Cyclin D1 interacts and collaborates with Ral GTPases enhancing cell detachment and motility

Abstract

Alterations in the levels of adhesion and motility of cells are critical events in the development of metastasis. Cyclin D1 (CycD1) is one of the most frequently amplified oncogenes in many types of cancers and it is also associated with the development of metastasis. Despite this, we still do not know which are all the relevant pathways by which CycD1 induces oncogenic processes. CycD1 functions can be either dependent or independent of the cyclin-dependent kinase Cdk4, and they affect several cellular aspects such as proliferation, cell attachment and migration. In this work, we reveal a novel function of CycD1 that fosters our understanding of the oncogenic potential of CycD1. We show that CycD1 binds to the small GTPases Ral A and B, which are involved, through exocyst regulation, in the progression of metastatic cancers, inducing anchorage-independent growth and cell survival of transformed cells. We show that CycD1 binds active Ral complexes and the exocyst protein Sec6, and co-localizes with Ral GTPases in trans-Golgi and exocyst-rich regions. We have also observed that CycD1–Cdk4 phosphorylates the Ral GEF Rgl2 ‘in vitro’ and that CycD1–Cdk4 activity stimulates accumulation of the Ral GTP active forms. In accordance with this, our data suggest that CycD1–Cdk4 enhances cell detachment and motility in collaboration with Ral GTPases. This new function may help explain the contribution of CycD1 to tumor spreading.

We thank N Agell, C Yeaman, JL Bos, M Matsuda, X Bustelo, P Crespo, MA del Pozo and J Herreros for the gift of plasmids and reagents, and also N Eritja,MBozic, JM Valdivielso, Lluı´s Fajas, Eloi Montanez and SaraWickstro¨m for technical advice. We are grateful to J Odajima and P Sicinski for providing CCND1 / and CCND1þ/þ immortalized MEFs, andMHendrix and E Seftor for providing R3327-50A cells. We thank S Rius, I Navarro, I Montoliu and MA Cornado´ for their technical assistance and the members of CYC lab for helpful discussions. This work was funded by the Ministry of Education and Science of Spain BFU2007-65640/BMC (Eloi Garı´) and BFU2007- 67929-C02-01 (Martı´ Aldea) and Consolider-Ingenio 2010 (CSD2007-00015). RMH Ferna´ ndez and M Ruiz-Miró were supported by predoctoral fellowships from the Ministry of Education and Science of Spain.