Resumen de Coordinación entre proliferación y polaridad celular. Funciones citoplasmáticas de ciclina d1
Rita Maria Fernandez Hernandez
Coordination between proliferation and processes that involve the establishment of cell polarity and adhesion and migration are critical for proper differentiation and tissue development. Abnormalities in the linkage of these processes can lead to the development of oncogenesis. Cell proliferation is regulated by the activity of the complexes formed by cyclins and cyclin-dependent-kinases. D cyclins are molecules that play a key role in regulating cell proliferation. Mammalian cells contain genes that encode three highly homologous type D cyclin (D1, D2, D3), which are expressed in a tissue-specific and are associated with Cdk4 or Cdk6 to form an active kinase complex that regulates the transcription of various clusters of genes. Among the three cyclins, cyclin D1 is the most studied, since the CycD1 gene is amplified in many human cancers. The aim of this study was the characterization of new functions CycD1-CDK4 complex, especially concerning the control of cell polarity. This objective is based on three initial assumptions: 1) the elimination of CycD1 in different cell lines and primary cultures causes abnormalities in the establishment of polarity, in particular in processes like the formation of neurites, adhesion to the extracellular matrix and migration, 2) it has been described the cytoplasmic accumulation of CycD1 in different conditions, eg during neuronal differentiation, and 3) in yeast cells is well established that the homologues of Cdk4-cyclin D1 coordinate the proliferation and polarized growth (budding) by modulating the activity of several regulatory proteins of GTPases. This background led us to hypothesize the existence of a cytoplasmic fraction of Cyc D1 that could participate in the coordination between proliferation and cell polarity, ie establishing decisions between processes as proliferate or migrate, proliferate or differentiate ....
In this study we have shown that in different growth conditions there is a fraction of cyclin D1 localized in the cytoplasm where it performs a new function. We have reported that cyclin D1 is a novel regulator of the Ral pathway. First we characterized the binding of CycD1-Cdk4 complex to Ral GTPases and CycD1 colocalization with Ral A and B in the cytoplasm, particularly in the trans-Golgi. This coincides with the fact that Ral GTPases control cell polarity processes, adhesion and migration, by regulating the trafficking of vesicles. For example, Ral activity is required for transport and endocytosis of integrins affect the dynamics of cell adhesion to the matrix. Here too, we describe that cyclin D1 interacts and colocalized with Ral active sites and ß1-integrin paxilina, proteins that regulate cell adhesion and motility. In addition, the activity of cyclin D1-Cdk4 complex promotes accumulation of active forms of Ral. Based on these data we propose that CycD1 regulates the dynamics of substrate adhesion and cell motility through the modulation of Ral activity. According to this possibility, we have shown that hyperactivation of the Ral pathway subverts the increased adhesion to the substrate and loss of motility caused by the elimination of CycD1. Regardless, in this work we have also identified by two-hybrid a new interactor of cyclin D1, the protein PALS1. This scaffold is involved in the regulation of cell-cell adhesion and cell migration. Finally, all data provided in this thesis has allowed us to describe a new role for cyclin D1 that provides a link between proliferation, polarity and differentiation and may help explain the contribution of cyclin D1 in tumor formation and development of metastasis.
