RESUMO
A chemical genetics approach identified a cellular target of several proapoptotic farnesyl transferase inhibitors (FTIs). Treatment with these FTIs caused p53-independent apoptosis in Caenorhabditis elegans, which was mimicked by knockdown of endosomal trafficking proteins, including Rab5, Rab7, the HOPS complex, and notably the enzyme Rab geranylgeranyl transferase (RabGGT). These FTIs were found to inhibit mammalian RabGGT with potencies that correlated with their proapoptotic activity. Knockdown of RabGGT induced apoptosis in mammalian cancer cell lines, and both RabGGT subunits were overexpressed in several tumor tissues. These findings validate RabGGT, and by extension endosomal function, as a therapeutically relevant target for modulation of apoptosis, and enhance our understanding of the mechanism of action of FTIs.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Alquil e Aril Transferases/metabolismo , Alquil e Aril Transferases/fisiologia , Animais , Antineoplásicos/farmacologia , Apoptose/genética , Apoptose/fisiologia , Caenorhabditis elegans/efeitos dos fármacos , Caenorhabditis elegans/enzimologia , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/fisiologia , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/fisiologia , Caspases/genética , Caspases/metabolismo , Caspases/fisiologia , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Expressão Gênica/genética , Células Germinativas/efeitos dos fármacos , Humanos , Mutagênese/genética , Neoplasias/enzimologia , Neoplasias/genética , Prenilação de Proteína/efeitos dos fármacos , Interferência de RNA , RNA de Cadeia Dupla/genética , RNA Interferente Pequeno/genética , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/fisiologia , Proteínas rab de Ligação ao GTP/genéticaRESUMO
We report here on a chemical genetic screen designed to address the mechanism of action of a small molecule. Small molecules that were active in models of urinary incontinence were tested on the nematode Caenorhabditis elegans, and the resulting phenotypes were used as readouts in a genetic screen to identify possible molecular targets. The mutations giving resistance to compound were found to affect members of the RGS protein/G-protein complex. Studies in mammalian systems confirmed that the small molecules inhibit muscarinic G-protein coupled receptor (GPCR) signaling involving G-alphaq (G-protein alpha subunit). Our studies suggest that the small molecules act at the level of the RGS/G-alphaq signaling complex, and define new mutations in both RGS and G-alphaq, including a unique hypo-adapation allele of G-alphaq. These findings suggest that therapeutics targeted to downstream components of GPCR signaling may be effective for treatment of diseases involving inappropriate receptor activation.