A new computational tool developed by Pacific Biosciences can detect genetic mutations that cause spinal muscular atrophy (SMA) with high accuracy and could help identify “hidden carriers” that are not being detected by current tests, a new study reports.
The study “ Comprehensive SMN 1 and SMN 2 profiling for spinal muscular atrophy analysis using long-read PacBio HiFi sequencing ,” was published in the American Journal of Human Genetics .
SMA is caused by mutations in the SMN1 gene , which provides instructions for making the SMN protein. The second gene, called SMN2 , has a nearly identical sequence to SMN1 , but due to a change in one nucleotide (one of the building blocks of DNA), it produces much less SMN protein – about 10% of the total.
Both SMN1 and SMN2 are on chromosome 5 and most people inherit two copies of each, one from each parent. However, a person can inherit additional copies of both genes. Extra copies of SMN2 are usually associated with a less severe course of the disease, since they can partially compensate for the mutated SMN1 ..
But extra copies of SMN1 can complicate genetic testing of SMA due to so-called “hidden carriers” – people with two healthy copies of SMN1 on one copy of chromosome 5, but no working versions of the gene on the other copy of the chromosome.
Technically, these individuals produce as much SMN protein as a healthy individual without any SMN1 mutations . But since a copy of chromosome 5 without any functional SMN1 can be passed on to their children, they are considered carriers of SMA.
Current genetic tests cannot accurately identify these latent carriers because they will only show two healthy copies of SMN1 .without differentiation in the arrangement of chromosomes. Covert carriers are particularly common among Africans, accounting for over a quarter of African SMA carriers.
The only way to identify hidden carriers is to analyze the pedigree, the study of patterns in the family tree. A particular genetic variation called g.27134T>G has been proposed as a marker for two copies of SMN1 on the same chromosome, but it is not very sensitive as it is also often present in people with one copy of SMN1 on each chromosome.
Search for “hidden carriers” of SMA
To better identify hidden carriers, a research team led by scientists from Pacific Biosciences has developed a computational tool called Paraphase to analyze SMN1 and SMN2 in more detail using genetic sequencing data. In these datasets, a person’s genetic code is represented in small chunks known as “reads”. The algorithm aligns reads with either SMN1 or SMN2
sequence given specific data to determine the difference between the two genes and identify SMA-causing mutations in SMN1. The program also predicts the number of copies of these genes based on counts of the number of reads corresponding to each sequence.
The researchers analyzed the genetic data of 438 people from five ethnic groups. Most were of European descent, but there were also data from mixed groups of Americans, Africans, South and East Asians.
Paraphase can identify, with over 99% accuracy, the copy number of SMN1 and SMN2 for each gene . In addition, all SMA patients and carriers were correctly identified by the algorithm, with no false negatives or false positives.
Combining haplotype, pedigree data
Since the SMN1 and SMN2 gene sequences are determined , the algorithm can also identify their haplotypes, a set of closely related DNA regions that tend to be inherited together.
Haplotype data were combined with pedigree data to search for markers of two copies of SMN1 on the same chromosome, with particular attention to African populations where this variation is most common.
The results showed that a certain combination of haplotypes, called S1-8+S1-9d, was found on two-thirds (21 of 31) of chromosomes with two copies of SMN1 in African patients. These haplotypes were rarely present when there was only one copy of SMN1. “We identified a common 2-copy SMN1 allele that accounts for 67.7% of 2-copy SMN1 alleles in Africans,” the researchers wrote.
Given the known prevalence of chromosomes without any functional SMN1, this means that the probability that an African carrying the S1-8+S1-9d haplotype is a concealed carrier is about 88%.
The haplotype markedly outperformed the g.27134T>G marker in identifying double copies of SMN1, since individuals with this marker have only about a 3% risk of being latent carriers.
“Based on our limited sample of 87 African alleles, we estimate that a positive test for these two haplotypes in an individual with two copies of SMN1 gives an occult carrier risk of 88.5%, significantly higher than the previously found marker [variant] g. .27134T>G,” the researchers said, noting that their study is limited to a relatively small number of samples, especially for non-European populations, and that more research is needed to confirm these results.
“For many years, researchers have known that SMN1 and SMN2 play a role in the occurrence and severity of SMA, but have not been able to accurately answer why and how people develop more or less severe forms of the disease,” writes Pacific Biosciences .
“With Paraphase, it is now possible to answer some of these long-standing questions and provide more accurate carrier screening.”