Over the past three decades, this addiction to scientific discovery has driven Tjian to prominence in gene-regulation research. Since 1973, he has focused on transcription, the process by which cells "read" information from their DNA to make RNA and eventually proteins. His investigations tackle a fundamental mystery of life. Add sperm to egg, and nine months later, you've got a miraculous, wailing being with billions of specialized cells, from red blood cells to neurons. But what makes a neuron a brain cell, and not a blood cell? It all depends on which genes are switched on or off.
"Somehow, we [humans] have evolved a system that allows us a very detailed, extraordinarily elaborate readout of information from the genetic blueprint to not only make a human being but then to maintain life for 80-something years," Tjian says. "During that entire time, every cell in your body has to do the right job. And that all boils down to which genes are being transcribed." To make each of the myriad proteins that carry out life's chores, cells must retrieve the right bits of genetic information accurately and at exactly the right time.
When Tjian, now 52, began studying transcription in mammalian cells, this fundamental biological process was a black box. What little was known had been pioneered by biochemist Robert G. Roeder, who had discovered the enzyme RNA polymerasethe heart of the engine that drives the cell's DNA-reading machineryin work he did at the University of Washington. Few other scientists thought transcription was worth studying. They figured that cells simply copied all of their genes into an intermediary moleculeheterogeneous nuclear RNA (hnRNA)and then just tossed out whatever wasn't needed. In other words, scientists envisioned the cell's DNA-scanning and RNA-making factory as largely automated and unregulated, with no direct role in specific gene expression.
That view has been shattered. Researchers have found that regulatory proteins, called transcription factors, guide RNA polymerase to scan only certain genes by telling the enzyme exactly where to start its work. The factory floor, it turns out, is teeming with more highly skilled laborers than anyone imagined. Drug makers now see the DNA-reading transcription machinery as a new target for treating illnesses such as asthma, cancer and heart disease, which arise in part from a shortage or excess of certain proteins. Novel medicines designed to tweak specific transcription factors could spur or inhibit the expression of genes for those proteins.
Tjian made his initial foray into the transcription field after finishing college at UC Berkeley in 1971 and spending a year at Oxford University. He began by exploring the DNA-reading process in bacteria during graduate school at Harvard University, earning a Ph.D. in 1976. He did postdoctoral research at Cold Spring Harbor Laboratory in New York at the invitation of Nobel laureate James Watson. In 1977, Tjian gained a reputation as a hot young scientist by isolating the first nonbacterial DNA-binding protein and transcription factor, called T-antigen. Returning to California in 1979 to become an assistant professor at UC Berkeley, he undertook the work for which he is best known: He and his group discovered Sp1, the first gene-specific transcription factor ever found in human cells.
Tjian isolated the gene for Sp1 and showed how the factor directly attaches to and interacts with DNA at a specific target site, says Richard Losick, a gene-regulation researcher at Harvard University who was Tjian's Ph.D.-thesis adviser. "Those were electrifying experiments back in those early days," Losick says, especially when one considers that the techniques then available were primitive by today's standards. Since then, Tjian and his lab colleagues have hunted down and purified more than 100 transcription factors. "In my view," Losick adds, "the two stars of the transcription field in that era were Bob Roeder and Tij."
East to West
"Tij" (pronounced "teej") is what friends call Tjian, who was born in Hong Kong in 1949, the youngest of nine children. After fleeing China during the Communist Revolution, his family bounced around Argentina and Brazil, and then moved to New Jersey in 1963. Six years later, as a UC Berkeley sophomore, Tjian boldly talked his way into the laboratory of biochemistry professor Daniel E. Koshland, Jr. Tjian visited Koshland and asked outright if he could join his research group. The scientist, already well known for his work on enzyme catalysis, said nothe kid hadn't even taken biochemistry yet. When Tjian persisted, Koshland gave him the weekend to read an advanced text called Protein Structure. Afterward, the professor quizzed him. "It was clear he hadn't just read it, he really understood it," Koshland recalls. "So I said, okay, you can be in my lab."
