The DNA Microarray: It's the Proof in the Protein Pudding

Image: ©1999-2002 Elsevier Science TESTING THE PUDDING: Using clustering of gene expression profiles, researchers showed that yer083c maybe implicated in cell wall biosynthesis. Direct testing of that hypothesis verified this connection. (Reprinted with permission Cell, 102:109-26, 2000.) As genomics morphs into proteomics and the quest to understand the biological functions of proteins accelerates, researchers continue to look for the best methods that will speed their understanding; one of them is the use of DNA microarrays. Stephen Friend, vice president of basic research at Merck Research Laboratories in West Point, Pa., and his research team demonstrated in this Hot Paper how to compare a gene expression profile in yeast cells to 300 "compendium" profiles, and then infer which of the cell's molecular pathways may have been enhanced or disrupted.1 "The concept of this was that you could get the equivalent of functional signatures from DNA or expression arrays," he says. The key is pattern recognition. The team generated expression profiles for 300 mutants and chemical treatments and then used this library as a reference when they captured array data. Most attempts to figure out a gene's function focuses on the gene itself. "People hadn't realized that you learned much more by using the genome as a sensor pad to listen to the changes that happen in a cell, as opposed to just the gene you cared about. What this method said was, if you want to follow the function of [a gene] ... capture what happened in the rest of the cell," says Friend. Data derived from the Science Watch/Hot Papers database and the Web of Science (ISI, Philadelphia) show that Hot Papers are cited 50 to 100 times more often than the average paper of the same type and age. T.R. Hughes et al., "Functional discovery via a compendium of expression profiles," Cell, 102:109-26, July 7, 2000. (Cited in 206 papers) The strategy's success surprised some who did not expect the pattern recognition to be sensitive enough to identify changes in specific genes. Another surprise: "The uniqueness of the signatures as you knocked out genes all in the same pathway. We went down the cholesterol lowering pathway, and got a different signal as we walked down," says Friend. That sensitivity could help researchers as they puzzle over the role of specific proteins in normal and diseased states. Friend's team created deletion mutants of several genes of unknown function and assigned them to roles in sterol metabolism, cell wall function, mitochondrial respiration, and protein synthesis. Since then, the assignment list has grown. "The compendium approach," Friend says, "is working quite well for more complex organisms, where you might have thought it would fracture into 10,000 compendiums that you would have had to build up. [In unpublished results], we've now used this technique in looking at blood from patients." Pediatrics professor Gary W. Litman, All Children's Hospital at the University of South Florida, St. Petersburg, says, "This is exceptionally important [work], particularly from the standpoint of potentially identifying functions of the large numbers of predicted proteins that are coming out of genome sequencing efforts." Litman adds that in most cases, commonly used targeted gene disruption methods have so far failed to identify a phenotype or lethality. Since the paper's publication, Friend's team analyzed samples from women recovering from breast cancer, using pattern matching to identify the metastatic potential at an early stage of the disease. The pattern of changes in fewer than 100 genes predicted patients who would have distant metastases, with 83% accuracy.2 Other researchers have performed a similar analysis on data from patients with ovarian cancer,3 and in another case, Friend's team used the compendium approach to identify the target of a protease inhibitor.4 The team also treated cells with the commonly used topical anesthetic dyclonine. Pattern matching suggested that it was impacting the Erg2p receptor, a yeast homolog of the human Sigma receptor, which interacts with neurosteroids and positively regulates potassium conductance. "That told us the type of protein that it was inhibiting, how it might be acting as a neuroactive drug," Friend says. Such results are not the be-all and end-all of the search process, but "it enriched what you were thinking. It does not lock onto the human target, but it gives you a lead as to what you might do as your next experiment in mammalian cells." Jim Kling (jkling@nasw.org) is a freelance writer in Washington, DC. References 1. T.R. Hughes et al., "Functional discovery via a compendium of expression profiles," Cell, 102:109-26, July 7, 2000. (Cited in 206 papers) 2. L.J. van't Veer et al., "Gene expression profiling predicts clinical outcome of breast cancer," Nature, 415:530-6, 2002. 3. E.F. Petricoin et al., "Use of proteomic patterns in serum to identify ovarian cancer," The Lancet, 359:572-7, 2002. 4. R. Ulrich, S.H. Friend, "Toxicogenomics and drug discovery: Will new technologies help us produce better drugs?" Nature Reviews, 1:84-8, 2002.

The DNA Microarray: It's the Proof in the Protein Pudding

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