Trinucleotide repeat expansion disorders
The mechanism involved in triplet repeat expansion and related disorders
The Huntington’s disease and fragile X syndrome are occurred due to the imbalance in trinucleotide repeats and we discussed both diseases in brief.
I researched the prevalence of both types of disease and I thought that it is important to cover the trinucleotide repeat expansion disorders or trinucleotide repeat expansion in detail.
During my experience in the research lab, I was worked on fragile X and Huntington’s disease so I thought that this topic is very important to explore.
In this article, we will discuss the trinucleotide repeat expansion mechanism and how trinucleotide repeats involved in disease.
Let us start with some basics,
Nitrogenous bases are the major ingredients of DNA. Remember a nucleotide is not a base. The nucleotide is made up of three important components: nitrogenous base, sugar and a phosphate.
For more detail read the article: Story of DNA
The junk DNA or non-coding DNA do not play any significant role in protein formation but it has a definite role in the regulation of gene expression.
This non-coding region of the genome is made up of repeats and located on the telomeric and centromeric region as microsatellite or minisatellites. However, some coding gene segments also contain repeat regions.
The repeats are used as a molecular marker in genetic studies ( for more detail on genetic markers read the article: Genetic markers). For example, the VNTR markers are 10 – 100 nucleotide long sequences having 5- 50 repeats and STRs contains 1-6 nucleotide long sequences with 50 repeats.
The non-coding region remains inactive as these sequences are methylated and cannot be recognized during gene expression.
For more detail on methylation read the article: Epigenetic class 1: What is epigenetic?
However, functionally it helps in the regulation of gene expression and it is very important to maintain the appropriate methylation pattern for the expression of a particular gene.
Hence the normal protein production depends on the proper function of coding and non-coding DNA.
In short, we can conclude that a mutation in a coding region inflict changes of protein sequence and function of protein whereas mutation in the non-coding region results in a change in the expression of that protein-coding gene.
Trinucleotide repeat expansion mechanism is not clearly explained as it occurs in both coding regions of a gene (for example Huntington’s disease) and non-coding regions (for example fragile X syndrome) of a gene.
Fore more detail on epigenetic alteration read the article: Epigenetic class 1: What is epigenetic?
Trinucleotide repeat expansion mechanism
Two of the most common mechanisms are scientifically accepted:
- Error in transcriptional mediated DNA repair pathway
- Strand slipping during DNA replication (called as DNA slippage)
During the transcriptional mediated DNA repair, under the regulation of some genes, the DNA repair mechanism repairs all the unmatched bases. However, the repeated sequences confuse the machinery and give a false signal to activate the mechanism for repairing mispaired bases.
The main function of the DNA repair mechanism during the transcription is to repairing the mismatch. Hence it can not recognize the repeat sequences or confused with it.
It leads to continuous expansion of that repeat.
Transcriptional mediated DNA repair mechanism involved in triplet repeat expansion is most prevalent in non-dividing cells because if it is occurred in dividing cells, the abnormal repeat numbers are recognized during cell division.
Petruska et.al.,1997 proposed a triplet repeat expansion mechanism with DNA slippage.
Strand slipping/ DNA slippage/ DNA strand slippage occurred during DNA replication. By their experiments on synthetic DNA, they concluded that “DNA looping promotes slipping of repeated DNA sequences during replications”.
For more detail on DNA replication read the article: DNA Replication class 1: General process of DNA replication.
During the strand separation stage of DNA replication, a loop is created at the site of repeat sequences. Therefore the repeats in the loop are not copied during replication.
If looping takes place in parental DNA strand than the loop is skipped from replication and results in decreased copy number of the repeated sequence.
But if the newly replicating DNA loops out, the repeat number is increased twice because the sequences in the loop are replicated twice in newly synthesized DNA.
Error in triplet repeat expansion during replication may occur in both somatic cells as well as germ cells. Hence the abnormally replicated repeats are inherited to the next generation.
Scientifically, the strand slipping mechanism is more appropriately defines the triplet repeat expansion mechanism than Error in the transcriptional mediated DNA repair pathway.
Genetic anticipation is most common in trinucleotide repeat expansion disorders. The severity of trinucleotide expansion disease is increased at the earlier onset of disease with each successive generations, is called as genetic anticipation of disease.
As the repeat number may cross the threshold length, it may cause the disease, however, the threshold length is different for both coding and non-coding region.
The permutational condition for TNR in the coding region is 29 to 35 in contrast, for a non-coding region is 55 to 200 (in general).
Huntington disease is an autosomal dominant disorder caused by the Huntingtin protein. It is a polyglutamine disease. The polyglutamine repeats are commonly observed within the gene. The gene is located on chromosome number 4.
For more detail on autosomal inheritance read the article: different type of inheritance.
It affects 1 in 10,000 individuals. The Huntington’s disease is associated with the motor, cognitive and behavioural symptoms hence it is called as a neurodegenerative disorder.
Common sign and symptoms are:
- Difficulty in organizing and focusing on a task
- Lacks on impulse control, acting without thinking
- Lack of awareness of one’s own behaviour
- Lack of behavioural flexibility
- Difficult in learning
- Feeling sadness and apathy
- Socially inactive
- Rigid muscle contraction
- Involuntary jerky movement
- Slow or abnormal eye movements
- Difficulty in swallowing and talking
- Suicidal thoughts
- Fatigue or loss of energy
Htt protein is naturally found in the neurone of the brain. The severity of the disease depends on the polyglutamine repeat numbers. Based on the diagnostic criteria, the CAG repeats are categorized into 4 classes:
• <26 triplet repeats of CAG- normal
• 27 to 35 triplet repeats of CAG – intermedia
• 36 to 39 triplet repeats of CAG – penetrance
• Above 40 triplet repeats of CAG- affected
Individuals with more than ~40 repeats of CAG results in Huntington’s disease. As the disease is further passed to the next generation in the early onset, the severity of disease will increase (called as Huntington’s anticipation).
