold binding to microfibrils, showing periodic double labeling at the ends of the interbead arms, and intermittent labeling at the bead. Beads and interbeads are labeled B and IB, respectively.
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Figure 1: Automated electron tomography images of negatively stained untensioned bovine zonular microfibrils. (A) A zero degree tilt image of a negatively stained microfibril. (B) XY slices at six different Z heights through a reconstruction showing a region of the same microfibril. Stain-penetrable space between the arms is seen in all Z slices and is therefore a 3-D feature. (C) Three Z slices shown in black and white using IMOD (Kremer et al.. 1996) compared with the same region of microfibril 3-D volume rendered (green) using AVS Express (Advanced Visual Systems Inc.). (D) Schematic diagram of a microfibril repeating unit with the mean measurements taken from a representative data set. For standard deviations, see Table . (E) Colloidal gold binding to microfibrils, showing periodic double labeling at the ends of the interbead arms, and intermittent labeling at the bead. Beads and interbeads are labeled B and IB, respectively.

Mentions: Negatively stained microfibrils showed repeating units of “beads” and “interbeads,” a diameter of 15–18 nm, and a mean periodicity of 57.5 nm (Fig. 1, a–d, and Table ). The interbeads often appeared to bow out between beads. Average bead height was 9 nm, calculated using 5 nm gold for comparison.


The supramolecular organization of fibrillin-rich microfibrils.

Baldock C, Koster AJ, Ziese U, Rock MJ, Sherratt MJ, Kadler KE, Shuttleworth CA, Kielty CM - J. Cell Biol. (2001)

Automated electron tomography images of negatively stained untensioned bovine zonular microfibrils. (A) A zero degree tilt image of a negatively stained microfibril. (B) XY slices at six different Z heights through a reconstruction showing a region of the same microfibril. Stain-penetrable space between the arms is seen in all Z slices and is therefore a 3-D feature. (C) Three Z slices shown in black and white using IMOD (Kremer et al.. 1996) compared with the same region of microfibril 3-D volume rendered (green) using AVS Express (Advanced Visual Systems Inc.). (D) Schematic diagram of a microfibril repeating unit with the mean measurements taken from a representative data set. For standard deviations, see Table . (E) Colloidal gold binding to microfibrils, showing periodic double labeling at the ends of the interbead arms, and intermittent labeling at the bead. Beads and interbeads are labeled B and IB, respectively.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2198817&req=5

Figure 1: Automated electron tomography images of negatively stained untensioned bovine zonular microfibrils. (A) A zero degree tilt image of a negatively stained microfibril. (B) XY slices at six different Z heights through a reconstruction showing a region of the same microfibril. Stain-penetrable space between the arms is seen in all Z slices and is therefore a 3-D feature. (C) Three Z slices shown in black and white using IMOD (Kremer et al.. 1996) compared with the same region of microfibril 3-D volume rendered (green) using AVS Express (Advanced Visual Systems Inc.). (D) Schematic diagram of a microfibril repeating unit with the mean measurements taken from a representative data set. For standard deviations, see Table . (E) Colloidal gold binding to microfibrils, showing periodic double labeling at the ends of the interbead arms, and intermittent labeling at the bead. Beads and interbeads are labeled B and IB, respectively.
Mentions: Negatively stained microfibrils showed repeating units of “beads” and “interbeads,” a diameter of 15–18 nm, and a mean periodicity of 57.5 nm (Fig. 1, a–d, and Table ). The interbeads often appeared to bow out between beads. Average bead height was 9 nm, calculated using 5 nm gold for comparison.

Bottom Line: Mass mapping shows that, in solution, microfibrils with periodicities of <70 and >140 nm are stable, but periodicities of approximately 100 nm are rare.Microfibrils comprise two in-register filaments with a longitudinal symmetry axis, with eight fibrillin molecules in cross section.We present a model of fibrillin alignment that fits all the data and indicates that microfibril extensibility follows conformation-dependent maturation from an initial head-to-tail alignment to a stable approximately one-third staggered arrangement.

View Article: PubMed Central - PubMed

Affiliation: Wellcome Trust Centre for Cell-Matrix Research, Schools of Biological Sciences and Medicine, University of Manchester, Manchester, M13 9PT, United Kingdom. clair.baldock@man.ac.uk

ABSTRACT
We propose a new model for the alignment of fibrillin molecules within fibrillin microfibrils. Automated electron tomography was used to generate three-dimensional microfibril reconstructions to 18.6-A resolution, which revealed many new organizational details of untensioned microfibrils, including heart-shaped beads from which two arms emerge, and interbead diameter variation. Antibody epitope mapping of untensioned microfibrils revealed the juxtaposition of epitopes at the COOH terminus and near the proline-rich region, and of two internal epitopes that would be 42-nm apart in unfolded molecules, which infers intramolecular folding. Colloidal gold binds microfibrils in the absence of antibody. Comparison of colloidal gold and antibody binding sites in untensioned microfibrils and those extended in vitro, and immunofluorescence studies of fibrillin deposition in cell layers, indicate conformation changes and intramolecular folding. Mass mapping shows that, in solution, microfibrils with periodicities of <70 and >140 nm are stable, but periodicities of approximately 100 nm are rare. Microfibrils comprise two in-register filaments with a longitudinal symmetry axis, with eight fibrillin molecules in cross section. We present a model of fibrillin alignment that fits all the data and indicates that microfibril extensibility follows conformation-dependent maturation from an initial head-to-tail alignment to a stable approximately one-third staggered arrangement.

Show MeSH
Related in: MedlinePlus