High-throughput sequencing technologies uncover a loss-of-function variant of the GATAB2B gene in a GAND patient
Clinical Genetics and Therapeutics
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Primary Categories:
- Clinical- Pediatric
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Secondary Categories:
- Clinical- Pediatric
Introduction
GATAB2B-associated neurodevelopmental disorder (GAND) is an autosomal dominant condition leading to characteristic facial features, global developmental delay with severe speech delay, intellectual disability and macrocephaly, caused by loss-of-function (LOF) variants in the GATAB2B. We report a case of GAND due to a heterozygous de novo duplication in GATAB2B, which long-read DNA and RNA sequencing proved a LOF effect due to transcriptional read-through of a partial second copy of the gene.
Case Presentation
The patient (RD0273) and parents were enrolled as part of an Institutional Review Board (IRB) approved study (IRB:11-00215: Rare Diseases) within the Steve and Cindy Rasmussen Institute for Genomic Medicine at Nationwide Children's Hospital. Informed consent was provided for all study participants.
The patient was born at term without complication. He was first noted with motor delay and hypotonia at 5-6 months, when he was not sitting and was not reaching for objects. At one year old, he did not crawl, or stand, or have any words. Brain MRI showed evidence of dysmyelination and type 1 Chiari malformation. At 3 years old, he pulled to stand. Swallow dysfunction and relative macrocephaly (88%ile for age) were noted.
Diagnostic Workup
Clinical exome sequencing, mitochondrial DNA sequencing, 11q11.2 (Prader-Will, Angelman critical region) methylation, and biochemical work-up - CDG, VLCFA, urine purine/pyrimidine - were undiagnostic.
Short-read Genome Sequencing (GS)
GS was performed on genomic DNA from the family using an Illumina NovaSeq6000 instrument according to manufacturer protocols. Reads were mapped to the GRCh37 reference sequence and secondary data analysis was performed using Churchill. VarScan v2.4.3 analyzed Copy number variant and runs of homozygosity.
RNA sequencing
RNA sequencing was performed using a NovaSeq6000 instrument (2 × 150 bp) according to manufacturer protocols.
Long Read GS
Genomic DNA from the proband was sheared using the Megaruptor 3 (Diagenode, Liege, Belgium) according to the manufacturer's protocol. PacBio high fidelity (HiFi) SMRTbell libraries were generated using the PacBio HiFi Express Template Prep Kit 3.0 protocol.
Short-read GS of the trio did not uncover any compelling SNV/indel variants, but VarScan 2 revealed a ~97 kbp duplication of exons 2-11 of the GATAD2B gene.
PacBio long-read sequencing of proband DNA (HiFi) and RNA (IsoSeq) showed that the tandem duplication occurs in the 3’ UTR of GATAD2B, causing a transcriptional read-through event. RNA sequencing showed that GATAD2B expression is in the normal range, but exhibits skewed (1024x coverage, 879 reads; 85.8%) allele frequency, consistent with haploinsufficiency caused by nonsense-mediated decay.
Treatment and Management
At the age of 5 years, child was diagnosed with Autism Spectrum Disorder. At age 6 years, he is still not interested in toilet training, has ongoing sleep issues, and is essentially non-verbal. The child is currently in kindergarten and receives physical/occupational/speech therapies.
Outcome and Follow-Up
Discussion of the result with the family is planned. If the family desires, the proband with be referred to the GAND support group and registry.
Discussion
NuRD regulates gene expression in many tissues and plays a role in genomic integrity, pluripotency, and neurodevelopment. To date, NuRD subunits GATAB2A, GATAB2B, CHD3 and CHD4, have been shown to be associated with overgrowth intellectual disability syndrome termed NuRDopathies. The phenotype of our case is consistent with GAND. GATAB2B is proposed as a causal gene for the symptoms of 1q21.3 microdeletion syndrome, in which a recurrent deletion results from non-homologous end-joining.
Conclusion
Our case illustrates how RNA-sequencing and long-read sequencing can prove the pathogenic hapoinsufficiency of the partial GATAD2B duplication causing GAND. This represents the first report of GAND caused by a de novo duplication. Specific diagnosis facilitated specific management and potential future treatment.
