Towards an understanding of the molecular events associated to the role of SPARC in tumor progression

ANDREA LLERA; MARIA SOLEDAD SOSA; SILVIA JUAREZ; JUAN PABLO ALBAR; MARIANO ALVAREZ; OSVALDO PODHAJCER

Beijing, China

Congreso; Human Proteome Organization 3rd. Annual World Congress; 2004

SPARC is a secreted glycoprotein that elicits changes in cell shape and proliferation. While its normal expression is restricted to tissues under remodeling, SPARC is overexpressed in different tumors, in association with tumor progression and metastasis. Our previous results showed that stable transfection of tumor cells with antisense SPARC DNA abolished tumorigenicity in an in vivo melanoma murine model. Along with traditional approaches to define a molecular pathway for SPARC-mediated tumor progression, we have started a proteomic analysis of proteins expressed by human melanoma cells with modulated SPARC expression, in order to identify putative SPARC-interacting proteins. We studied conditioned medium from MEL-LES clone 1-D (L-1D, showing an 80% decrease of SPARC mRNA and protein expression), as compared with MEL-LES cells transfected with control plasmid (L-CMV). Proteomic analysis was done by two-dimensional electrophoresis followed by tryptic digestion of spots and MALDI-TOF peptide fingerprinting analysis. We have identified a total of 81 unique proteins, including some related to SPARC function, extracellular matrix structural proteins and oncoproteins. We have also detected 24 differentially expressed proteins (Student T-test p<0.05) in the non-tumorigenic clone L-1D versus tumorigenic L-CMV cells, including known SPARC interactors such as type I collagen. We are currently validating those most biologically significant candidates using Northern blotting, real time PCR and/or immunoblotting.