Terminator in the Lab

The world is organized by headlines. That’s just the way we humans like to do things, to keep life simple and digestible at a glance. Whether the day’s banner news concerns a movie star landing an Oscar for best actress (shyly murmuring, “First, I’d like to thank…”) or a third baseman punching a line drive over the left-field wall to secure a playoff win, we don’t much care to have the whole supporting cast of players trotted out for their share of hard-earned applause.

Science is no different. Few ventures that involve laboratory research are anything but large, complex team efforts, yet who can name the technicians and assistants who sweated the details for any recent major scientific prize? As a general rule, we don’t make time for those in the shadows, despite their often essential contribution to the ultimate success of the work.

Russian émigré Alexander (“Sasha”) Poltorak, who earned his doctorate in chemistry at St. Petersburg Technical Institute and is now an assistant professor of pathology in the Sackler School of Graduate Biomedical Sciences, played just such a vital supportive role in groundbreaking work involving the triggers of the human immune system in the mid-1990s.

The geneticist Bruce Beutler had been looking for someone to assist him in his research at the Howard Hughes Medical Institute (and later at University of Texas Southwestern Medical Center) in Dallas. A mutual friend of the two men put them in touch; they quickly clicked over their shared scientific appetites (“I knew of his work, and I was fascinated by it,” Poltorak says of Beutler), and by 1993, the brilliant young Russian scientist, then in his early 30s, had moved to the U.S. to take a job as a research associate in Beutler’s lab. His wife, chemist Irina Smirnova, worked beside him in the lab.

“He was absolutely outstanding and had superb cloning skills,” Beutler said of Poltorak during an interview a few weeks after he won the 2011 Nobel Prize in medicine. Beutler went on to describe Poltorak as “the point of the spear” in his laboratory’s aggressive six-year hunt for a mutant gene that would reveal—precisely and for the first time—just how the innate immune system functions at the molecular level. The quest was anything but a walk in the park. “He had to solve problems daily in our endless search,” Beutler says of Poltorak.

The Double Moat of Immunity

The major questions were simply put. As Beutler asks at the top of his laboratory’s website, “How do we ‘know’ when we have an infection? Where are the receptors that alert us?”

Remember that the human immune system has two levels of defense against pathogenic intruders to ward off infection and keep the body safe—something like a double moat around a castle flying the flag of Good Health.

The first defensive level, innate immunity, involves sensors that trigger inflammation to help block the assault. (This was the area of Beutler’s and Poltorak’s interest.) If the invaders break through this first line, the second level of defense, adaptive immunity, works to rebuff the infection through an array of antibodies and killer cells.

Jules A. Hoffmann, the French molecular biologist who shared the 2011 Nobel Prize with Beutler (and immunologist Ralph M. Steinman, who died just days before the prize was announced), had already narrowed the hunt, using fruit flies to scope out the insects’ innate immunity receptors.

In 1996, Hoffmann found that flies with mutations in certain genes, including the so-called Toll gene, were unable to mount an effective defense against bacteria or fungi. As a consequence, these Toll mutants died. Hoffmann reasoned that if the Toll gene was broken, and the flies got sick, maybe the insects’ protective mechanism—their sensor, in other words—was located in the Toll gene. And he was able to prove this through further testing.

Working at about the same time, Beutler and Poltorak went a slightly different way, choosing mice for their research subjects. This was largely because of the depth of genetics research that already had been done on mouse strains at the Jackson Laboratory, and that stood as part of the scientific literature going back to 1968. That knowledge supplied a baseline for their own investigations, which involved a hunt for receptor proteins in the mouse gene whose job was to “recognize” and connect with the bacterial product lipopolysaccharide, or LPS.

If the receptor were functioning properly, LPS would be recognized by the immune system and a measured level of beneficial inflammation would result. If not, LPS levels would rise, followed by septic shock. Septic shock, of course, causes tens of thousands of deaths in the U.S. each year, according to conservative estimates.