When Tjian graduated, the professor took him along on a year-long research sabbatical at Oxford. "He was very dedicated, a really hard worker," Koshland says. "He's a very inventive and imaginative scientist. He's willing to do new things, even if everybody else says it's risky."
Those qualities, and a now-legendary technical prowess, sped Tjian through graduate school at Harvard in just three and a half years. "He's the most gifted experimentalist I've encountered," Losick says. "He was fearless and had a kind of special intelligence for making things happen on the bench-top."
Find the Activator
As a young professor at Berkeley, Tjian began tackling the mystery of transcription in animal cells. His work focused on a nagging question. Scientists knew that RNA polymerase must touch down on a gene's "promoter"a set of nucleotides punctuating the start of every genebefore chugging down the DNA track to read it and manufacture messenger RNA (mRNA). However, given that the polymerase can read any DNA it encounters, how was it able to choose the correct gene to read?
Tjian had already helped prove that in bacteria, proteins called sigma factors plugged into the RNA polymerase and preferentially instructed it to transcribe specific genes. Now, he and his research team began searching for a mammalian version of the sigma factors by studying how a virus called SV40 infects monkey and human cells. Like all viruses, SV40 replicates by hijacking the transcription machinery of the host cell. The researchers took a human cell extract containing the transcription apparatus and separated out its different proteins. When they mixed all the components back together with SV40 DNA in a test tube, they were able to recreate the virus' action: The host machinery was tricked into scanning viral, rather than human, DNA.
The scientists hoped to find the specific human transcription factor co-opted by the virus to pull off this stunt. So, in a series of experiments, they began testing the cell proteins one by one to see whether the same reaction would fail to occur if the protein wasn't added. "It's as if you have a whole bagful of stuff and pick out molecules from that pool one by one, asking, 'Can I throw that one away, or do I really need it?' " says Tjian. "It was just like looking for the needle in the haystack. It was really remarkable it even worked."
The culprit factor, Sp1, emerged from Tjian's lab in 1983 in work carried out by postdoc William Dynan. However, the protein wasn't what anyone expected. Unlike a sigma factor, it didn't bind to the polymerase. Instead, it stuck to a specific stretch of DNA called the GC box. Another year of studies revealed that Sp1 was an "activator," a transcription factor that speeds up the reading of specific genes. ("Repressors," on the other hand, slow or stop transcription.)
In 1985, another Tjian postdoc, James Kadonaga, purified Sp1 extracted from human cellsno mean feat, because cells contain vanishingly small quantities of transcription factors. Using a technique called affinity chromatography, Kadonaga, a chemist, attached copies of the GC box to tiny beads, which he then loaded into a glass cylinder. When he poured a crude protein extract through the column, only Sp1 stuck to the beads.
It was a breakthrough that galvanized efforts worldwide to find similar factors, says Losick. "Tjian's lab pioneered the methodology for hunting down these rare proteins and really set the standard for the entire field." Since then, many hundreds have been identified, and results from the Human Genome Project now suggest that anywhere from 2,000 to 3,000 of our approximately 30,000 genes code for such factors.
Building a Bridge
With activator-protein Sp1 in hand, Tjian's group had yet another puzzle to solve. Given that the GC box lies upstream from the gene it regulates and RNA polymerase interacts directly with the gene, how did Sp1 communicate with RNA polymerase?
By the mid-1980s, Roeder and his colleagues at The Rockefeller University had detected several other basic, or basal, components that, along with RNA polymerase, made up the core engine of the transcription machine. Scientists thought that cells of all types used this same engine to scan genes at a low level, like a car that can only move in first gear. To crank up or halt readout of particular genes, however, each different cell type would rely on its own, customized set of activators and repressors.
