A BRAT1 synonymous mutation affecting exon splicing leads to lethal neonatal rigidity and multifocal seizure syndrome: A Chinese case report.

doi:10.21203/rs.3.rs-2229166/v1

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Case Report

A BRAT1 synonymous mutation affecting exon splicing leads to lethal neonatal rigidity and multifocal seizure syndrome: A Chinese case report.

https://doi.org/10.21203/rs.3.rs-2229166/v1

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Rigidity and Multifocal Seizure Syndrome, Lethal Neonatal (RMFSL, OMIM#614498), is caused by mutations of the BRAT1 gene. Our patient had the typical syndromes of RMFSL, and Trio whole exome sequencing (trio-WES) identified a homozygous synonymous variant (BRAT1:C.1395(exon10)G > C). Given that the pathogenicity of synonymous mutation (p.Thr465Thr) is most likely underestimated, a further transcriptional study of the father showed that C.1395(exon10)G > C mutation would result in abnormal splicing, which caused the exon 10 skipping and affected protein features. We first confirmed the pathogenicity of the synonymous mutation of BRAT1 using a transcriptional study.

Medical Genetics

Neurobiology of Disease

BRAT1

RMFSL

case report

synonymous mutation

exon splicing

RMFSL is an autosomal-recessive epileptic encephalopathy caused by variants of the BRAT1 gene [2, 5, 10]. Previous studies have demonstrated that BRAT1 maintains mitochondrial homeostasis and DNA damage response[2, 3, 9]. However, the underlying pathogenic mechanisms of these BRAT1 mutations are not well understood. In the past few years, several homozygous missense mutations and compound heterozygous mutations in BRAT1 have been identified among cases with RMFSL[2–5, 7, 8]. RMFSL is a fatal neurological disorder with episodic jerking in utero, lack of psychomotor development, early onset of rigidity, frequent multifocal epilepsy, and early death[7]. Here, we report a case with a severe phenotype of RMFSL and first confirm the pathogenicity of synonymous mutation of BRAT1 using a transcriptional study.

The patient was a full-term male infant with average weight, head circumference, and length. He had unexplained cyanosis and convulsions just after cesarean delivery. The baby had been in a coma and needed respiratory support, with cortical thumbs, hypertonia, limb myoclonic, and facial twitch. The most invasive operation could trigger muscle spasms. His heart rate and blood oxygen saturation occasionally drop to very low levels. We pumped midazolam continuously and gradually increased the dose to cease the seizures. The seizures were getting worse daily, exhibiting resistance to many antiepileptic drugs. The combination of midazolam (7ug/kg.min), phenobarbital (10mg/kg.d), topiramate (5mg/kg.d), and levetiracetam (20mg/kg.d) could not work later. Tandem mass of serum amino acid and urine organic acid analyses, TORCH test results were negative. Heart rate variability weakened in a dynamic electrocardiogram. Cranial magnetic resonance imaging (MRI) showed an insufficient volume of bilateral temporal and frontal lobes (Fig. 1A). His parents were not consanguineous in marriage. The baby's sister was healthy, while his brother died with the same symptoms as him. Unfortunately, despite all efforts, the patient died soon after the ventilator withdrawal in the neonatal period.

Trio-WES identified one homozygous mutation in the BRAT1 gene (c.1395 (exon10) G > C), which locates in the chr7:2580613. Further genetic analysis revealed that the unaffected parents of the patient were heterozygous carriers of the mutation of c.1395 (exon10) G > C, and the sister was wild-type (Fig. 1B). A synonymous mutation is easily ignored in the analysis of pathogenic variants. Considering the result was a synonymous mutation (p.Thr465Thr), we further conducted a transcriptional study to confirm the effect of sequence changes on RNA splicing. First, the mRNA was extracted from the father’s blood sample and reverse transcribed into cDNA. Then, Sanger sequencing sequenced the targeted regions (exon8-13). The results showed that the mutation caused exon 10 to skip (Fig. 1C). The gross deletion of exon 10 led to truncated proteins, which could influence the structure and function of the BRAT1 protein (Fig. 1D).

The BRAT1 gene locates on chromosome 7p22. The BRAT1 gene is thought to be involved in mitochondrial homeostasis and DNA damage response[1] and is essential in regulating mitochondrial function and cell proliferation[9]. The mutation of BRAT1 could result from two kinds of phenotypes that inherit an autosomal recessive. One is a neurodevelopmental disorder with cerebellar atrophy and with or without seizures (OMIM:618056); the other is rigidity and multifocal seizure syndrome, lethal neonatal(OMIM: 614498). The most severe RMFSL symptoms are refractory seizures, hypertonia, and early death. EEG can reveal bilateral temporal and central spike activity, multifocal seizures, background slowing, and absent posterior dominant rhythm[5]. Cardiopulmonary arrest and pneumonia are the most common causes of death.