Notably, the polyglutamine repeat numbers occur within the coding region of the HTT gene.
An interesting research stated that the severity of the disease further depends on the parental inheritance.
If the mutant allele is inherited from the father, the risk of earlier onset is higher as compared to maternal inheritance. The chance of CAG repeats expanding though sperm is higher because the mutant allele is more unstable if it is transmitted through sperm.
There are three possible ways proposed by which the Htt protein causes the Huntington’s disease,
Sadri-Vakili and Cha, 2006, reported that mutant Htt proteins involve in alteration of transcription by Histone modification. Histone helps in the packaging of DNA. If it is loosely packed, it promotes the transcription.
When the mutant HTT gene (having abnormal CAG- polyglutamine triplet repeats) interacts with histones, it makes the histone- chromatin structure more tightly packed which resist the transcription.
The HTT gene is not properly transcribed and causes neurodegenerative disease.
Verma H, et.al., 2007 reported that Htt protein interacts with HAP-1 intercellular protein and results in earlier onset of disease.
Normally Htt protein is work independently in the brain. The protein is involved in most of the brain functions like cognition, movement and understanding.
In contrast, HAP-1 protein is involved in organelles transport and trafficking (like mitochondria and endoplasmic reticulum), when the mutant form of Htt protein binds with the HAP-1, it alters the function of the HAP-1 protein.
It leads to insufficient energy production in the brain, results in the neurodegenerative spectrum of disease ( because of lack of energy the brain cells can not function properly).
Another interesting Huntington’s disease mechanism is explained by Cummings and Zoghbi in 2000. They stated that the mutant Htt protein interferes with protein folding. The misfolding of protein which results in abnormal function of Htt protein in brain cells. The mutant misfolded protein now interacts with the wrong receptors and aggregates.
Aggregation affects the normal function of the cell and its structure. However, all three mechanisms are not fully studied.
Fragile X syndrome
The fragile X CGG repeats are located on the 5’-P untranslated region which means in the non-coding region. It is caused by the FMR1 gene located on chromosome X.
FXS is X linked dominant disorder which is associated majorly with mental retardation. It is caused by the change in the expression of the FMR1 gene because it occurs in the non-coding region of a gene.
Sings and symptoms of FXS:
- Mentally retarded
- Intellectual disability from mild to severe
- Speech delay, sensory integration problems
- Attention deficit
- Flat feet, long prominent ears
- Large face
- Low muscle tone
- Larger testis in male
- Hyper-extensible joints
The FXS disease spectrum in the female is different from the male. It is X linked dominant, 50% of males may affect by FXS. The disease is more severe in males. It affects 1 in 1,250 males and 1 in 2,500 females.
Female contains two X chromosome one active and another inactive if the mutant copy of FMR-1 gene is present on inactive X chromosome than she will remain normal. However, the severity of the disease varies from female to female.
Additionally, the intellectual disability is also less in females as compared to male. however, some affected females have severe symptoms as compared to other females because of the length of the CGG repeat. The more the mutant CGG repeats, the more severe FXS.
The range of CGG repeats in fragile X syndrome is as listed:
• 1-40 range of CGG repeats is a normal state
• 40- 200 pre-mutation
• More than 200 CGG repeats disease state
The incomplete penetrance is one of the possibility when considering premutation. The pre-mutation is a condition in which an individual is most probably passes the mutant allele to their child.
However, if the pre-mutant allele passes from female to their child, the chance of Fragile X syndrome is higher as compared to male.
The variation emerges due to the difference in the process of egg and sperm formation. Even some individuals can have a mutant allele but do not shows any symptoms of the disease.
Timchenko and Caskey, 1996 reported two mechanisms of FXS inheritance.
Transcriptional silencing through methylation, abnormal repeats of CGG results in the change in methylation pattern of the gene FMR-1. The gene becomes transcriptionally inactive due to hypermethylation.
In another mechanism, the translation initiation is decreased. The non-coding regions of DNA contain sequences which can be recognized by some of the enzymes and proteins involved in translation.
If the non-coding region is abnormally repeated the enzyme cannot recognize the substrate sequence and the initiation of translation is decreased.
Either mechanism is most probably involved in mutant FMRP. FMRP is fragile X mental retardation protein which is encoded by the FMR-1 gene. It is highly expressed in neurones of brain, testis and ovaries.
List of triplet repeat expansion disorders in coding regions:
HTT exon 1
ATN1 exon 5
Oculopharyngeal muscular dystrophy
PABPN1 exon 1
More than 11
Spinocerebral ataxia 1
ATXN1 exon 1
ATXN2 exon 1
ATXN3 exon 8
CACNA1A exon 47
ATXN7 exon 3
TBP exon 3
List of trinucleotide repeat expansion disorders in non-coding regions:
Fragile X syndrome
FMR1, 5’ UTR
Myotonic dystrophy 1
DMPK, 3’ UTR
More than 50
AFF2, 5’ UTR
More than 200
FXN, intron 1
ATXN80S, 3’ UTR
Each trinucleotide repeat expansion disorders has a different mechanism of inheritance. FXS results from the loss of function or reduced expression of the FMR1 gene while HD results from the change in function of the HTT gene.
No clear universal mechanism for all types of trinucleotide repeat expansion disorder is postulated yet.
Article written and reviewed by: Tushar Chauhan