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The molecular machinery of mitochondrial morphology and inheritance

Benedikt Westermann

Introduction

Each type of eukaryotic cell possesses a characteristic three-dimensional structure. Maintenance of its architecture and duplication during cell division depend on active transport of organelles along the cytoskeleton as well as continuous fission and fusion events of intracellular membranes. Mitochondria are essential organelles which are often located at sites of high energy consumption in the cell. They cannot be formed de novo, but have to be inherited from the mother to the daughter cell during cell division. There is mounting evidence that positioning and transport of mitochondria are controlled by the cytoskeleton and depend on fusion and fission of their membranes.

We use the budding yeast Saccharomyces cerevisiae and the filamentous fungus Neurospora crassa to study these processes. Both are excellent model organisms since genetic and biochemical techniques can be readily combined. The genomes of both fungi are completely sequenced, genetic engineering is extremely simple in yeast, and Neurospora has great advantages for biochemistry. Mitochondria can be easily visualized in living cells using the green fluorescent protein fused to a mitochondrial targeting sequence (Westermann and Neupert, 2000; Fuchs et al., 2002).

Why is it important to study mitochondrial dynamics?

First of all, because it comprises a fascinating basic cell biological problem. By studying mitochondrial morphology and inheritance we can learn a lot about many different aspects of organellar biogenesis, such as partitioning of organelles during cell division, transport of membranes along the cytoskeleton, intracellular membrane fusion, regulatory processes etc.

Mitochondrial dynamics is directly important for cellular function. Just to give a few examples: Fusion of mitochondria is essential for the development of spermatids in Drosophila, mitochondrial division is important for apoptosis (Jagasia et al., 2005), and mitochondrial fusion is required in human cells to allow extensive complementation of mitochondrial genomes as a defense against the accumulation of oxidative damage during cellular ageing.

Last but not least, mitochondrial morphogenesis is pathologically relevant. Aberrant mitochondria are associated with cardiomyopathy and cancer, and OPA1, a gene coding for a protein important for mitochondrial morphogenesis, is mutated in patients suffering from childhood blindness.

Projects

Fusion of mitochondria

Fusion of mitochondria has been observed in a great variety of different cell types. In yeast and many mammalian cell types, mitochondria form a giant branched network, the continuity of which is maintained by a balanced frequency of fission and fusion events (for review see: Westermann, 2002; Westermann, 2003). The first known protein component mediating mitochondrial membrane fusion was identified in Drosophila: The Fzo protein is the founding member of a novel conserved protein familiy of large membrane-bound GTPases. Its homolog in yeast, Fzo1, is essential for the biogenesis of functional mitochondria. It is part of a high-molecular weight complex in the mitochondrial outer membrane (Rapaport et al., 1998). In contrast to other intracellular membrane fusion events – such as fusion of enveloped viruses with host cell membranes or SNARE-mediated vesicle fusion in the secretory pathway (Weber et al., 1998) – mitochondria face a topological problem: Each organelle is bounded by two membranes; hence, four membranes have to be fused in a coordinated manner. This dilemma is solved by an interaction of the outer membrane fusion machinery with the inner membrane of mitochondria. This connection of the membranes is mediated by Fzo1 and allows a coordinated fusion of the double membranes (Fritz et al., 2001). Further elucidation of the molecular mechanisms of mitochondrial fusion will shed light not only on an important aspect of the biogenesis of this organelle, but may reveal also mechanistic aspects of the fusion of cellular membranes in general.

Interaction of mitochondria with the cytoskeleton

The interaction of mitochondria with the cytoskeleton plays a key role in their motility, positioning, and morphology (Westermann and Prokisch, 2002). In Saccharomyces cerevisiae, the branched mitochondrial network below the cell cortex is maintained by an interaction with the actin cytoskeleton. In Neurospora, however, mitochondria are transported by kinesin-related motor proteins along microtubules (Fuchs et al., 2002; Fuchs and Westermann, 2005), as it is the case in many mammalian cell types. One key component of mitochondrial morphology in Neurospora is the MMM1 protein, an integral protein of the outer membrane. Knock out mutants harbor only few but extremely enlarged mitochondria with defective motility (Prokisch et al., 2000). It is a matter of debate whether MMM1 is a component of a mitochondrial receptor protein complex that recruits molecular motor proteins to the organelle. The identification and functional characterization of the molecular components mediating mitochondrial transport in Neurospora may also contribute to our understanding of the mechanisms of organelle transport in higher eukaryotic cells.

