RESUMEN
RhoBTB1 is an atypical Rho GTPase with two BTB domains in addition to its Rho domain. Although most Rho GTPases regulate actin cytoskeletal dynamics, RhoBTB1 is not known to affect cell shape or motility. We report that RhoBTB1 depletion increases prostate cancer cell invasion and induces elongation in Matrigel, a phenotype similar to that induced by depletion of ROCK1 and ROCK2. We demonstrate that RhoBTB1 associates with ROCK1 and ROCK2 and its association with ROCK1 is via its Rho domain. The Rho domain binds to the coiled-coil region of ROCK1 close to its kinase domain. We identify two amino acids within the Rho domain that alter RhoBTB1 association with ROCK1. RhoBTB1 is a substrate for ROCK1, and mutation of putative phosphorylation sites reduces its association with Cullin3, a scaffold for ubiquitin ligases. We propose that RhoBTB1 suppresses cancer cell invasion through interacting with ROCKs, which in turn regulate its association with Cullin3. Via Cullin3, RhoBTB1 has the potential to affect protein degradation.
Asunto(s)
Proteínas de Neoplasias/metabolismo , Neoplasias de la Próstata/metabolismo , Proteínas de Unión al GTP rho/metabolismo , Quinasas Asociadas a rho/metabolismo , Animales , Células COS , Chlorocebus aethiops , Proteínas Cullin/genética , Proteínas Cullin/metabolismo , Células HeLa , Humanos , Masculino , Invasividad Neoplásica , Proteínas de Neoplasias/genética , Células PC-3 , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/patología , Proteínas de Unión al GTP rho/genética , Quinasas Asociadas a rho/genéticaRESUMEN
Cancer cells interact with endothelial cells during the process of metastatic spreading. Here, we use a small interfering RNA screen targeting Rho GTPases in cancer cells to identify Cdc42 as a critical regulator of cancer cell-endothelial cell interactions and transendothelial migration. We find that Cdc42 regulates ß1 integrin expression at the transcriptional level via the transcription factor serum response factor (SRF). ß1 integrin is the main target for Cdc42-mediating interaction of cancer cells with endothelial cells and the underlying extracellular matrix, as exogenous ß1 integrin expression was sufficient to rescue the Cdc42-silencing phenotype. We show that Cdc42 was required in vivo for cancer cell spreading and protrusion extension along blood vessels and retention in the lungs. Interestingly, transient Cdc42 depletion was sufficient to decrease experimental lung metastases, which suggests that its role in endothelial attachment is important for metastasis. By identifying ß1 integrin as a transcriptional target of Cdc42, our results provide new insight into Cdc42 function.
Asunto(s)
Integrina beta1/metabolismo , Migración Transendotelial y Transepitelial , Proteína de Unión al GTP cdc42/metabolismo , Animales , Adhesión Celular , Línea Celular Tumoral , Células Cultivadas , Células Endoteliales/citología , Células Endoteliales/metabolismo , Fibronectinas/metabolismo , Regulación Neoplásica de la Expresión Génica , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Ratones , Ratones SCID , Metástasis de la Neoplasia , Trasplante de Neoplasias , Transporte de Proteínas , Factor de Respuesta Sérica/metabolismo , Transcripción Genética , Migración Transendotelial y Transepitelial/genética , Proteína de Unión al GTP cdc42/deficiencia , Proteína de Unión al GTP rac1/metabolismo , Proteína de Unión al GTP rhoA/metabolismoRESUMEN
BACKGROUND: There is no evidence to date on whether transcriptional regulators are able to shift the balance between mitochondrial fusion and fission events through selective control of gene expression. METHODOLOGY/PRINCIPAL FINDINGS: Here, we demonstrate that reduced mitochondrial size observed in knock-out mice for the transcriptional regulator PGC-1beta is associated with a selective reduction in Mitofusin 2 (Mfn2) expression, a mitochondrial fusion protein. This decrease in Mfn2 is specific since expression of the remaining components of mitochondrial fusion and fission machinery were not affected. Furthermore, PGC-1beta increases mitochondrial fusion and elongates mitochondrial tubules. This PGC-1beta-induced elongation specifically requires Mfn2 as this process is absent in Mfn2-ablated cells. Finally, we show that PGC-1beta increases Mfn2 promoter activity and transcription by coactivating the nuclear receptor Estrogen Related Receptor alpha (ERRalpha). CONCLUSIONS/SIGNIFICANCE: Taken together, our data reveal a novel mechanism by which mammalian cells control mitochondrial fusion. In addition, we describe a novel role of PGC-1beta in mitochondrial physiology, namely the control of mitochondrial fusion mainly through Mfn2.