Design of a retroviral-mediated ecdysone-inducible system and its application to the expression profiling of the PTEN tumor suppressor.
We have engineered the ecdysone-inducible mammalian expression system for general retroviral delivery to cultured mammalian cells. We inducibly expressed PTEN in the glioblastoma cell line, U87MG, lacking this gene. Because nearly all cells are recruited on induction, we find both up- and down-regulated genes by cDNA microarray analysis. The changes we see are similar to those observed after treatment with LY294002, an inhibitor of phosphatidylinositol 3-OH kinase, fully consistent with the model that PTEN antagonizes phosphatidylinositol 3-OH kinase. Both treatments result in suppressed expression of the transforming growth factor (TGF)-beta gene and the genes of the cholesterol biosynthesis pathway. Our results illustrate the power of using a fully inducible expression system in conjunction with cDNA microarray analysis for exploring gene function.
Stolarov
J
J
Cold Spring Harbor Laboratory, 1 Bungtown Road, P.O. Box 100, Cold Spring Harbor, NY 11724, USA.
Chang
K
K
Reiner
A
A
Rodgers
L
L
Hannon
G J
GJ
Wigler
M H
MH
Mittal
V
V
D004440:Ecdysone / Q000096:biosynthesis*; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D016147:Genes, Tumor Suppressor; D005822:Genetic Vectors; D006801:Humans; D051059:PTEN Phosphohydrolase; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics*; D012190:Retroviridae / Q000235:genetics*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D025521:Tumor Suppressor Proteins / Q000235:genetics*
D025521:Tumor Suppressor Proteins; D004440:Ecdysone; D010744:Phosphoric Monoester Hydrolases; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human