Tumor suppressor Smad4 mediates downregulation of the anti-adhesive invasion-promoting matricellular protein SPARC: Landscaping activity of Smad4 as revealed by a "secretome" analysis.
We have demonstrated previously that restoration of Smad4 expression in Smad4-deficient SW480 human colon carcinoma cells was adequate to suppress tumorigenicity and invasive potential, whereas cell growth in vitro was not affected. Here we show that Smad4-positive and Smad4-negative SW480 cells deposit extracellular matrices in tissue culture which are functionally different with respect to their adhesiveness. We present a "differential secretomics analysis" as the most direct approach to identify the underlying alterations. The protein composition of conditioned media produced by Smad4-positive and Smad4-negative SW480 cells was compared by two-dimensional (2-D) gel electrophoresis. A major group of protein spots was detected in media derived from Smad4-negative cells, only, which were identified as "secreted protein, acidic and rich in cysteins" (SPARC) by mass spectrometry. SPARC expression in SW480 cells was suppressed by Smad4 at the level of transcription. SPARC is a glycoprotein of the extracellular matrix, characterized as an anti-adhesive and invasion-promoting protein. Differential secretomics appeared as a powerful method to identify a novel Smad4 target gene, which may be one of the players involved in reduced adhesiveness of extracellular matrices and thus consistent with Smad4's emerging role as an invasion suppressor.
Volmer
Martin W
MW
Department of Internal Medicine, IMBL, Ruhr-Universität Bochum, Germany.
Radacz
Yvonne
Y
Hahn
Stephan A
SA
Klein-Scory
Susanne
S
Stühler
Kai
K
Zapatka
Marc
M
Schmiegel
Wolff
W
Meyer
Helmut E
HE
Schwarte-Waldhoff
Irmgard
I
D000595:Amino Acid Sequence; D002448:Cell Adhesion; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms; D017077:Culture Media, Conditioned / Q000032:analysis; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D005109:Extracellular Matrix / Q000737:chemistry*; D015972:Gene Expression Regulation, Neoplastic; D016147:Genes, Tumor Suppressor; D006801:Humans; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009361:Neoplasm Invasiveness; D009363:Neoplasm Proteins / Q000032:analysis* / Q000378:metabolism; D015676:Osteonectin / Q000737:chemistry / Q000378:metabolism*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D016622:Silver Staining; D051901:Smad4 Protein; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D015534:Trans-Activators / Q000378:metabolism*; D014158:Transcription, Genetic; D025521:Tumor Suppressor Proteins / Q000032:analysis* / Q000378:metabolism
D017077:Culture Media, Conditioned; D004268:DNA-Binding Proteins; D009363:Neoplasm Proteins; D015676:Osteonectin; C497796:SMAD4 protein, human; D051901:Smad4 Protein; D015534:Trans-Activators; D025521:Tumor Suppressor Proteins