One critical basal transcription factor that Roeder's group found was TFIID. Its main ingredient was thought to be a proteindubbed TBPknown to seek out and bind to the TATA box that lies within many gene promoters. As expected, when researchers put copies of purified TBP into a test tube with DNA, RNA polymerase and other basal components, transcription levels were low. In theory, adding an activator to the same mixture should have kicked transcription into high gear. A postdoc in Tjian's lab, B. Franklin Pugh, was the first to attempt this, using Sp1. But in a flummoxing turn of events, it didn't work. Something was missing. "We called this missing thing a co-activator," Tjian says.
Pugh and Tjian proposed in 1990 that co-activators were a third new class of transcription proteins that serve as a bridge between activators and the RNA polymerase. Other researchers were skeptical, but two years later, the Berkeley scientists purified a cluster of proteins called TAFs (TBP-associated factors) that work along with TBPand, within this grouping, they found the first co-activators. These newly discovered helpers joined up with TBP at the promoter, forming what's now called the TFIID complex, which binds to the TATA box. One end of Sp1 gloms onto the GC box while the other end makes contact with the co-activators. Dozens of co-activators have been identified in the decade since.
From there, Tjian pursued another hunch. He began pondering whether the so-called basal machinery was a misnomer. Maybe, he suggested, different kinds of cells or tissues had specialized versions of that machinery. Again, Tjian's intuition paid off. In studies of fruit flies, mice and humans, his team identified proteins that functioned similarly to TBP. Called TBP-related factors or TRFs, these new factors show up only in certain tissues, such as testicular and ovarian cells. And in separate work published in Science last fall (2001), the group showed that one kind of co-activator, named TAFII105, works selectively in ovarian tissue. When senior postdoc Richard Freiman and graduate student Shane Albright deleted the gene encoding for TAFII105 in female mice, they found that the mutant rodents were all infertile. DNA-screening tests showed that TAFII105 controls genes responsible for proper oocyte formation.
Where do the layers of intricacy end? No one knows, but Tjian keeps digging. One of his current missions is to decipher the structures of TFIID and other co-factors to see exactly how they come together. Another is to learn how transcription factors navigate along and interact with DNA during normal cellular activity, in particular when DNA hasn't yet unwound from its naturally coiled-up state, chromatin. In 1999, collaborating with Berkeley biochemist and HHMI investigator Eva Nogales, Tjian's team created the first three-dimensional images of the transcription engine, the TFIID complex, by using the state-of-the-art techniques of electron microscopy and single-particle image analysis. This work showed that the TFIID complex is shaped like a squat, three-pronged pitchfork that can dock around DNA whether it is in chromatin or single-strand form.
Given the wealth of data his group churns out, it might seem that Tjian commands an army of scientists. In fact, fewer than 20 researchers share the four large interconnected rooms of his lab in Koshland Hall, named after his former mentor. Somehow, he passes on his talents to his staff, Losick says. "They get beautiful data and are able to pull off very complicated experiments."
Tjian himself is too busy these days to do hands-on lab work; his crammed schedule includes lecturing in undergraduate biology and chairing the chancellor's strategic-planning council for UC Berkeley's biological sciences programs. Nevertheless, postdocs and students say he's always available. "He expects the best of us and is extremely supportive," says Andreas Ladurner, a senior postdoc from Italy.
At the same time, the professor is up-front about the level of work he expects from everyone in his lab. "The nine-to-five thing doesn't exist here," he says. "Holidays don't exist. And they know that if I'm in town, I'm going to be here Saturdays and Sundays." What do his wife, Claudia (an attorney), and two daughters (ages 22 and 16) have to say about his workaholic habits? "They think I'm nuts. They've had to put up with me forever like this."
Tjian isn't all work, no play, however, and he does occasionally leave town. Every year, the biochemist spends several weeks pursuing his recreational passionfly fishingin such far-flung places as Russia and New Zealand. Often, he goes with close friend and biotech star David Goeddel, of Genentech fame. It was through fishing with Goeddel, in fact, that Tjian came to launch a biotech company, Tularik, Inc. (see "Spawning a Start-up").