Profiling the activity of G proteins in patient-derived tissues by rapid affinity-capture of signal transduction proteins (GRASP).
The next phase in molecular medicine will require the ability to identify signal transduction events inside a cell, in the biologic context of the disease-host interface and at a given point in time. New technologies are needed to profile the activity of these signaling pathways in patient tissue rather than cultured cell lines since the tumor-host microenvironment influences the cellular proteome. We introduce such a technology, rapid affinity capture of signaling proteins (GRASP), to investigate the activity of signaling pathways from patient-derived carcinomas and benign epithelial surfaces and apply it to studying important signaling events in ovarian carcinoma. During the progression from benign ovarian epithelium to invasive carcinoma, there is loss of repression of Rho A as evidenced by its dissociation from its inhibitor, Rho Guanine Nucleotide Dissociation Inhibitor (RhoGDI). GRASP is more informative than simply profiling transcript or protein levels. Furthermore, GRASP coupled with mass spectrometry allowed us to identify a protein-binding partner of RhoGDI, demonstrating the power of this technology in the discovery of potentially novel protein-protein interactions. GRASP represents an advance in the field of proteomics as it detects protein interactions present in cells as they exist in their native tissue microenvironment.
Laboratory of Pathology, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA. bermand@mail.nih.gov
David M
DM
Berman
Ie-Ming
IeM
Shih
Lorri-Anne
LA
Burke
Timothy D
TD
Veenstra
Yingming
Y
Zhao
Thomas P
TP
Conrads
Sung Won
SW
Kwon
Van
V
Hoang
Li-Rong
LR
Yu
Ming
M
Zhou
Robert J
RJ
Kurman
Emanuel F
EF
Petricoin
Lance A
LA
Liotta
D002277:Carcinoma / Q000378:metabolism; D004848:Epithelium / Q000378:metabolism; D005260:Female; D019204:GTP-Binding Proteins / Q000737:chemistry*; D006801:Humans; D008163:Luminescent Measurements; D008274:Magnesium / Q000494:pharmacology; D008280:Magnetics; D013058:Mass Spectrometry / Q000379:methods; D010051:Ovarian Neoplasms / Q000378:metabolism; D010053:Ovary / Q000378:metabolism; D020543:Proteome; D040901:Proteomics / Q000379:methods*; D015398:Signal Transduction; D020741:rho GTP-Binding Proteins / Q000378:metabolism
D020543:Proteome; D019204:GTP-Binding Proteins; D020741:rho GTP-Binding Proteins; D008274:Magnesium