Molecular dynamics simulations of the glucocorticoid receptor DNA-binding domain suggest a role of the lever-arm mobility in transcriptional output.

One of the first and essential steps in gene expression regulation involves the recruitment of transcription factors (TFs) to specific response elements located at enhancers and/or promoters of targeted genes. These DNA elements have a certain variability in both sequence and length, which may affect the final transcriptional output. The molecular mechanisms in which TFs integrate the subtle differences within specific recognition sequences to offer different transcriptional responses is still largely unknown. Here we used molecular dynamics simulations to study the DNA binding behavior of the glucocorticoid receptor (GR), a ligand-regulated TF with pleiotropic effects in almost all cells. By comparing the behavior of the wild type receptor and a well characterized Ala477Thr substitution within the rat GR DNA binding domain, we found that the region that connects the two-zinc fingers (i.e. the lever arm) would likely play a key role in GR transcriptional output

The A477T mutation alters the GR DBD dimerization interface.

<p>The A477T mutation alters the GR DBD dimerization interface.</p

Complexes that are more active have lever-arms with a reduced mobility.

<p>a) RMSF values of chain A residues of S1, S2, S3 and S4 systems. b) RMSF values of chain B residues of S1, S2, S3 and S4 systems. The secondary structure of GR DBD is schematized along the x-axis. c) Principal Component Analysis of MD trajectories: projection on the first two eigenvectors of lever arm and D-loop CA atoms in S1, S2, S3 and S4 systems.</p

The absence of the GR DBD dimer partner provokes a strong dynamical alteration on the D-loop.

<p>a) RMSF values of the S1 system, chain A in black and chain B in red, S1A system in purple and S1B system in yellow. The secondary structure of the GR DBD is schematized along the x-axis. b) Principal component analysis of the MD trajectories: projection on the first two eigenvectors in S1, chain A in black and chain B in red, S1A system in purple and S1B system in yellow. c) Average structures of S1, chain A in black and chain B in red, S1A system in purple and S1B system in yellow. d) Time evolution of secondary structure propensities for the S1 system.</p

Simulated GR DBD/DNA complexes.

<p>Simulated GR DBD/DNA complexes.</p

The GRE spacer affects the minor groove width.

<p>Variation of the average minor groove width along the oligomer in S1 (black), S2 (green), S3 (blue) and S4 (magenta) (x = G for S1 and S3 systems, x = A for S2 and S4 systems).</p

The GRE spacer sequence alters the propeller and the slide base-pair parameters.

<p>Time evolution of propeller and slide values for S1 (black), S2 (green), S3 (blue) and S4 (magenta) systems. Schematic representations were extracted from Ref. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0189588#pone.0189588.ref024" target="_blank">24</a>].</p

The A477T mutation affects the dimerization interface.

<p>Overall view of the average structures of the S1 (a) and S2 (b) systems showing the location of the dimerization zinc fingers and residues 477.</p

The GR DBD/Fkbp5 complex.

<p>a) Primary and secondary structure of the GR DBD. b) Representation of the crystal structure of the GR DBD/Fkbp5 complex (pdb:3g6p, chain A).</p

Complexes that are more active have lever-arms with a reduced mobility.

<p>a) RMSF values of chain A residues of S1, S2, S3 and S4 systems. b) RMSF values of chain B residues of S1, S2, S3 and S4 systems. The secondary structure of GR DBD is schematized along the x-axis. c) Principal Component Analysis of MD trajectories: projection on the first two eigenvectors of lever arm and D-loop CA atoms in S1, S2, S3 and S4 systems.</p