Regulation of gene transcription by retinoic acid receptor heterodimer and its coregulators

Regulation of gene transcription by retinoic acid receptor heterodimer and its coregulators (PIs: N. Sibille & P. Bernadó)

People Involved: N. Sibille, P. Bernadó, F. Allemand, A. Fournet, T. Cordeiro, L. Sénicourt, A. Sagar

Nuclear Receptors (NRs) are transcription factors with a direct role in regulating the expression of hormone-response genes. Agonist binding to NRs triggers transcription through a cascade of events that is initiated by the exchange of the repression complex by the activation one. The assembly of the two large complexes is mediated by intrinsically disordered coregulator proteins that, simultaneously, also interact with the NRs. Coregulators are named coactivators or corepressors when they mediate the activation or repression complexes, respectively. In collaboration with A. le Maire, W. Bourguet and P. Germain we have dissected the structural bases of the interaction between both families of coregulators with the Retinoic Acid Receptor heterodimer (RAR/RXR). Funded by the Labex EpiGenMed, we have studied the interaction of the intrinsically disordered NCoR (corepressor) and TIF2 (coactivator) with RAR/RXR, which form highly dynamic and malleable complexes.

We reached to the following conclusions for the corepressor NCoR (Cordeiro et al., 2019): (i) Small ligands induce disorder to order transitions in the C-
terminal helix of the Ligand Binding Domain (LBD) of both RAR and RXR, and they occur
independently of the presence of coregulator proteins.
(ii) We demonstrated that the NR binding region of NCoR is intrinsically disordered, but it encompasses three highly conserved regions that are partially structured that are linked to RAR/RXR recognition. (iii) The NCoR complex with RAR/RXR is highly plastic as N-CoR is bound to the heterodimer in a singly- and a doubly-bound form (Fig. 3). The relative populations of these complexes can be modulated with small ligands and point mutations.
The main results derived from the study of the coactivator TIF2 are (Sénicourt et al.,
in preparation): (i) We demonstrated by NMR that the NR binding region of Tif2 is intrinsically disordered, but it encompasses three highly conserved regions that are partially structured. (ii) In addition of the three well-known NR interacting sites, we have highlighted importance of a flanking region in strengthening the interaction with RAR/RXR. Initially identified by NMR, this flanking region has been successfully co-crystalized with the nuclear receptor, displaying a helix-turn-helix motif. Further studies will be performed to dissect the relevance of this interaction.
These results provide the structural and functional clues for the regulation of gene transcription and suggest mechanistic pathways for the transition from basal (inactive) state to the transcriptionally active one.

Main collaborators: A. LeMaire, P. Germain, W. Bourguet, A. Barducci (CBS, Montpellier)
Grant: Labex EpiGenMed

Computational tools for modeling and data analyses

Computational tools for modeling and data analyses (PIs: P. Bernadó & N. Sibille)

People Involved: P. Bernadó, N. Sibille, F. Allemand, F. Herranz-Trillo, A. Estana, A. Sagar

During the present contract the group has developed an intense activity in developing new tools for the structural modeling of flexible systems as well as the comprehensive analysis of complex SAXS data. In the following paragraphs the most relevant results on these two research lines will be described.


- Structural modeling of Disordered Proteins: Structural modeling of IDPs is extremely challenging as their special features are coded in the conformational bias at the residue-level. In order to solve of these challenges, we have developed a tripeptide database from high-resolution structures in collaboration with Juan Cortés that allows to build realistic IDP ensemble models for these proteins (Estaña et al, 2019). The resulting ensembles are in agreement with extensive experimental NMR and SAXS data. The enhanced performance of this approach is based on the inclusion of the specific chemical and structural properties of the neighboring residues. The tripeptide database is extremely rich and can also be used to decipher the folding pathway of small structural elements of proteins (Estaña et al., 2019) and the prediction of partially structured elements in IDPs from their sequences (Estaña et al., in preparation).

teamA2 fig4 disorder pred

- Chemometric analysis of SAXS data from complex biomolecular systems: Many biological systems present distinct species that are in equilibrium. These polydisperse systems are extremely difficult to characterize as the experimental observables are population-weighted averages. SAXS is very sensitive to the overall size and shape of particles and, therefore, it can monitor changes when the equilibrium is perturbed by modifying the experimental conditions (concentration, pH, time...). We have developed a chemometric approach, which we call COSMiCS, to decompose large population-weighted SAXS datasets. COSMiCS uses multiple SAXS data representations to decrease the degeneracy of possible solutions. As a proof of concept, we applied COSMiCS to study the fibrillation of insulin and α-synuclein by SAXS for which the population and structure of the three main species (monomer, oligomer and fibril) in equilibrium were derived (Herranz-Trillo et al., 2017).

Main collaborators: J. Cortés (LAAS, Toulouse), B. Vestergaard (U. Copenhagen), R. Tauler (CSIC, Bercelona)

Autres projets du groupe

 

RDC under pressure (Nathalie Sibille)

   Collaboration : Christian Roumestand and Cathy Royer, CBS, Montpellier


   Accurate description of protein folding and stability, in normal and pathological conditions, has been a major area of biophysical chemistry for over 50 years. Although much progress has been made in recent years to achieve this objective, a number of important questions remain. The physical basis of pressure effects on the structure and stability of the protein remained a mystery unlike the effects of temperature.
   We have recently characterized several alignment media that are amenable to high-pressure. This enables the measurement of RDCs in a broad range of pressures that will enable the structural characterization of protein states emerging under these conditions. For more details see :
    Sibille N*, Delarole M, Royer C, Roumestand C. Measuring Residual Dipolar Couplings at High Hydrostatic Pressure : Robustness of Alignment Media to High Pressure. J Biomol NMR. (2014).

 

 

Advanced Computational Approaches to study protein motions. (Pau Bernadó)


Collaboration : Juan Cortés (LAAS – Toulouse)

   The study of flexible systems at atomic level requires detailed description of the structure and the mechanisms inducing their fluctuations. In that context, our group is interested in advanced computational approaches allowing the exploration of the conformational space in an efficient manner to study mobility in broad time-scales. We are interested in Normal Mode approaches that perturb 3D structures in a physically meaningful manner. Derived conformations can be evaluated using SAXS data, which are exquisitely sensitive to large conformational fluctuations.
    Robotics-inspired algorithms have been demonstrated to be very efficient in sampling the 3D space available to macromolecules. In collaboration with Juan Cortés (LAAS-Toulouse) we aim to apply these approaches to study conformational transitions in IDPs.

Connexion