Celik Y et al.[2] reported a Turkey female baby with the same symptoms as our case. She died from multiorgan failure at ten months old. The exon sequence detected a homozygous mutation (c.2230_2237dupAACATGC)[2]. Compound heterozygous mutation in BRAT1 has different clinical manifestations and severity. Two siblings from Japan with compound heterozygous mutations in BRAT1 had intractable seizures from the neonatal period and died of pneumonia at three months and one year old, respectively[8]. Four compound heterozygous BRAT1 mutation cases with mild or moderate symptoms extend the clinical spectrum of BRAT1-related diseases[10]. A 10-year-old girl with severe intellectual disability, rigidity, ataxia or dyspraxia, and cerebellar atrophy has a biallelic variant in the BRAT1, which was confirmed significantly decrease BRAT1 transcript levels and pathogenicity[6]. Table 1 lists some of the mutation information of the BRAT1 gene and corresponding clinical phenotypes.

We collected the reported BRAT1 mutations and found that many different mutations could cause RMFSL (Fig. 1E). Among them, frameshift mutation accounted for the most proportion, which often severely impacted protein function. In addition, the same BRAT1 splice site synonymous mutation had been reported in a Chinese female case and predicted to cause splice site disruption[7]. Using a transcriptional study, we first confirmed that the synonymous mutation caused the exon 10 to skip, which could influence the structure and function of the BRAT1 protein.

In conclusion, the homozygous synonymous mutation of BRAT1 causing splice site disruption in our patient led to RMFSL. Trio-WES is a powerful approach to clarify the underlying molecular of unexplained intractable seizures, and it is vital for guiding future fertility.

CONFLICT OF INTEREST

The authors declare no conflict of interest. The Ethics Committee of The First Affiliated Hospital of Anhui Medical University approved this case report. In addition, the patient’s guardian approved sharing the case presentation without exposing privacy.

ACKNOWLEDGEMENTS

We always thank the patient and his parents for participating in this study.

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Qi Y, Ji X, Ding H, Liu L, Zhang Y, Yin A (2022) Novel Biallelic Variant in the BRAT1 Gene Caused Nonprogressive Cerebellar Ataxia Syndrome. FRONT GENET 13. DOI:10.3389/fgene.2022.821587
Randy H Van Ommeren AFGS (2018) BRAT1 Mutation: The First Reported Case of Chinese Origin and Review of the Literature. J Neuropathol Exp Neurol 12: 1071–1078. DOI:10.1093/jnen/nly093
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Table 1

Characteristics of individuals with some reported BRAT1 mutations

	Mutations type of BRAT1	Gene and Protein variant	Clinical Phenotypes	Age at death of each patient
Celik Y, et al.[2]	homozygous	c.2230_2237dupAACACTGC (p.S747Tfs*36)	RMFSL	Ten months
Hanes I, et al.[3]	compound heterozygous	c.294dupA (p.Leu99Thrfs*92); c.1825C>T (p.Arg609Trp)	Delayed RMFSL	3 years 8 months
Van de Pol et al.[4]	homozygous	c.638dupA (p.Val214Glyfs*189)	RMFSL	3 months/17months/2 months
Puffenberger et al[5]	homozygous	c.638_639insA (p.Val214Glyfs*189)	RMFSL	One month in one， and< 4 months in others
Qi Y et al.[6]	compound heterozygous	c.1014A > C (p.Pro338 = ); c.706delC (p.Leu236Cysfs*5)	Very mild phenotype	At the time of publication, the patient was more than five years old.
Van Ommeren RH, et al.[7]	homozygous	c.1395G>C （p.Thr465Thr）	RMFSL	10weeks
Saitsu et al.[8]	compound heterozygous	Patient1. c.176T>C (p.Leu59Pro); Patient 2. c.962_963delTC (p.Leu321Profs*81)	RMFSL	21 months/3months
Srivastava S, et al.[10]	compound heterozygous	Patient1-2. c.638dupA (p.Val214Glyfs*189); c.803+1G>C	Mild phenotype	10 years /6years
		Patient3. c.638dupA (p.Val214Glyfs*189); c. 419T>C (p.Leu140Pro)	Moderate phenotype	4 years and 4 months
		Patient4.c.171delG(p.Glu57Aspfs*7); c. 419T>C (p.Leu140Pro)	Moderate phenotype	15months

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A BRAT1 synonymous mutation affecting exon splicing leads to lethal neonatal rigidity and multifocal seizure syndrome: A Chinese case report.

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