In bacteria, as in most prokaryotes, the regulatory region, called the promoter, resides within a stretch of nucleotides located a short distance—often as few as 10 nucleotides—in front of (upstream from) the start of the coding region. For transcription to proceed accurately and efficiently, RNA polymerase must attach to the promoter. Once it is so positioned, it slides over to the start of the coding region and rides along the DNA, like a train on a track, constructing an RNA replica of the coding sequence. Except in very long genes, the number of RNA molecules made at any moment depends mainly on the rate at which molecules of RNA polymerase attach to the promoter and initiate transcription.
Interestingly, RNA polymerase is a rather promiscuous molecule, unable to distinguish between the promoter and other DNA sequences. To direct the enzyme to promoters of specific genes, bacteria produce a variety of proteins, known as sigma factors, that bind to RNA polymerase. The resulting complexes are able to recognize and attach to selected nucleotide sequences in promoters. In this way, sigma factors program RNA polymerase to bypass all nonpromoter sequences and to linger only at designated promoters.
Considering the importance of sigma factors to the differential activation of genes in bacteria, my colleagues and I began our inquiry into the human transcription apparatus by searching for sigmalike molecules in human cells. But we had underestimated the complexity of the machinery that had evolved to retrieve genetic information from our elaborate genome. It soon became apparent that human sigma factors might not exist or might not take the same form as they do in bacteria.
If there were no simple sigma factors in eukaryotes, how did such cells ensure that RNA polymerase transcribed the right genes at the right time and at the right rate? We began to see glimmerings of an answer once the unusual design of eukaryotic genes was delineated.
By 1983 investigators had established that three kinds of genetic elements, consisting of discrete sequences of nucleotides, control the ability of RNA polymerase to initiate transcription in all eukaryotes—from the single-celled yeast to complex multicellular organisms. One of these elements, generally located close to the coding region, had been found to function much like a bacterial promoter. Called a core promoter, it is the site from which the polymerase begins its journey along the coding region. Many genes in a cell have similar core promoters.
Walter Schaffner of the University of Zurich and Steven Lanier McKnight of the Carnegie Institution of Washington, among others, had additionally identi fied an unusual set of regulatory elements called enhancers, which facilitate transcription. These sequences can be located thousands of nucleotides upstream or downstream from the core promoter—that is, incredibly far from it. And subsequent studies had uncovered the existence of silencers, which help to inhibit transcription and, again, can be located a long distance from the core promoter.
In a somewhat imperfect analogy, if the core promoter were the ignition switch of a car engine, enhancers would act as the accelerator, and silencers as the brakes. Eukaryotic genes can include several enhancers and silencers, and two genes may contain some identical enhancer or silencer elements, but no two genes are precisely alike in the combination of enhancers and silencers they carry. This arrangement enables cells to control transcription of every gene individually.
Discovery of these elements led to two related—and, at the time, highly surprising—conclusions. It was evident that enhancers and silencers could not control the activity of RNA polymerase by themselves. Presumably they served as docking sites for a large family of proteins. The proteins that bound to enhancers and silencers—now called activators and repressors—then carried stimulatory or repressive messages directly or indirectly to RNA polymerase (that is, pressed on the accelerator or on the brakes). It also seemed likely that the rate at which a gene was transcribed would be dictated by the combined activity of all the proteins—or transcription factors—bound to its various regulatory elements.