you are here » research » programs

Cell matrix signaling

Nicolai Sidenius

[IFOM]

Nicolai Sidenius, Ph.D.
c/o IFOM-IEO Campus
Via Adamello, 16 - 20139 Milan, Italy
T +39 02 574303.261 (Office) - +39 02 574303.328 (Lab)
F +39 02 574303.231
nicolai.sideniusmailifom.eu

Research project

Background:

The extracellular matrix (ECM) is the complex structural scaffold surrounding and supporting all cells in all tissue and organs. The appropriate dynamic regulation of cell–ECM interactions is fundamental for the successful outcome of physiological processes including embryonic development, morphogenesis and tissue homoeostasis, as well as a devastating factor in the invasion and dissemination of cancer cells in the process of tumor metastasis – the number one cause of mortality in this disease.

The urokinase-type plasminogen activator (uPA)-receptor (uPAR) is a GPI-anchored plasma membrane receptor highly expressed in inflammatory processes and virtually in all human cancers types. Since the discovery of uPAR about 20 years ago extensive evidence obtained by in vitro experimentation, using in vivo animal models, as well as from correlative clinical studies, suggests an important functional link between the over-expression of components of the uPA/uPAR-system and the malignant phenotype of cancer cells including their increased proliferation, migration, and invasion of the surrounding tissue. In the context of cell-ECM interactions uPAR appear to play a dual role. On one side uPAR promotes the degradation and turnover of the extracellular matrix by promoting extracellular protolysis (through the regulation of the plasminogen activation protease cascade) as well as by the internalization of inactive uPA/uPA-inhibitor complexes. On the other side uPAR has been shown to directly modulate the strength and location of cell-ECM interactions through its direct contact with the extracellular matrix protein vitronectin (Vn) and through the regulation of the activity of members of the integrin family of adhesion receptors. The latter processes typically occur independently of the catalytic activity of uPA and are often referred to as the non-proteolytic functions of uPAR. These non-proteolytic function of uPAR is historically the key research area of our lab.

Current lines of research in the lab:

  1. Indentification of the molecular mechanisms of signal transduction induced by cell adhesion to the extracellular matrix.
    Our long-standing interest in the molecular basis for uPAR’s ability to induce signal transduction has over the last few years led to a focused effort aimed at understanding, at the molecular level, how membrane receptors translate extracellular signals (i.e. ligand binding) into defined intracellular biochemical signaling events. Of specific interest is the mechanism by which membrane receptors lacking transmembrane and cytoplasmic domains signal. This type of receptors, including uPAR, cannot transduce signals by propagation of a conformational change over the plasma membrane and must therefore rely on different mechanisms. The prevailing paradigm is that these adhesion receptors utilize direct lateral interactions with other signaling competent receptors (such as integrins or receptor tyrosine kinases) to transmit the signal. We have, however, recently documented that direct lateral interaction with other membrane receptors play no functional role in the ability of uPAR to signal and that the signaling mechanism downstream of uPAR is likely to be shared with most, if not all, membrane receptors with high-affinity ligands in the extracellular matrix. Our findings strongly question the importance of the prevailing paradigm of “adapter mediated” uPAR signaling, but the real mechanism still has to be established. In the lab we have some promising “leads” on this mechanism and these are currently being exploited using state-of-the-art “genetics-in-cell-culture” and optical imaging techniques.

  2. The importance of the uPAR/Vn-interaction in vivo.
    Our demonstration of the pivotal importance of the direct molecular interaction with Vn for the function of uPAR in vitro clearly identifies this interaction as a promising target for the pharmaceutical interference with cancer invasion and metastasis (and in other diseases where uPA/uPAR is known or suspected to be of functional relevance). To address the importance of this interaction in vivo we are currently conducting a systematic comparison of the phenotypes of uPAR and Vn null mice in a variety of processes where these genes have been published to be of functional relevance including inflammation, tumor invasion and metastasis, as well as in HSC/HPC mobilization (in collaboration with Dr. Marc Tjwa, Leuven, Belgium). If functional and relevant phenocopying will be observed between these two lines of null mice the final evidence for importance of the molecular interaction between the two proteins will be provided by the generation of a uPAR knock-in mouse in which we have specifically and selectively disrupted the Vn-binding activity of the receptor.

  3. Determination of the structural basis for the regulation of the molecular interaction between uPAR and Vn.
    Several crystal structures of the bi-molecular complex between uPAR and uPA, as well as a structure of the ternary complex between uPAR, uPA and Vn have been published, providing a certain degree of insight into the structural basis for the molecular interactions between these three proteins. Despite this structural insight the published structures, besides roughly defining the molecular interfaces, fall short of explaining the biological activity of uPAR as a cell adhesion receptor for vitronectin and the importance of uPA in this process. Firstly, efficient uPAR binding to Vn requires dimerization while uPAR is monomeric in the crystal structure(s). Secondly, the structures only partially explain the structural requirements to uPAR and Vn as determined by complete alanine scans of binding regions of these proteins. Thirdly, the stoichiometry of the component observed in the crystal structures, as well as the experimentally determined affinities, are entirely insufficient to explain the uPA-dependence of soluble uPAR binding and cell adhesion to matrix Vn. To fill this knowledge gab a series of structural and biochemical approaches are being exploited in the lab to determine the stoichiometry and topology of the high-affinity uPAR/uPA/Vn-interaction as well as to understand its regulation on the surface of living cells by single molecule imaging/nano-spectroscopy techniques conducted in the collaboration with the Caiolfa group at the San Raffaele Scientific Institute in Milan, Italy.
update: Oct 2008
more info about this group:
 
 
 
last update: February 14, 2012 . Copyright © IFOM & IEO . Campus IFOM-IEO . Via Adamello 16 . 20139 Milan Italy
webmasteratifom-ieo-campus.it - optimized: 1024x768, supported browsers: IE6+ . Safari 4+ . Firefox 2+ . Opera 8+ . Netscape 7+