GeneBlocs are powerful tools to study and delineate signal transduction processes that regulate cell growth and transformation.
The study of signal transduction processes using antisense oligonucleotides is often complicated by low intracellular stability of the antisense reagents or by nonspecific effects that cause toxicity. Here, we introduce a new class of antisense molecules, so-called GeneBlocs, which are characterized by improved stability, high target RNA specificity, and low toxicity. GeneBlocs allow for efficient downregulation of mRNA expression at nanomolar concentrations, and they do not interfere with cell proliferation. We demonstrate these beneficial properties using a positive readout system. GeneBloc-mediated inhibition of tumor suppressor PTEN (phosphatase and tension homologue detected on chromosome 10) expression leads to hyperactivation of the phosphatidylinositol (PI) 3-kinase pathway, thereby mimicking the loss of PTEN function and its early consequences observed in mammalian cancer cells. Specifically, cells treated with PTEN GeneBlocs show functional activation of Akt, a downstream effector of PI 3-kinase signaling, and exhibit enhanced proliferation when seeded on a basement membrane matrix. In addition, GeneBlocs targeting the catalytic subunit of PI 3-kinase, p110, specifically inhibit signal transduction of endogenous or recombinant PI 3-kinase. This demonstrates that GeneBlocs are powerful tools to analyze and to modulate signal transduction processes and, therefore, represent alternative reagents for the validation of gene function.
Sternberger
Maria
M
atugen AG, Berlin, Germany.
Schmiedeknecht
Anett
A
Kretschmer
Anny
A
Gebhardt
Frank
F
Leenders
Frauke
F
Czauderna
Frank
F
Von Carlowitz
Ira
I
Engle
Mike
M
Giese
Klaus
K
Beigelman
Leonid
L
Klippel
Anke
A
D000818:Animals; D017209:Apoptosis / Q000528:radiation effects; D001483:Base Sequence; D002455:Cell Division / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D002471:Cell Transformation, Neoplastic / Q000235:genetics*; D004789:Enzyme Activation; D015870:Gene Expression; D005821:Genetic Techniques; D006801:Humans; D016376:Oligonucleotides, Antisense / Q000737:chemistry* / Q000494:pharmacology; D051059:PTEN Phosphohydrolase; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000037:antagonists & inhibitors* / Q000235:genetics; D010766:Phosphorylation; D051381:Rats; D016196:S Phase / Q000187:drug effects; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology*; D025521:Tumor Suppressor Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics; D014466:Ultraviolet Rays
D016376:Oligonucleotides, Antisense; D025521:Tumor Suppressor Proteins; D019869:Phosphatidylinositol 3-Kinases; D010744:Phosphoric Monoester Hydrolases; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human