GATAB2B-associated neurodevelopmental disorder (GAND) is an autosomal dominant condition leading to characteristic facial features, global developmental delay with severe speech delay, intellectual disability and macrocephaly, caused by loss-of-function (LOF) variants in the GATAB2B. We report a case of GAND due to a heterozygous de novo duplication in GATAB2B, which long-read DNA and RNA sequencing proved a LOF effect due to transcriptional read-through of a partial second copy of the gene.
Case Presentation
The patient (RD0273) and parents were enrolled as part of an Institutional Review Board (IRB) approved study (IRB:11-00215: Rare Diseases) within the Steve and Cindy Rasmussen Institute for Genomic Medicine at Nationwide Children's Hospital. Informed consent was provided for all study participants.
The patient was born at term without complication. He was first noted with motor delay and hypotonia at 5-6 months, when he was not sitting and was not reaching for objects. At one year old, he did not crawl, or stand, or have any words. Brain MRI showed evidence of dysmyelination and type 1 Chiari malformation. At 3 years old, he pulled to stand. Swallow dysfunction and relative macrocephaly (88%ile for age) were noted.
Diagnostic Workup
Clinical exome sequencing, mitochondrial DNA sequencing, 11q11.2 (Prader-Will, Angelman critical region) methylation, and biochemical work-up - CDG, VLCFA, urine purine/pyrimidine - were undiagnostic.
Short-read Genome Sequencing (GS)
GS was performed on genomic DNA from the family using an Illumina NovaSeq6000 instrument according to manufacturer protocols. Reads were mapped to the GRCh37 reference sequence and secondary data analysis was performed using Churchill. VarScan v2.4.3 analyzed Copy number variant and runs of homozygosity.
RNA sequencing
RNA sequencing was performed using a NovaSeq6000 instrument (2 × 150 bp) according to manufacturer protocols.
Long Read GS
Genomic DNA from the proband was sheared using the Megaruptor 3 (Diagenode, Liege, Belgium) according to the manufacturer's protocol. PacBio high fidelity (HiFi) SMRTbell libraries were generated using the PacBio HiFi Express Template Prep Kit 3.0 protocol.
Short-read GS of the trio did not uncover any compelling SNV/indel variants, but VarScan 2 revealed a ~97 kbp duplication of exons 2-11 of the GATAD2B gene.
PacBio long-read sequencing of proband DNA (HiFi) and RNA (IsoSeq) showed that the tandem duplication occurs in the 3’ UTR of GATAD2B, causing a transcriptional read-through event. RNA sequencing showed that GATAD2B expression is in the normal range, but exhibits skewed (1024x coverage, 879 reads; 85.8%) allele frequency, consistent with haploinsufficiency caused by nonsense-mediated decay.
Treatment and Management
At the age of 5 years, child was diagnosed with Autism Spectrum Disorder. At age 6 years, he is still not interested in toilet training, has ongoing sleep issues, and is essentially non-verbal. The child is currently in kindergarten and receives physical/occupational/speech therapies.
Outcome and Follow-Up
Discussion of the result with the family is planned. If the family desires, the proband with be referred to the GAND support group and registry.
Discussion
NuRD regulates gene expression in many tissues and plays a role in genomic integrity, pluripotency, and neurodevelopment. To date, NuRD subunits GATAB2A, GATAB2B, CHD3 and CHD4, have been shown to be associated with overgrowth intellectual disability syndrome termed NuRDopathies. The phenotype of our case is consistent with GAND. GATAB2B is proposed as a causal gene for the symptoms of 1q21.3 microdeletion syndrome, in which a recurrent deletion results from non-homologous end-joining.
Conclusion
Our case illustrates how RNA-sequencing and long-read sequencing can prove the pathogenic hapoinsufficiency of the partial GATAD2B duplication causing GAND. This represents the first report of GAND caused by a de novo duplication. Specific diagnosis facilitated specific management and potential future treatment.