Investigating the essential roles of Ubiquitin-like molecules and the Ubiquitin Proteasome System

Manuel S. Rodriguez

Education

  • 1997: Ph.D. Microbiology (Virology). Paris VII University-Institut Pasteur.
  • 1989: BSc in Biology. National Autonomous University of Mexico.

Research Appointments

  • 2012: Group Leader at Inbiomed, San Sebastian, Spain.
  • 2005: Group Leader at CIC bioGUNE, Bilbao, Spain.
  • 2001: Charge de recherche 1 / Centre National de la Recherche Scientifique). Institut Jacques Monod / CNRS, France.
  • 1997: Postdoctoral Fellow at Biomolecular Sciences St. Andrews University- MRC fellow UK.

Research Interests

We are interested on the role of ubiquitin and ubiquitin like molecules such as SUMO and NEDD8 in the regulation of physiological and pathological processes, more specifically during the transition of one stade to the other.

  • Using molecular traps to capture ubiquitin or ubiquitin like proteins, we isolate individual protein models (eg. P53 and NF-ï�«B molecules) or globally modified proteins in response to physiological signals or chemical compounds that are used in distict patient therapies.
  • We also develop new technology or implement new applications of the technology we have already available that could be used to better assez the proceses regulated by the family of ubiquitin-like modifiers and the ubiquitin proteasome system.

SUMOylations Figure 1. Principle of the ubiquitin-traps (TUBEs). TUBEs efficiently purify ubiquitylated proteins from cell extracts. (a) Schematic illustrating traditional GST pulldown, with use of DUB inhibitors IAA and NEM. (b) Schematic illustrating method of lysis in presence of TUBEs and absence of IAA/NEM. TUBEs are allowed to form complexes with poly-ubiquitylated proteins during lysis, and subsequently intact complexes are pulled down using glutathione agarose beads. (c) HEK 293 cells were pretreated with proteasome inhibitor, and stimulated with TNF-α. Cells were lysed in presence IAA/NEM, lysates were clarified and incubated with glutathione agarose beads bound to 20 ug GST-fusion protein. Eluted proteins were immunoblotted for IκBα and poly-ubiquitin. (d) HEK 293 cells were treated as above, and subjected to pulldown using the method described in (b ), adding 20 ug of purifed GST-fusion protein to lysis reaction. Asterisk indicates band corresponding to IκBα on reblotted membrane.

Figure 2. Implementations for Mass Spectrometry identification of Ubiquitylated proteins and interacting cellular factors using TUBEs. 

Selected Publications

Da Silva-Ferrada E, Torres-Ramos M, Aillet F, Campagna M, Matute C, Rivas C, Rodríguez MS, Lang V. (2011) Role of Monoubiquitylation on the Control of IκBα Degradation and NF-κB Activity. PLoS One. 6(10): Oct 12.

http://www.ncbi.nlm.nih.gov/pubmed/22022389
González-Santamaría J, Campagna M, García MA, Marcos-Villar L, González D, Gallego P, Lopitz-Otsoa F, Guerra S, Rodríguez MS, Esteban M, Rivas C. (2011) Regulation of vaccinia virus e3 protein by small ubiquitin-like modifier proteins.
J Virol. 2011 Dec;85(24):12890-900. 
http://www.ncbi.nlm.nih.gov/pubmed/21957283
Sánchez, J., Talamillo, A., Lopitz-Otsoa, F., Pérez, C., Hjerpe, R., Sutherland, J. D., Herboso, L., Rodríguez, M. S. and Barrio, R. Sumoylation modulates the activity of spalt-like proteins during wing development in Drosophila. JBC 285, 33 25841-9 (2010)
http://www.ncbi.nlm.nih.gov/pubmed/20562097
Hjerpe R., Aillet F., Torres-Ramos M., Lang V., Farrás R., Hay RT., and Rodríguez M.S. (2010) Oligomerization conditions Mdm2-mediated efficient p53 poly-ubiquitylation but not its proteasomal degradation.  Int. J. Biochem & Cell Biology 42(5):725-735. 
http://www.ncbi.nlm.nih.gov/pubmed/20080206
Hjerpe R., Aillet F., Lopitz F., Lang V., England., P. and Rodríguez M.S. (2009) Effiecient protection and isolation of ubiquitylated proteins using Tandem Ubiquitin Binding Entities (TUBEs).  EMBO reports. 10(11):1250-8 
http://www.ncbi.nlm.nih.gov/pubmed/19798103
Tatham M., Rodriguez M.S., Xirodimas D., and Hay RT. Detection of protein SUMOylation in vivo. (2009) Nature Protocols 9, 1363.  CITED:1
http://www.ncbi.nlm.nih.gov/pubmed/19730420
Da Silva-Ferrada E, Torres-Ramos M, Aillet F, Campagna M, Matute C, Rivas C, Rodríguez MS, Lang V. (2011) Role of Monoubiquitylation on the Control of IκBα Degradation and NF-κB Activity. PLoS One. 6(10): Oct 12.http://www.ncbi.nlm.nih.gov/pubmed/22022389

González-Santamaría J, Campagna M, García MA, Marcos-Villar L, González D, Gallego P, Lopitz-Otsoa F, Guerra S, Rodríguez MS, Esteban M, Rivas C. (2011) Regulation of vaccinia virus e3 protein by small ubiquitin-like modifier proteins.J Virol. 2011 Dec;85(24):12890-900. http://www.ncbi.nlm.nih.gov/pubmed/21957283

Sánchez, J., Talamillo, A., Lopitz-Otsoa, F., Pérez, C., Hjerpe, R., Sutherland, J. D., Herboso, L., Rodríguez, M. S. and Barrio, R. Sumoylation modulates the activity of spalt-like proteins during wing development in Drosophila. JBC 285, 33 25841-9 (2010)http://www.ncbi.nlm.nih.gov/pubmed/20562097

Hjerpe R., Aillet F., Torres-Ramos M., Lang V., Farrás R., Hay RT., and Rodríguez M.S. (2010) Oligomerization conditions Mdm2-mediated efficient p53 poly-ubiquitylation but not its proteasomal degradation.  Int. J. Biochem & Cell Biology 42(5):725-735. http://www.ncbi.nlm.nih.gov/pubmed/20080206

Hjerpe R., Aillet F., Lopitz F., Lang V., England., P. and Rodríguez M.S. (2009) Effiecient protection and isolation of ubiquitylated proteins using Tandem Ubiquitin Binding Entities (TUBEs).  EMBO reports. 10(11):1250-8 http://www.ncbi.nlm.nih.gov/pubmed/19798103

Tatham M., Rodriguez M.S., Xirodimas D., and Hay RT. Detection of protein SUMOylation in vivo. (2009) Nature Protocols 9, 1363. CITED:1http://www.ncbi.nlm.nih.gov/pubmed/19730420

Partner

Inbiomed

Fellow

Lucia Pirone

Collaborators

Rosa Barrio

LUMC

GSK

Project

In vivo requirements of the UBL modifier SUMO on SALL proteins function during steroidogenesis: D. melanogaster as a model system.