August 13, 1999
Single Switch Triggers Two Immune System Genes
Two genes critical to the immune system's
adaptability in battling viruses, bacteria and other invaders receive on and
off signals from a single DNA segment, HHMI researchers have found. The
discovery explains how the two genes work in concert and hints at how the
genes have managed to remain partners for more than 450 million years since
they first appeared in cartilagenous fish as part of an adaptive immune
system.
In an article in the August 13, 1999, issue of the journal
Science
, HHMI investigator
Michel
Nussenzweig
and colleagues at The Rockefeller University reported that
neighboring genes, called
RAG1
and
RAG2
, are switched on in
concert by a single genetic control signal nestled near
RAG2
on the
chromosome. The discovery is like finding that a light switch in one house
also controls lights in a house across the street.
“The big question is how do you regulate something that lands in the genome in such a way that both of these gene products, which are required for the transposase activity, in fact are coordinately regulated.”
Michel C. Nussenzweig
The
RAG1
and
RAG2
genes produce proteins that somehow join
to form a "transposase," an enzyme that helps snip apart and rearrange genes
that code for two critical weapons in the immune system's arsenal. These
weapons are protein antibodies produced by the immune system's B cells and
receptors found on the surface of T cells.
Antibodies are molecules that roam the bloodstream, recognizing and
attaching to invaders and marking them for destruction. Similarly, the
receptors on T cellsthe immune system's infectionfighting "soldiers"are those that recognize antigens, fragments of foreign proteins, so thatthe T cells can attack the viral or bacterial proteins.
The RAG1/RAG2 recombinases' ability to rearrange the genes for such
antibodies and receptors is crucial to the immune system's flexibility in
generating a nearly infinite variety of weapons against infections and
malignant cells. HHMI investigator
David Schatz
discovered RAG1 and RAG2 in
1989, and in 1998 with colleagues at Yale University, he showed that RAG1 and
RAG2 had transposase activity.
"Before our studies, we knew that the two genes were controlled
coordinately," said Nussenzweig. "However, we knew very little about their
control elements." He added that previous experiments testing small segments
of DNA were inconclusive in defining the
RAG1/RAG2
control mechanism.
In the current work, however, Nussenzweig and his colleagues used
specially modified bacterial artificial chromosomes (BACs) to carry large
pieces of DNA into mouse cells. HHMI investigator
Nathaniel Heintz
, also at
Rockefeller, invented the BAC modification technique.
Using BAC to shuttle DNA into the mice, the researchers created
transgenic mice that possessed fluorescently-tagged
RAG
genes and
extended DNA segments adjacent to the
RAG
genes that the researchers
suspected carried the
RAG
controller. In addition, Nussenzweig's team
used the BACs to introduce genes coding for two fluorescent proteins-one
that would shine green wherever RAG1 appeared, the other yellow in cells
expressing RAG2.
By inserting such test segments into mice and looking for fluorescent B
and T cells, the scientists could determine whether those segments contained
a control sequence that properly switched on the RAG genes.
"We used the marker genes to show us where the RAGs were being expressed,
and then we used recombination technology to knock off pieces of the
bacterial artificial chromosomes to reveal where the control elements might
be," explained Nussenzweig.
After many such experiments with various constructed segments, the
scientists narrowed down the control sequence to a small piece of DNA next
to one end of the
RAG2
gene.
The discovery that the neighboring
RAG
genes are controlled by a
single switch has important implications for understanding how the two
genes evolved as part of the immune system, said Nussenzweig.
"The configuration of these genes makes scientists believe that they were
inserted into the genome together, as a 'transposable element,'" he said.
What's more, said Nussenzweig, the finding may explain why, over the 450
million years since the genes first appeared, evolution has required them to
remain closely spaced in the genome.
"The big question is how do you regulate something that lands in the
genome in such a way that both of these gene products, which are required
for the transposase activity, in fact are coordinately regulated?"
The key to the genes' long-lived marriage might lie in the mechanism by
which the control sequence activates both genes at once, said Nussenzweig.
In the
Science
paper, he and his colleagues propose two
possibilities: First, the control element may bounce back and forth between
the promoter regions of the two genes and activate them; alternatively, the
controller may touch both simultaneously.
Although Nussenzweig emphasizes that there are no data that reveal which
is the correct mechanism, the theory of simultaneous activation might better
explain why evolution has refused to allow the two genes to separate and
still remain functional partners in the transposase enzyme. He and his
colleagues plan experiments that they hope will reveal the control mechanism
for RAG1 and RAG2.
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