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Novel invadopodia components revealed by differential proteomic analysis
Francesca Attanasio, Giusi Caldieri, Giada Giacchettia, Remcovan Horssen, Bé Wieringa, Roberto Buccione, 2010. European Journal of Cell Biology Vol. 90, Issues 2-3, February-March 2011, Pages 115-127.

Tumour growth and dissemination throughout the body to form metastases, the main cause of cancer-related mortality, depend on the ability of tumour cells to move through/invade the very dense meshwork formed by the extracellular matrix (ECM), in which the cells that make up tissues and organs are embedded. The controlled degradation of specific components of the ECM is an essential step in tumour cell invasion. This process can be recapitulated in the “test tube” using tumour cell lines that form protrusive structures called invadopodia that have the ability to locally degrade and penetrate the ECM.

Our understanding of the molecular composition of invadopodia has rapidly advanced in the last few years, but is far from complete. In this paper, we describe a novel approach to accelerate the discovery process. In detail, we were able to prepare tumour cell fragments enriched in invadopodia which were then analyzed by proteomic techniques. This led to the discovery of new protein components belonging to different functional groups including those related to the production of energy, cell movement and secretion of proteins.

We expect these findings to open further avenues for the molecular study of invasive growth behaviour of cancer cells.

Figure Legend:

The G protein Beta subunit is a very important protein that mediates communication between the extracellular and intracellular environments. We first discovered it to be associated with invadopodia in our proteomics analysis and then validated the finding by fluorescence microscopy as shown here. Invadopodia (labeled in blue) are positioned over areas of digested extracellular matrix (dark holes in the green background) and contain the G protein Beta subunit (red).

Podosome rings generate forces that drive saltatory osteoclast migration.

Shiqiong Hu , Emmanuelle Planus , Dan Georgess, Christophe Place , Xianghui Wang , Corinne Albiges-Rizo , Pierre Jurdic, and Jean-Christophe Géminard. Mol Biol Cell. 2011 Jul 7. [Epub ahead of print]

Osteoclasts are huge cells containing several nuclei formed by fusion of mononucleated cells belonging to the monocyte/macrophage lineage. They are found on the bone surface, where they remove the old bone matrix in order to allow bone rejuvenation by osteoblasts. In order to adhere to their substrates, to migrate and to resorb, they are equipped of structures called podosomes, which are loosely organized on artificial substrates (plastic or glass) but condensed on calcium apatite mineral. Podosomes are made of polymerized actin organized as dense dots, and are surrounded by a loose network of polymerized actin. They are very dynamic structures with a short life-time (2 to 3 minutes), that self organize with different patterns during osteoclast life.

Podosomes are thought to be responsible for cell adhesion and migration. In this article we show that osteoclast podosomes are major anchoring sites exerting tensions on the substrate. They appear in areas where membranes are expanding and disappear in areas where membranes retract. Videomicroscopy has shown that podosome dynamics can move latex beads dispersed in a soft substrate, thus demonstrating podosome-mediated tension forces. Finally, analyzing the migration dynamic of several osteoclasts, we were able to show that actin-containing podosomes are major driving forces for osteoclast migration. A mathematical model of osteoclast migration provides unexpected results: osteoclasts are moving by jumps and this is reminiscent of the way that osteoclasts are moving to resorb bone surfaces.

Figure legend:

Dynamic images of an osteoclast spreading on the plastic surface of a petri dish. Podosomes are in green; they are present at the periphery of the cell in the spreading area.

t= 0  rounded non-adherent osteoclasts are seeded in the petri dish; Then images of the same cells 12minutes (t= 12 mn) and 20 minutes (t=20mn) after seeding.

 

The kinesin KIF9 and reggie/flotillin proteins regulate matrix degradation by macrophage podosomes.

Susanne Cornfine, Mirko Himmel, Petra Kopp, Karim el Azzouzi, Christiane Wiesner, Marcus Krüger, Thomas Rudel, Stefan Linder, 2010. Molecular Biology of the Cell Vol. 22, Issue 2, January 15, 2011, Pages 202-215.

Macrophages are immune cells that form the first line of defense against infectious agents such as bacteria or viruses. To get to sites of infection, macrophages have to migrate through the dense meshwork of the extracellular matrix that connects cells within tissues. Podosomes, adhesion structures with the ability to locally degrade matrix material, play a key role in this migration.

We show here that matrix degradation by macrophage podosomes depends on contact with the microtubule network of cells. Microtubules form a “cellular railway system”, which enables the delivery of cargo molecules through transport by tiny motors walking on these fibers. This study identifies the motor protein KIF9 as an essential regulator of matrix degradation at podosomes. KIF9 delivers cargo vesicles, which contain proteins of the reggie family, a group of membrane-associated proteins. We could also identify a distinct region of KIF9 as the binding site for reggie-1. Interaction of both proteins through this region is critical for regulating matrix degradation at podosomes.

Figure legend:

Vesicles containing the motor protein KIF9 (green) are associated with microtubules (red), which form the "cellular railway system".

 

 

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