Poker Faces

Beutler’s lab used an approach called “positional cloning” in its attempt to figure out precisely where in the mouse’s genome the LPS receptor lay hidden. This meant taking plate after plate of cell samples, activating them with bacterial LPS and determining the levels of inflammatory response in those cells by means of fluorescence, then cross-checking these results against known phenotypes and running the combined results through computer software to sort out likely locations for the hidden gene.

“You are looking for a calculated probability,” says Poltorak. “We were able to say, ‘This place has the best association with the phenotype, so the gene is likely to be found there.’ ”

Beutler and Poltorak logged 14-hour days—and often more than that—for six years. In retrospect, both men agree the daily grind was intense. “We were looking for one nucleotide change out of many millions,” Beutler explains. “If you miss it, you miss it. You don’t get a second chance. Beyond our immediate goal, we were trying to assemble a map of an area [in the mouse genome]. Mistakes could be fatal to the process, so that exerted a great deal of pressure.”

Competitive pressure added more weight. Poltorak notes that Beutler’s team was aware of two other laboratories pursuing work closely linked to their own—everybody steaming forward as fast as they could with the understanding that whoever cracked the riddle first would garner the world’s glory.

How far along the trail the competitors stood, whether miles ahead or a shade behind, was impossible to know for sure, despite periodic (and of course unreliable) reports from the other camps. “It’s like poker,” says Poltorak about the ongoing scientific rivalry. “You never really show your cards.”

The struggle took its toll. Poltorak remembers one morning during this time when he drove to the lab ready to plunge into the day’s work and couldn’t get out of his car. He was simply overwhelmed—intellectually, emotionally—and slumped near total collapse at the wheel.

“That was one of the worst moments,” he says now with a tight shake of his head, seated in his spotless lab on the fifth floor of the Jaharis Center. “I thought: What am I doing? I’ve been cloning this stupid gene in my sleep for years.”

“Obsession Is a Very Good Thing”

But there was steel within the man. Poltorak had an unlikely but potent model in mind, he admits, for how to proceed under extreme duress: the 1984 sci-fi movie Terminator, starring Arnold Schwarzenegger as a cyborg assassin sent back in time by the forces of Skynet to kill future human resistance leader Sarah Connor.

The Terminator battles through one brutal assault after another, including pipe bombs and exploding tank trucks full of gasoline, to nearly succeed at his goal. (The hydraulic press gets him in the end.) Poltorak wanted to be just as unstoppable in his own way as the Terminator had been in his. “I felt like I was on a mission,” he shrugs.

“We were obsessed, me and Bruce,” Poltorak continues in his light Russian accent, “determined to do or die. And obsession is a very good thing to have in medicine. We need to have crazy people—that’s crazy in a good way—to do the work.” He hopes his recent success will carry a tonic message for the current batch of doctoral students and post-docs whom he supervises in his lab. As he puts it, “Hard work can pay off, but you must do the work.”

The crowning achievement of those laser-focused years came on Dec. 11, 1998, when Science published an article heralding the Beutler lab findings: “Defective LPS Signaling in C3H/HeJ and C57BL/10ScCr Mice: Mutations in Tlr4 Gene.” Alexander Poltorak was the lead author, with Bruce Beutler appearing last among 14 names. (Irina Smirnova is listed third.)

“It’s a point of honor, and he deserved that,” Beutler says of Poltorak’s pre-eminence. “It’s true that I was the PI and the director of the lab, but he explored 2.6 million bases of DNA and did so relentlessly until we found the mutation.” A landmark event, the paper drew some 4,500 citations in ensuing years and set the stage for the Nobel Prize.

Poltorak notes during our interview that he did get his share of the limelight: he went to Stockholm in December for the prize ceremony. And then, unexpectedly, he delivers the quick universal gesture that says it all: two thumbs up.

This story first appeared in the Winter 2012 issue of Tufts Medicine magazine.

Bruce Morgan can be reached at bruce.morgan@tufts.edu.

A note regarding the above references to the Sackler School: In December 2019, Tufts University announced that it would remove the Sackler name from all programs and facilities—including the Sackler School of Graduate Biomedical Sciences. At that time, the school was renamed the Graduate School of Biomedical Sciences.