- Fragile X
- Treatment & Intervention
- Support the NFXF
Most of us have heard of CGG repeats, those patterns of DNA molecules that are counted when testing for Fragile X (FMR1) mutations. (For background, see the Q & A with Dr. Karen Usdin on page 16.) We then hear a number like 400 for full mutations or 80 for premutations and that’s it, nothing else matters but that magic number. We envision a long tract of CGGCGGCGGCGG over and over again, as the genetic counselor or doctor described. But lately you may also have begun hearing about “AGG interruptions,” which in some way are related to Fragile X inheritance. This article will offer an overview of their importance.
To review some genetics, DNA is a strand of chemicals, called nucleotides. The nucleotides that make up DNA are Adenine, Thymine, Cytosine and Guanine (abbreviated as A, T, C and G). So a CGG repeat is a triplet of cytosine, guanine and guanine. The pattern of nucleotides (like ATCGATCG, etc.) makes up a gene that instructs the cell on how to make or regulate proteins.
In the 1990s, after the discovery of the FMR1 gene and the CGG repeat expansion that causes Fragile X syndrome, Fragile X experts began talking about a different DNA pattern called an “AGG interruption,” which occurs between about every nine or 10 CGGs. In the normal Fragile X gene (five to 45 repeats), you might have a 30-CGG-repeat pattern that has 10 CGGs, one AGG, then 10 CGGs, then one AGG, then another 10 CGGs. These AGG interruptions act as a sort of anchor, keeping the 30-repeat FMR1 gene stable, like fence posts every 10 feet in a long fence.
As DNA samples were being studied by Fragile X researchers, they found that while some individuals had AGG interruptions every nine or 10 CGG repeats, some lost them as the repeats got bigger. So a person might have 10 CGGs then one AGG, 10 CGGs then one AGG, then 40 CGGs in an individual who has 60 CGG repeats.
This led to two questions regarding the presence or pattern of AGG interruptions: 1) Do the number or placement of AGG interruptions affect the stability of the FMR1 gene?
2) “Why do some premutations expand and some not?”
A recent study by Dr. Sarah Nolin at Institute of Basic Research in New York, Dr. Elizabeth Berry-Kravis at Rush Medical Center in Chicago and Asuragen Inc. in
Austin, Texas was presented in a poster at the annual meeting of the American College on Medical Genetics earlier this year. The purpose of the study was to see how AGG interruptions affect the stability of the premutation, and also to develop (and use) the technology that would identify these AGG interruptions in Fragile X testing.
The study found that if there were 33 or fewer CGGs in a row beyond the last AGG, then the premutation usually didn’t expand. But when there were 39 or more CGGs in a row after the last AGG, without any AGG interruptions, then the premutation was usually unstable (expanding by one or more CGG repeats). This helps us understand why some premutations with, for example, 62 CGG repeats, appear stable over generations, whereas some with the same number of 62 repeats expand over subsequent generations. It’s because the one with a long tract (>39 repeats) of CGG repeats without an
AGG “anchor” is more likely to expand than one with properly interspersed AGG interruptions.
The technology that was developed to identify the presence and placement of the AGG interruptions is new and exciting. Asuragen, in cooperation with the UC Davis M.I.N.D.
Institute, has developed a unique type of PC test (see below) called “Amplidex.” It can identify the AGG interruptions in both males and females and in both normal and expanded FMR1 genes.
For scientific reasons, labs have had a hard time finding AGG interruptions, especially in women. This new technology overcomes these hurdles. Variations on a genetic testing method called polymerase chain reaction, or PCR, reveal the number and location of AGG interruptions. Three different PCR studies done together show very different patterns of the CGG repeat region when AGG interruptions are present. Specially trained experts in a laboratory can review all three studies to solve the problem of where the AGG interruptions are located; the method is similar to solving a brainteaser puzzle.
This test is currently available only for research studies. Eventually it may be available for clinical use. So it is possible that in addition to the “magic number” of CGG repeats, the presence and placement of AGG interruptions will inform your risk for expansion of a premutation.
Why it Matters
There are a number of reasons why this issue of AGG interruptions is important.
First, if you or a family member have a premutation that hasn’t expanded to a full mutation, this technology may give you more accurate risk figures for that premutation to expand.
Second, it is possible that the number of AGG interruptions may contribute to or affect the risk for FXPOI or FXTAS. The risk for FXPOI is about 20–25 percent in female premutation carriers; the risk for FXTAS is about 30–40 percent in male and 5–8 percent in female carriers. At this point we don’t have good “predictors” of who may develop these two conditions. It is possible that this technology identifying AGG interruptions may lead us to better predict the group at highest (and lowest) risk for these Fragile X-associated Disorders.
Third, it is possible that our knowledge of AGG interruptions might lead to better understanding of the small subset of children with premutations who have developmental disorders like autism, ADHD, etc. We know this is clearly a small group, since most premutation carriers exhibit normal development and cognitive function. However, there is a small subset of (mostly male) children in the premutation range who have developmental/behavioral disorders, and this new technology might give us some clues as to why they are affected, even though most carriers are not.
All of which leads to the possibility that you may soon hear, “I have 65 repeats with three AGG interruptions, ya-hoo!”