Next-generation sequencing corroborates a probable de novo GNPTG variation previously detected by Sanger sequencing.

Letter to the Editor

Mucolipidosis III (MLIII) is an autosomal recessive disease caused by pathogenic variations in the GNPTAB (MLIII alpha/beta) or GNPTG (MLIII gamma) genes. GNPTAB and GNPTG encode, respectively, the α/β and γ subunits of GlcNAc-1-phosphotransferase, the enzyme responsible for catalyzing the addition of a mannose-6-phosphate residue to lysosomal hydrolases, allowing their entry into lysosomes [1], [2].

In 2014, our group published a paper describing an MLIII gamma patient, born to a non-consanguineous couple, who carried the variations c.244_247dupGAGT at exon 3 and 328G > T at exon 5, both detected by Sanger sequencing of GNPTG. Analysis of parental DNA using the same technique showed c.328G > T in the father and no alterations in the mother, despite using different pairs of primers and different tissues. Maternity was confirmed, and c.244_247dupGAGT was thus considered a de novo mutation [3].

Next-generation sequencing (NGS) has proven more sensitive than Sanger sequencing for the detection of mosaicism when the variants are located in exons, as in the present case [4], [5], [6], [7], [8], [9], [10]. However, NGS has several limitations, such as incomplete acceptable coverage of coding regions due to regions rich in CpG islands or repetitive sequences [11]. Thus, we decided to analyze DNA samples obtained from the blood of the patient and her mother using the Ion 314™ Chip Kit v2 and sequence GNPTG with the Ion PGM Hi-Q Sequencing Kit (Thermo Fisher Scientific). The fragment of GNPTG where c.[244_247dupGAGT] is located was amplified 406 and 1047 times in the maternal and patient samples, respectively. No alteration was identified in the maternal sample, while both were identified in the patient, confirming our previous findings. As we failed to detect mosaicism using this technique, the possibility of a de novo event is even higher. Our findings corroborate the hypothesis that de novo variations may be more frequent than expected [12], and speak strongly in favor of always investigating the parents of a child presenting with an autosomal recessive disease in order to confirm heterozygous status.

Conflict of interest

The authors declare no conflict of interest.