CABIMER

 

 

 

 

 

 

 

 

 

 

 

In the last years our group has been interested in the study of molecular mechanisms that regulate the association between the Golgi apparatus and the centrosome in mammalian cells. The association Golgi/Centrosome plays an essential role in diverse cellular processes of great physiological importance such as cell polarity, cell division, cell migration and cell differentiation. In this respect, the most widely accepted view is that during interphase the centrosome is the only microtubule organising centre in nonpolarized proliferating and migrating cells. In this type of cells, microtubules appear organized in a characteristic radial pattern with the minus ends anchored to centrosome in the cell centre and the plus ends growing towards the cell periphery. The Golgi Apparatus, whose integrity depends on the microtubule network, is organized arou nd the centrosome and colocalizes with the minus ends of the microtubules. Several relevant contributions in the last years have modified this vision.

Firstly, the Golgi Apparatus possesses its own molecular machinery capable of nucleating and regulating a subpopulation of microtubules, probably involved in its own dynamics but also in general functioning of the cells.  Such machinery includes i) the protein AKAP450, that we have identified as responsible for the nucleation of microtubules that are produced at the cis face of the AG (Rivero et al., 2009);  ii) the CLASPs proteins that stabilize the plus ends  of the microtubules thus permitting lengthening; iii) the protein CAP350, a centrosomic protein which participates in the selective stabilization of the microtubules associated to the Golgi Apparatus y iv) the protein GMAP210, which participates in the microtubule-dependent fusion of Golgi elements to form a single organelle and assure its pericentrosomal position (Cárdenas et al., 2009).

Secondly, the Golgi morphology and the position of Golgi Apparatus/centrosome region depend on other factors, such as the cell adhesion. Thus, when the cells grow in infinite substrate, the Golgi Apparatus extends to considerable distances of the centrosome whereas in restricted adhesion conditions, i.e., when cells grow on micropatterns of fibronectin/collagen, an experimental condition that imitates a tissue, the Golgi Appartus appear as a ring whose centre is the centrosome. This suggests that there exists a crosstalk between the centre and the periphery of the cell that regulate the structure and probably the function of the Golgi Apparatus.

Thirdly, the Golgi Apparatus/Centrosome nexus is broken and re-established in each cell cycle by mechanisms that remained mostly unknown. Golgi ribbon are fragmented in mitotic cells and this fragmentation is a cue for regulating entry into mitosis. Our aim is to gain insights into the molecular machinery that controls the relationship of the Golgi Apparatus/Centrosome between them and with the cell periphery by characterising new molecular complexes that participate in several processes with special attention to the adhesion/migration and to the cell division.


The interface Golgi Apparatus/Centrosome probably involves structural and biochemical mechanisms which are dependent or not of MTs. The number of proteins that participates in this interaction constantly increases. In general, these are large proteins with large domains folded as coiled-coil structures that establish interactions among themselves and with other types of regulatory proteins. Our hypothesis is that the establishment of interactions between proteins located in the pericentriolar material, in the Golgi Apparatus or both would allow the construction of a network on which the GA is organized. The dynamics of this network would depend not only on cytoskeleton activity but also on membrane trafficking. AKAP450 (Rivero et al., EMBO J, 2009), GMAP210 (Rios et al., Cell, 2004 ) and CAP350 (Andersen et al., 2003) are examples of such proteins.

We are using cell micropatterning, recombinant antibodies and live imaging technologies to assess the role of AKAP450 and GMAP210 in polarized membrane trafficking, in the breaking of Golgi Apparatus/Centrosome connection at the onset of mitosis, and in the dynamics of actin cytoskeleton. We are also characterizing several molecular complexes involving these proteins that have been revealed by a two-hybrid screening.

Finally,Golgi fragmentation has been observed in several neurodegenerative diseases including amyotrophic lateral sclerosis, Alzheimer and Parkinson’s disease (PD). We want to study  the relationship between Golgi fragmentation and alpha-synuclein aggregation and to determine whether fragmentation is a conditioning step in the neurodegenerative process. We will focus in an experimental model of PD using the human neuroblast SH-SY5Y cell line which will be transfected with wild or mutant alpha-synuclein DNA and co-treated with the reversible proteasome inhibitor MG132 or the mitochondrial complex I inhibitor rotenone.

