Researchers at the Whitehead Institute and Corning Inc. have invented a powerful new microarray technique that can decipher the function of master switches in a cell by identifying the circuit, or the set of genes, they control across the entire genome.
The researchers show that the technique can correctly identify the circuits controlled by two known master switches in yeast. In addition, the technique allows researchers to unravel in a week what takes years to achieve by conventional methods.
"We are very excited by these results because they suggest that our technique can be used to create a "user's manual" for the cell's master controls, a booklet that matches the master switches to the circuits they control in the genome," said Professor of Biology and Whitehead member Richard Young, who led the study.
The technique, published in the December 22 issue of Science, also gives researchers new scientific muscle needed to piece together the master wiring diagram -- the controls and the circuits -- that operate the complicated machinery of life.
Creating such a diagram represents the next step toward using the information from the Human Genome Project. Although the Human Genome Project will soon provide researchers a catalog of all the genes that make up a human being, it will in many ways be analogous to having the complete parts list for a Boeing 777, say researchers. The information does not tell us anything about putting all the parts together, nor does it tell us how the cockpit controls function to make the plane fly.
"Our technique creates the documentation needed to put the parts together and identifies how the major controls are connected to these parts," said Professor Young. Such information will be fundamental to finding the genetic basis of diseases and for discovering better drugs.
The technique also will help solve many unanswered questions in cancer research, he said. Malfunctioning of master switches has been shown to lead to cancer, but little is known about the nature of the circuitry they control.
The genome's master switches are DNA-binding proteins called gene activators. In humans, there are about 1,000 such activators controlling important functions in life, including cell growth and development. Some of the best known of these switches -- the p53 protein, for example -- are those that play a role in cancer. Others play a role during development, designating which cells become nerve or muscle cells, for instance.
Scientists know the identity of nearly 600 master switches and know the function of at least 250 of those; their hope has been to find the set of all genes controlled by the master switches so they could crack open the genetic basis of health and disease.
However, finding all the genes -- i.e., the circuitry -- directly controlled by any given master switch has been a painstakingly long and tedious process, involving years of biochemical and molecular experiments. The new technique reported in Science provides a way to get the data in a global fashion and will allow researchers to do in a week what would have taken years to achieve.
"Our technique could conceivably be used in human cells to create a map matching up the master switches with the circuits they control," said Bing Ren, a postdoctoral associate in the Young lab.
Although DNA arrays are useful in determining a cell's expression profile (a snapshot of which genes are turned on and off in a cell), they represent an overall picture and capture the cell's state at a moment in time. One perturbation in the environment or a slight change in the tumor of which the cell is part could trigger a cascade of changes, all of which are captured in the snapshot. Such information is invaluable to researchers, but when it comes to identifying the one crucial master switch, finding it from DNA arrays can be like finding a needle in the haystack.
In this study, the Young lab scientists created a technique to overcome this problem. The technique involves first fixing DNA-binding proteins in living cells to their binding sites using chemical cross-linking methods and then breaking open the cells to create a molecular soup of DNA-protein complexes. Specific antibodies coupled with magnetic beads are then used to fish out DNA fragments cross-linked to proteins of interest. This provides researchers with a pure population of DNA-protein complexes. Unhooking the cross-linked DNA from the protein leaves them with DNA fragments that bind to proteins of interest. The researchers then label these fragments with fluorescent dye and hybridize them to a DNA array containing genomic DNA from yeast to reveal their identity.
"Our goal is to use this technique to find the circuits controlled by the 200 or so master switches in yeast and then develop analogous techniques in humans," said Professor Young.
This work was supported by Corning Inc., the National Institutes of Health, the Helen Hay Whitney Foundation, the National Cancer Institute of Canada, the National Science Foundation, the Howard Hughes Medical Institute, the European Molecular Biology Organization and the Human Frontier Science Foundation.
A version of this article appeared in MIT Tech Talk on January 10, 2001.