Turning off Genes

MOLECULAR BIOLOGY

A new way to control gene expression has been devised: using synthetic peptide nucleic acids (PNAs) or RNAs to interact directly with genomic DNA.

Research assistant professor of pharmacology Bethany A. Janowski, professor of pharmacology and biochemistry David R. Corey, and coworkers at the University of Texas Southwestern Medical Center, Dallas, show that the expression of specific genes in cells can be shut down by using "antigene" PNAs and RNAs to block transcription of chromosomal DNA (Nat. Chem. Biol., published online July 31, dx.doi.org/10.1038/nchembio724 and 725).

A number of means for inhibiting gene expression are already available, including polyamides and triple-helix-forming agents, which bind double-stranded DNA, and antisense RNA and RNA interference (RNAi) agents, which bind to or induce cleavage of mRNA. Targeting a single-stranded part of chromosomal DNA with antigenes could complement the use of existing techniques--for studying gene function and for treating diseases by curbing errant genes.

"The bottom line is that chromosomal DNA is much more accessible to sequence-specific recognition by synthetic agents and RNA than we had previously believed," Corey says.

Antigene agents recognize genomic DNA when its guard is down. Single-stranded DNA is exposed momentarily when RNA polymerase initiates transcription by opening an approximately 20-base-pair segment of double-stranded genomic DNA, resulting in a transcription start-site structure called the "open complex." Antigene agents of complementary sequence interact with one of the open complex's single DNA strands, causing transcription and gene expression to be blocked.

Because every gene has a transcription start site, the technique may be widely applicable. So far, Corey and coworkers have been able to inhibit the expression of nine genes (of nine tested) in cancer cells.

It's "a major advance in the field," says associate professor of chemistry Bruce Armitage of Carnegie Mellon University, a specialist in DNA recognition. The demonstration of "substantial knockdown of several different genes having different promoters is very encouraging."

Associate professor of chemistry and chemical biology David R. Liu of Harvard University, who specializes in biomolecular evolution, adds that "applications for tailor-made, target-specific regulators of gene expression based on this work naturally come to mind."

Corey and coworkers inhibited the expression of human progesterone receptor B (hPR-B), a key reproductive protein, by directing PNA to the gene's transcription start site. Blocking expression of that protein also reduced expression of a variant, hPR-A, revealing a previously unknown regulatory relationship between the two receptors. The researchers then inhibited expression of those receptors and others by using double-stranded RNAs complementary to sequences in the genes' transcription start sites.

The work builds in part on a study by the late professor of chemistry and biological chemistry David S. Sigman of the University of California, Los Angeles, and coworkers, who reported in 2001 that oligonucleotides complementary to one strand of open complexes inhibited transcription by bacterial polymerase in cell-free assays.

Corey and coworkers speculate that the antigene RNA phenomenon may reflect a previously unappreciated natural process. An exciting topic for further research, Corey says, will be "to determine whether the human body or viruses and bacteria make RNA sequences like this to control gene expression."