Electron tomography of mitochondria after the arrest of protein import associated with Tom19 depletion

doi:10.1078/0171-9335-00149

European Journal of Cell Biology

Volume 80, Issue 2, February 2001, Pages 139-150

https://doi.org/10.1078/0171-9335-00149 Get rights and content

Summary

In a mutant form of Neurospora crassa, in which sheltered RIP (repeat induced point mutation) was used to deplete Tom19, protein transport through the TOM/TIM pathway is arrested by the addition of p-fluorophenylalanine (FPA). Using intermediate-voltage electron tomography, we have generated three-dimensional reconstructions of 28 FPA-treated mitochondria at four time points (0 – 32 h) after the addition of FPA. We determined that the cristae surface area and volume were lost in a roughly linear manner. A decrease in mitochondrial volume was not observed until after 16 h of FPA treatment. The inner boundary membrane did not appear to shrink or contract away from the outer membrane. Interestingly, the close apposition of these membranes remained over the entire periphery, even after all of the cristae had disappeared. The different dynamics of the shrinkage of cristae membrane and inner boundary membrane has implications for compartmentalization of electron transport proteins. Two structurally distinct types of contact sites were observed, consistent with recently published work. We determined that the cristae in the untreated (control) mitochondria are all lamellar. The cristae of FPA-treated mitochondria retain the lamellar morphology as they reduce in size and do not adopt tubular shapes. Importantly, the crista junctions exhibit tubular as well as slot-like connections to the inner boundary membrane, persisting until the cristae disappear, indicating that their stability is not dependent on continuous protein import through the complex containing Tom19.

References (38)

E. Bullitt et al.
The yeast spindle pole body is assembled around a central crystal of Spc42p
Cell
(1997)
J. Frank
Approaches to large-scale structures
Curr. Opin. Struct. Biol.
(1995)
T.G. Frey et al.
The internal structure of mitochondria
Trends Biochem. Sci.
(2000)
R.A. Horowitz et al.
Automated electron microscope tomography of frozen-hydrated chromatin: The irregular three-dimensional zigzag architecture persists in compact, isolated fibers
J. Struct. Biol.
(1997)
A.J. Koster et al.
Perspectives of molecular and cellular electron tomography
J. Struct. Biol.
(1997)
C.A. Mannella et al.
Reconsidering mitochondrial structure: new views of an old organelle
Trends Biochem. Sci.
(1997)
D. Nicastro et al.
Cryoelectron tomography of Neurospora mitochondria
J. Struct. Biol.
(2000)
P. Penczek et al.
Double-tilt electron tomography
Ultramicroscopy
(1995)
G. Perkins et al.
Recent structural insight into mitochondria gained by microscopy
Micron
(2000)
G. Perkins et al.
Electron tomography of neuronal mitochondria: Three-dimensional structure and organization of cristae and membrane contacts
J. Struct. Biol.
(1997)

G. Perkins et al.

Electron tomography of large, multicomponent biological structures

J. Struct. Biol.

(1997)

J.P. Schroeter et al.

SUPRIM: Easily modified image processing software

J. Struct. Biol.

(1996)

F.S. Sjostrand et al.

Alongrange ordered structure in mitochondrial cristae revealed by a pathological structural modification.

J. Ultrastruct. Mol. Struct. Res.

(1988)

G.P. Dempsey et al.

A copper block method for freezing non-cryoprotected tissue to produce ice-crystal-free regions for electron microscopy.II Evaluation using freeze fracturing with a cryo-ultramicrotome

J. Microsc.

(1976)

G.Y. Fan et al.

Conditions for electron tomographic data acquisition

J. Electron Microsc.

(1995)

J. Frank et al.

Alignment by cross-correlation.

T.G. Frey

Cytochrome c oxidase: Structural studies by electron microscopy of two-dimensional crystals

Microsc. Res. Tech.

(1994)

C.R. Hackenbrock

Ultrastructural bases for metabolically linked mechanical activity in mitochondria I. Reversible ultrastructural changes with change in metabolic steady state in isolated liver mitochondria

J. Cell Biol.

(1966)

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Electron tomography of mitochondria after the arrest of protein import associated with Tom19 depletion

Summary

Cell

Curr. Opin. Struct. Biol.

Trends Biochem. Sci.

J. Struct. Biol.

J. Struct. Biol.

Trends Biochem. Sci.

J. Struct. Biol.

Ultramicroscopy

Micron

J. Struct. Biol.

J. Struct. Biol.

J. Struct. Biol.

J. Ultrastruct. Mol. Struct. Res.

A copper block method for freezing non-cryoprotected tissue to produce ice-crystal-free regions for electron microscopy.II Evaluation using freeze fracturing with a cryo-ultramicrotome

J. Microsc.

Conditions for electron tomographic data acquisition

J. Electron Microsc.

Alignment by cross-correlation.

Cytochrome c oxidase: Structural studies by electron microscopy of two-dimensional crystals

Microsc. Res. Tech.

Ultrastructural bases for metabolically linked mechanical activity in mitochondria I. Reversible ultrastructural changes with change in metabolic steady state in isolated liver mitochondria

J. Cell Biol.