Hurtado L, Caballero C, Gavilan MP, Cardenas J, Bornens M, Rios RM. Disconnecting the Golgi ribbon from the centrosome prevents directional cell migration and ciliogenesis. J Cell Biol. 2011 May 30;193(5):917-33. Epub 2011 May 23.

Kierszenbaum AL, Rivkin E, Tres LL, Yoder BK, Haycraft CJ, Bornens M, Rios RM. GMAP210 and IFT88 are present in the spermatid golgi apparatus and participate in the development of the acrosome-acroplaxome complex, head-tail coupling apparatus and tail. Dev Dyn. 2011 Mar;240(3):723-36. doi: 10.1002/dvdy.22563. Epub 2011 Feb 10.

Chakarova CF, Khanna H, Shah AZ, Patil SB, Sedmak T, Murga-Zamalloa CA, Papaioannou MG, Nagel-Wolfrum K, Lopez I, Munro P, Cheetham M, Koenekoop RK, Rios RM, Matter K, Wolfrum U, Swaroop A, Bhattacharya SS. TOPORS, implicated in retinal degeneration, is a cilia-centrosomal protein. Hum Mol Genet. 2011 Mar 1;20(5):975-87. Epub 2010 Dec 15.

Gavilán MP, Pintado C, Gavilán E, Jiménez S, Ríos RM, Vitorica J, Castaño A, Ruano D. Dysfunction of the unfolded protein response increases neurodegeneration in aged rat hippocampus following proteasome inhibition. Aging Cell. 2009 Dec;8(6):654-65. Epub 2009 Sep 11.

 

Rivero S, Cardenas J, Bornens M, Rios RM. Microtubule nucleation at the cis-side of the Golgi apparatus requires AKAP450 and GM130.   EMBO J. 2009, 28(8):1016-28. Comment in: A. Kodani and C. Sutterlin. EMBO J. 2009, 28:995-996

Nozawa, K., Ikeda, K., Satoh, M., Reeves, W.H., Stewart, C., Li, Y.C., Yen, T.J., Rios, R.M., Takamori, K., Ogawa, H., Sekigawa, I., Takasaki, Y., and Chan, E.K.L. Autoantibody to nuclear antigen NA14 is an independent marker primarily for Sjogren’s syndrome. Frontiers Biosci. 2009, 14, 3733-3739.

G. Egea and Rios R.M. The role of the cytoskeleton in the structure and function of the Golgi apparatus. In The Golgi apparatus. State of the art 110 years after Camilo Golgi’s discovery. Editors: A. Mironov, M. Pavelka, and A. Luini. Springer-Verlag, 2008, 270-299

Rios R.M., Sanchís A, Tassin AM, Fedriani C and Bornens M. GMAP-210 recruits gamma-tubulin complexes to cis-Golgi membranes and is required for Golgi ribbon formation. Cell, 2004, 118(3):323-333
Comment in: A.D. Linstedt. Cell, 2004, 118(3):271-272

Rios RM and Bornens M. The Golgi Apparatus at the cell center. Curr. Opin. Cell Biol. 2003, 15(1), 60-66

Pfannenschmid, F., Wimmer V.C., Rios R.M., Geimer S.,  Kröckel U., Leiherer A., Haller K., Nìmcová, Y., and Mages, W. Chlamydomonas DIP13 and human NA14: a new class of proteins associated with microtubule structures is involved in cell division. J. Cell Sci. 2003, 116:1449-1462

Pernet-Gallay K, Antony C, Johannes L, Bornens M, Goud B, Rios R.M. The Overexpression of GMAP-210 Blocks Anterograde and Retrograde Transport Between the ER and the Golgi Apparatus.   Traffic 3, 822-832,  2002

Infante, C., Ramos-Morales, F., Fedriani, C., Bornens, M. and R.M. Rios.  GMAP-210, a cis-Golgi network-associated protein, is a minus end microtubule-binding protein  J. Cell Biol. 145, 83-98, 1999

Ramos-Morales, F., C. Infante, C. Fedriani, M. Bornens and R.M. Rios. NA14 is a novel autoantigen with a coiled-coil domain. J. Biol. Chem. 273, 1634-1639, 1999