Case Report: A Novel KMT2E Splice Site Variant as a Cause of O'Donnell-Luria-Rodan Syndrome in a Male Patient

Abstract

Background: O'Donnell-Luria-Rodan (ODLURO) syndrome is an autosomal dominant systemic disorder characterized by global developmental delay caused by mutations in the KMT2E gene. The aim of this study was to investigate the role of KMT2E mutations as a cause of ODLURO syndrome in a Chinese boy.

Methods: We reported the clinical course of a Chinese boy who was diagnosed with ODLURO syndrome by the whole exome sequencing. We extracted genomic DNA of the proband and parents, gene variations were screened using whole-exome sequencing, followed by validation using direct Sanger sequencing. The effect of mRNA splicing variants were analyzed through a minigene splice assay and in vitro reverse transcription PCR (RT-PCR).

Results: The proband presented with recurrent seizures and developmental delay. Using genetic analysis, we identified that the proband carried a de novo heterozygous splicing variant (c.1248+1G>T) in the KMT2E gene. In vivo transcript analysis showed that the proband did not carry any KMT2E mRNA transcript, while a specific exon11-exon13 (440 bp) transcript was detected in the unaffected parents. The in vitro minigene splice assay conducted in HEK293 cells confirmed that the c.1248+1G>T variant resulted in exon 12 skipping, which in turn caused an alteration in KMT2E mRNA splicing. The mutant transcript created a premature stop codon at the 378 amino acid position that could have been caused nonsense-mediated mRNA decay (NMD).

Conclusion: We verified the pathogenic effect of the KMT2E c.1248+1G>T splicing variant, which disturbed normal mRNA splicing and caused mRNA decay. Our findings suggest that splice variants play an important role in the molecular basis of ODLURO, and that careful molecular profiling of these patients could play an essential role in tailoring of personalized treatment options soon.

Keywords: KMT2E; O'Donnell-Luria-Rodan's syndrome; abnormal mRNA splicing; epilepsy; splice variant.

Figure 1

KMT2E gene variant and auxiliary examination of the patient. (A) Schematic presentation of the familial pedigrees. Males and females are designated by squares and circles, respectively. The shapes filled with shadow indicate affected individuals with the KMT2E variant; the arrow indicates the proband. (B) Direct sequencing identified the novel KMT2E (c.1248+1G>T) variant (C) Brain MRI of the patient showing delayed myelination (D) The EEG reports of the patients, from top to bottom, representing the background slow wave, general fast wave rhythm during sleep, and general fast wave rhythm during the waking state.

Figure 2

Effect of the KMT2E gene c.1248+1G>T variant determined by minigene assays. (A) The pSPL3 vector contains 2 exons SD6 and SA2, and a functional intron. All of the indicated fragments were separately cloned into the XhoI and BamHI cloning sites of the pSPL3 vector. SD6 and SA2 primers were designed for RT-PCR amplification of cDNA sequences generated by transfected HEK293 cells. (B) Gel electrophoresis of the RT–PCR products of minigene transcripts in HEK293 cells. Lane 1: marker; Lane 2 and 3: pSPL3 (263 bp); Lane 4 and 5: E12-wide type (381 bp); Lane 6 and 7: c.1248+1G>T (263 bp). The two fragments were directly sequenced (right panel). Sequencing analysis of the cDNA showed that the shorter transcript lacked a sequence corresponding to exon 12 of the KMT2E gene. (C) Exon 12 and adjacent structures in the KMT2 gene. The arrow shows the location of the c.1248+1G>T splice variant and its effect at the amino acid level. The variant caused by a premature stop codon at amino position 378.