Fred Alt has built a career making sense of the immune
system—specifically, the diverse antibodies that fight off
invading molecules, from viruses to cancer cells to pollen.

Photograph by Jared Leeds

The Twists and Turns of Immunity

The diversity of the immune system’s antibodies depends on the three-dimensional structure of DNA.

Imagine that the only road connecting two cities had rollercoaster-inspired loops. Cars and trucks attempting to make the journey would plummet off the highway. It would surely discourage direct traffic between the towns.

In the cell, where molecular machines constantly travel along strands of genetic material, such uninviting loops and curls are commonplace. They help keep proteins from accessing genes the cell doesn’t need at the time. Later, when the cell needs the genes to be activated, the dizzying roundabouts can be straightened out to support molecular traffic.

In the late 1970s, when HHMI investigator Fred Alt at Children’s Hospital Boston started studying the human immune system, he had no idea this higher-order structure of DNA even existed. Today, he’s found that the way DNA is packaged into three-dimensional structures is crucial to every immune process he’s looked at.

Alt studies how the immune system generates antibodies against any potential invader to the body—from a virus to a cancer cell or bit of pollen. When an antibody recognizes an invader, it signals the rest of the immune system to destroy it. This means millions of different antibodies are somehow encoded in the DNA of immune cells to recognize all types of invading molecules.

“It’s important for a cell to make only one type of antibody at a time,” says Alt. In immunology, that quality is called specificity. “But it’s also important that different cells make different antibodies.” This is diversity.

If the immune system had enough genes for every possible antibody the body might need, our genomes would be exponentially larger than they are. Instead, immune cells combine different bits of genes in different ways to make millions of unique combinations. Three gene segments—dubbed variable (or V, for short), diversity (D), and joining (J)—are the basis for these permutations.

In 1984, Alt discovered that immune cells always combine a D and J segment before adding a V. Cells early in their development, he found, contained partial antibody genes composed of only Ds and Js. When Alt grew those cells in the lab, the antibody genes added a V later in the cell’s development. He has since worked out some of the proteins responsible for combining these gene segments in different ways. And he’s pinpointed how the cell makes sure only one antibody is produced—by suppressing alternate antibody genes once a combination is successfully produced.

But Alt wanted to know how the cell ensured the order in which the genes were combined. The hint came when he was looking at the DNA between D and V gene segments. He noticed a stretch of DNA that’s found elsewhere in the genome and is known to regulate DNA transcription. A protein called CTCF that attaches to certain DNA sequences to regulate gene expression can bind to the region. It folds the surrounding DNA into loops.

Learn about the process of copying DNA into messenger RNA (mRNA). Video: ©HHMI.

Alt wanted to know the role of the so-called CTCF-binding element in controlling the assembly of antibody genes. So he mutated the binding element, with drastic results.

“We saw a very unordered pattern of antibody gene generation,” says Alt. Segments from the V genes joined with Ds before the Js were tacked on. And only a few of the possible V segments were used, leading to fewer unique antibody possibilities. Moreover, cells lost the ability to ensure production of only one antibody at a time. Both specificity and diversity were obliterated. His group, led by postdoctoral fellow Chunguang Guo, published the observations September 11, 2011, in Nature.

“This site impacts every regulatory process that we’ve been studying for 30 years,” Alt says of the CTCF.

The CTCF protein is normally bound to the CTCF-binding site during development. Alt’s team discovered that the antibody gene forms a number of loops that require the bound protein. Transcription proteins—which travel along DNA and use it as a blueprint for RNA strands—get stuck between the D and V segments because of these loops, Alt thinks. The arrangement allows developing cells to produce antibodies with only the D and J regions; the V segment is added later.

“Genes have incredibly complex three-dimensional organization,” Alt says. “And we’re learning that all of biology depends on that organization.”

Alt’s next task is to uncover the other proteins responsible for controlling the CTCF site. Now that he’s found a master control site that keeps the V from being added to antibodies too early, he wants to know what signals the cell to finally add the V segment. Stay tuned.

Scientist Profile

Boston Children’s Hospital
Cancer Biology, Immunology

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