Probing Gene Activation

Photo: DNA

Switching on a gene -
researchers copied the process;
© NCI Visuals Online

“The new methodology allows us to examine the steps that turn on individual genes in order to figure out what part of the process breaks down in diseases like cancer,” says lead researcher Danny Reinberg.

DNA, in its simplest form, is a long double-stranded helix. Inside the cell’s nucleus, however, the helix is further twisted and wrapped around protein complexes to form much more compact fibres called chromatin.

The degree of compaction is dynamic and is part of the way cells control gene transcription. When a gene is inactive, its chromatin is packed together more tightly than when the gene is actively transcribed. Although scientists have known about these changes for years, they didn’t know how the cell regulated the shift from the most highly compact fibre, which is thirty nanometres across, to the less compact one, which is just eleven nanometres.

They wanted to understand the process, but faced a major hurdle: How to recreate a thirty nanometre fibre in a test tube? The researchers have cleared that hurdle, allowing the team to begin teasing apart the steps to unwind the fibre and start transcription in a test gene.

Once the fibre was in the assay tubes, the team found that a DNA regulatory protein called the retinoic acid receptor (RAR) could access its binding site on the DNA, even when the chromatin was in its most tightly wound state. When researchers added the hormone that stimulates RAR to the system, things started to change. The hormone-bound RAR began to unwind the chromatin and move aside large protein complexes, called nucleosomes, to make room for other DNA unwinding proteins and transcription factors.

As they added in more and more factors, the DNA continued to loosen up, until finally, the team could see transcription start from their test gene PEPCK, which is controlled by RAR. Significantly, electron microscopy showed that the 30 nanometre fibre formed in the test tube resembles the one in cells.; Source: New York University Langone Medical Center