Task 2

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Examples of membrane organization diversity across evolution: A, Gloeobacter violaceus (Rippka et al., 1974); B, Synechocystis 6803 (Liberton et al., 2006) ; C, Chlamydomonas reinhardii (http://www.stolaf.edu/people/giannini/cell/nuc.htm); D, Yeast Saccharomyces cerevisiae. (E-G) Intracellular membrane proliferation upon over expression of membrane proteins in Escherichia coli: E, chemotaxis receptor Tsr (Lefman et al., 2004); F, F1Fo ATP synthase complex (Von Meyenburg et al., 1984), G, H, AtpF of F1Fo ATPase (Arechaga et al., 2000).

 

In living cells, bioenergetic processes take place in membranes. In eukaryotic cells, respiratory complexes are found in mitochondria cristae and chloroplasts thylakoid. Little is known about membrane biogenesis, maintenance and remodelling. Labex Dynamo is focussing on three model systems : the dynamic of mitochondrial fusion and fission in the yeast Saccharomyces cerevisiae, the biogenesis of thylakoid membranes of Chlamydomonas Reinhardii, and internal membrane proliferation in Escherichia coli.  A related topic supported by DYNAMO is the structural organisation of membrane protein complexes.

 

 

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Haemophilus influenzae is a Gram-negative bacterium whose unique host is human. Being heme auxotroph, it displays at the cell surface several systems that allow it to acquire heme from human hemoproteins. Among these systems we are studying the Hxu (Hemopexin utilisation) system. The Hxu system allows heme acquisition from Hemopexin, a serum glycoprotein which binds heme with one of the highest known affinities and is involved in heme recycling after hemolysis, protecting in this way the body from the oxidative damage that an be caused by free heme.

The Hxu system is made up of three outer membrane proteins: the TonB dependent heme receptor Hxu C, and the two partner secretion system (TPS) proteins HxuB and A. The aim of my PhD work is to structurally characterize HxuA alone and in complex with Hemopexin, and to try to understand in detail the mechanism of heme uptake. For this purpose we are using a combined approach that involves molecular biology, biochemistry, electron microscopy and protein crystallography.

 

 

Tethered mitochondria: Mitochondrial enriched fractions purified from wild type cells were centrifuged to be brought in close proximity and subjected to in vitro fusion. Resulted reactions were analyzed by Transmission Electron Microscopy at the Max Planck Institute of Biophysics (Frankfurt, Germany). On this image, note the region of contact between two adjacent mitochondria about to fuse their outer membranes.

Mitochondria constitute a real and remarkably dynamic network whose morphology is conditioned by a constant equilibrium between frequent fission and fusion events of their membranes. These processes are essential to shape the ultra-structure of the mitochondrial compartment and are thus also crucial for all mitochondrial functions. Consequently, defects in mitochondrial fusion and fission are associated with numerous pathologies and severe neurodegenerative syndromes especially. Unique among intracellular membrane fusion systems, that of mitochondria does not require SNAREs but Dynamin-Related Proteins (DRPs). DRPs are large GTPases with a particular ability to bind and shape the form of biological membranes and especially promote their fission. How DRPs can also catalyze mixing of lipid bilayers remains however poorly understood. Allying cutting edge cell biology and biochemistry approaches, task 2 of DYNAMO addresses this outstanding question by focusing on the yeast DRPs that promote tethering of mitochondria and fusion of their outer membranes.

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