Leber congenital amaurosis with early-onset severe macular atrophy and optic atrophy is likely pathognomonic of NMNAT1 mutations

Abstract

Purpose: The nuclear nicotinamide mononucleotide adenyltransferase 1 (NMNAT1) encodes a homohexameric NAD- synthesizing enzyme as well as a chaperone that protects against neuronal activity-induced degeneration. NMNAT1 mutations have been reported to cause a highly specific Leber congenital amaurosis (LCA) phenotype characterized by severe neonatal neurodegeneration of the central retina with early-onset optic atrophy. The purpose of the present study was to search for second NMNAT1 disease alleles in single heterozygote patients harboring the NMNAT1 phenotype.

Methods: Sixteen sporadic cases and two sibs harboring single heterozygote NMNAT1 mutations were screened for copy number variations (CNV) using quantitative multiplex PCR of short fluorescent fragments (QMPSF) and qPCR digital droplet; and for mutations affecting regulatory elements or the splicing using Sanger sequencing of genomic DNA and/or lymphoblastoid cDNA. NMNAT1 haplotypes were constructed using SNP and microsatellite segregation analysis to search for shared ancestral alleles.

Results: cDNA sequencing and/or CNV analysis allowed identifying a one-exon duplication in the two sibs, and 1 or 2-exon deletions in 4/17 unrelated patients. Sanger sequencing of non-coding NMNAT1 regions allowed identifying two rare variants possibly affecting a 5’ regulatory elements in 6/17 index cases. One out of the two variants was shared by 5 index cases originating from a French Indian Ocean Island (La Réunion). All five individuals harbored a common haplotype at the NMNAT1 locus supporting the transmission of an ancestral allele. Interestingly, none of the two variants could be detected at the cDNA level, suggesting down regulation or instability of the mRNAs harboring these changes. Two other 5’ changes were identified in 2/17 individuals which analysis is ongoing.

Conclusions: Here, we report that at least 14/18 LCA individuals with single heterozygote NMNAT1 mutations carried a second disease allele undetected by Sanger sequencing of the NMNAT1 exome. This result suggests that severe neonatal neurodegeneration of the central retina with early-onset optic atrophy is pathognomonic of NMNAT1 mutations. In-depth molecular analysis of the gene and surrounding regulatory elements should be considered in all patients harboring this highly specific LCA phenotype