Unlocking the great cell Mediator

Using high-powered electron microscopy (EM), scientists at the Scripps Research Institute in La Jolla, Calif., have uncovered one of the most important cellular mechanisms of gene expression, the Mediator, according to a study published today in the journal Cell. The researchers’ new map of cellular machinery is key to a deeper understanding of cellular processes, including the production of proteins. 

Francisco J. Asturias, PhD, associate professor at the institute, and colleagues zeroed in on the Mediator, which informs the activity of genes in the cells of everything from plants to humans. This is the first validated model of its structure and functionality in a human cell. 

The Mediator concept was first presented almost 20 years ago by biologists at Stanford University. The discovery led to a Nobel Prize. However, a functional model of the Mediator has eluded scientists until now.

Making out such a model is a highly complex endeavor. Researchers have had to create a 3D scheme of all the subunit proteins as well and account for Mediator conformations that take place in order to communicate between all of the transcription machinery. As an example, a model of the Mediator in yeast has 25 different subunits of protein. The human version includes 30 subunits. This is a mammoth task for such a small machine—far too complex for even high-resolution imaging techniques such as X-ray crystallography or nuclear magnetic resonance spectroscopy.

“The multisubunit Mediator, comprising about 30 distinct proteins, plays an essential role in gene expression regulation by acting as a bridge between DNA-binding transcription factors and the RNA polymerase II (RNAPII) transcription machinery,” wrote Asturias et al. “Efforts to uncover the Mediator mechanism have been hindered by a poor understanding of its structure, subunit organization, and conformational rearrangements. By overcoming biochemical and image analysis hurdles, we obtained accurate EM structures of yeast and human Mediators. Subunit localization experiments, docking of partial X-ray structures, and biochemical analyses resulted in comprehensive mapping of yeast Mediator subunits and a complete reinterpretation of our previous Mediator organization model.”

Research of Mediator particles led to the gathering of approximately 85,000 EM images in categories describing conformation. These were then averaged and the result is the most comprehensive 3D model of the Mediator structure available today with a resolution of about 18 Angstroms, or 1.8 billionths of a meter.