Microarrays for protease detection in tissues and cells.
Expression of a given protease and of the endogenous inhibitors that regulate protease activity can be readily determined at the transcript level by using whole genome microarray chips. In the case of proteases and protease inhibitors, however, determining which cells are expressing them is often critical to understanding the functional roles of the proteases. For example, in cancer many of the proteases are derived from cells that are found in the microenvironment surrounding the tumor, e.g., fibroblasts and inflammatory cells. Proteases from both fibroblasts and inflammatory cells have been implicated in malignant progression. Therefore, it is important to recognize the origin of these molecules if one is to develop effective therapies. In this regard, mouse transgenic models and xenograft models in which human tumor cells are implanted in mice are useful tools. To profile human and mouse proteases, protease inhibitors, and protease interactors, we have developed in partnership with Affymetrix a custom, single platform, dual species chip: the Hu/Mu ProtIn chip. The Hu/Mu ProtIn chip has been validated for its ability to identify human and mouse transcripts in single species specimens and to identify and distinguish between human and mouse transcripts in dual species specimens such as xenografts. In the latter specimens, the Hu/Mu ProtIn chip has enabled us to identify host (mouse) proteases that play a protective role in development of lung tumors. Here we outline a protocol for using the Hu/Mu ProtIn chip to profile proteases, protease inhibitors, and protease interactors in tissues and cells.
Moin
Kamiar
K
Department of Pharmacology, Wayne State University, 540 E. Canfield, Detroit, MI, USA.
Schwartz
Donald
D
Mullins
Stefanie R
SR
Sloane
Bonnie F
BF
D000818:Animals; D002478:Cells, Cultured; D020869:Gene Expression Profiling / Q000379:methods; D006801:Humans; D051379:Mice; D010447:Peptide Hydrolases / Q000032:analysis* / Q000235:genetics*; D011480:Protease Inhibitors / Q000032:analysis; D040081:Protein Array Analysis / Q000379:methods*; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D014183:Transplantation, Heterologous
D011480:Protease Inhibitors; D012313:RNA; D010447:Peptide Hydrolases