Genetic basis of mitochondrial function and morphology in yeast

The understanding of the processes underlying organellar function and inheritance requires the identification and characterization of the molecular components involved. During the past few decades, many of the proteins required for establishment and maintenance of mitochondrial structure have been identified in yeast. The advent of the post-genomic era allows us to conduct systematic genome-wide screens to define whole complements of genes associated with particular functions. With the systematic screening of comprehensive mutant libraries covering ca. 90% of the Saccharomyces cerevisiae genome, we obtained a comprehensive picture of the cellular processes and molecular components required for mitochondrial function and structure in a simple eukaryotic cell (Dimmer et al., 2002; Altmann and Westermann, 2005). Among the newly identified proteins is the first known component involved in division of the mitochondrial inner membrane (Messerschmitt et al., 2003), a novel subunit of a ubiquitin ligase that is involved in regulation of the turnover of the mitochondrial fusion machinery (Fritz et al., 2003), and a novel family of inner membrane proteins coordinationg inheritance of mitochondria and mitochondrial DNA (Dimmer et al., 2005). Furthermore, filamentous actin, proteasome-mediated protein degradation, the mitochondrial protein import machinery, the vesicular trafficking system and ergosterol biosynthesis are of fundamental importance for maintenance of mitochondrial morphology (Altmann and Westermann, 2005). It will be a major challenge for the future to establish the molecular functions of the other newly discovered proteins and to unravel the interactions among the components of mitochondrial behavior. These studies will certainly improve our understanding of the mechanisms that shape this complex double membrane-bounded organelle.

Selected publications

Altmann, K. and Westermann, B. (2005). Role of essential genes in mitochondrial morphogenesis in Saccharomyces cerevisiae. Mol. Biol. Cell 16: 5410-5417.

Dimmer, K.S., Fritz, S., Fuchs, F., Messerschmitt, M., Weinbach, N., Neupert, W., and Westermann, B. (2002). Genetic basis of mitochondrial function and morphology in Saccharomyces cerevisiae. Mol. Biol. Cell 13: 847-853.

Dimmer, K.S., Jakobs, S., Vogel, F., Altmann, K., and Westermann, B. (2005). Mdm31 and Mdm32 are inner membrane proteins required for maintenance of mitochondrial shape and stability of mitochondrial DNA nucleoids in yeast. J. Cell Biol. 168:103-115.

Fritz, S., Rapaport, D., Klanner, E., Neupert, W., and Westermann, B. (2001). Connection of the mitochondrial outer and inner membranes by Fzo1 is critical for organellar fusion. J. Cell Biol. 152: 683-692.

Fritz, S., Weinbach, N., and Westermann, B. (2003). Mdm30 is an F-box protein required for maintenance of fusion-competent mitochondria in yeast. Mol. Biol. Cell. 14: 2303-2313.

Fuchs, F., Prokisch, H., Neupert, W., and Westermann, B. (2002). Interaction of mitochondria with microtubules in the filamentous fungus Neurospora crassa. J. Cell Sci. 115: 1931-1937.

Fuchs, F. and Westermann, B. (2005). Role of Unc104/KIF1-related motor proteins in mitochondrial transport in Neurospora crassa. Mol. Biol. Cell 16:153-161.

Jagasia, R., Grote, P., Westermann, B., and Conradt, B. (2005). DRP-1-mediated mitochondrial fragmentation during EGL-1-induced cell death in C. elegans. Nature 433: 754-760.

Jakobs, S., Martini, N., Schauss, A., Egner, F., Westermann, B., and Hell, S.W. (2003). Spatial and temporal dynamics of budding yeast mitochondria lacking the division component Fis1p. J. Cell Sci., 116: 2005-2014.

Messerschmitt, M., Jakobs, S., Vogel, F., Fritz, S., Dimmer, K.S., Neupert, W., and Westermann, B. (2003). The inner membrane protein Mdm33 controls mitochondrial morphology in yeast. J. Cell Biol. 160: 553-564.

Prokisch, H., Neupert, W., and Westermann, B. (2000). Role of MMM1 in maintaining mitochondrial morphology in Neurospora crassa. Mol. Biol. Cell 11: 2961-2971.

Rapaport, D., Brunner, M., Neupert, W., and Westermann, B. (1998). Fzo1p is a mitochondrial outer membrane protein essential for the biogenesis of functional mitochondria in Saccharomyces cerevisiae. J. Biol. Chem. 273: 20150-20155.

Weber, T., Zemelman, B.V., McNew, J.A., Westermann, B., Gmachl, M., Parlati, F., Söllner, T.H., and Rothman, J.E. (1998). SNAREpins: Minimal machinery for membrane fusion. Cell 92: 759-772.

Westermann, B. (2002). Merging mitochondria matters. Cellular role and molecular machinery of mitochondrial fusion. EMBO Rep. 3: 527-531 (review article).

Westermann, B. (2003). Mitochondrial membrane fusion. Biochim. Biophys. Acta. 1641: 195-202 (review article).

Westermann, B. and Neupert, W. (2000). Mitochondria-targeted green fluorescent proteins: convenient tools for the study of organelle biogenesis in Saccharomyces cerevisiae. Yeast 16: 1421-1427.

Westermann, B. and Prokisch, H. (2002). Mitochondrial dynamics in filamentous fungi. Fungal Genet. Biol. 36: 91-97 (review article).