Identification and characterization of 15 novel GALC gene mutations causing Krabbe disease

The characterization of the underlying GALC gene lesions was performed in 30 unrelated patients affected by Krabbe disease, an autosomal recessive leukodystrophy caused by the deficiency of lysosomal enzyme galactocerebrosidase. The GALC mutational spectrum comprised 33 distinct mutant (including 15 previously unreported) alleles. With the exception of 4 novel missense mutations that replaced evolutionarily highly conserved residues (p.P318R, p.G323R, p.I384T, p.Y490N), most of the newly described lesions altered mRNA processing. These included 7 frameshift mutations (c.61delG, c.408delA, c.521delA, c.1171_1175delCATTCinsA, c.1405_1407delCTCinsT, c.302_308dupAAATAGG, c.1819_1826dupGTTACAGG), 3 nonsense mutations (p.R69X, p.K88X, p.R127X) one of which (p.K88X) mediated the skipping of exon 2, and a splicing mutation (c.1489+1G>A) which induced the partial skipping of exon 13. In addition, 6 previously unreported GALC polymorphisms were identified. The functional significance of the novel GALC missense mutations and polymorphisms was investigated using the MutPred analysis tool. This study, reporting one of the largest genotype-phenotype analyses of the GALC gene so far performed in a European Krabbe disease cohort, revealed that the Italian GALC mutational profile differs significantly from other populations of European origin. This is due in part to a GALC missense substitution (p.G553R) that occurs at high frequency on a common founder haplotype background in patients originating from the Naples region.     

Complex cystic fibrosis allele R334W-R1158X results in reduced levels of correctly processed mRNA in a pancreatic sufficient patient

CFTR alleles containing two mutations have been very rarely found in cystic fibrosis (CF) patients. They provide an opportunity to study the effect of two in cis-interacting gene defects on gene expression. Here, we describe a three-generation CF family with a complex CFTR allele that has not been previously described, containing the missense mutation R334W in exon 7 and the nonsense mutation R1158X in exon 19. Lymphocyte RNA analysis showed that (1) the mRNA corresponding to the complex allele is present although at markedly reduced levels; and (2) the nonsense mutation does not lead to detectable skipping of exon 19. The clinical picture of the patients with the genotype R334W-R1158X/ΔF508 is characterized by pancreatic sufficiency and an atypical course of the disease. © 1996 Wiley-Liss, Inc.     

Three new adenosine deaminase mutations that define a splicing enhancer and cause severe and partial phenotypes: implications for evolution of a CpG hotspot and expression of a transduced ADA cDNA

We report three novel adenosine deaminase (ADA) mutations withinteresting implications. A Somali child with severe combinedimmunodeficiency disease (SCID) had reduced ADA mRNA in T cellsand was homozygous for the nonsense mutation Q3X. Unexpectedly,her healthy father was a compound ADA heterozygote whose secondallele carried a ‘partial’ mutation, R142Q, dueto a GA transition of a CpG dinucleotide. A CT transition ofthe same CpG produced a nonsense mutation, R142X, in two homozygousCanadian Mennonite infants with SCID. The severe and healthyphenotypes associated with R142X and R142Q, the high frequencyof ‘partial’ ADA mutations arising from CpGs inhealthy individuals of African descent and the presence of CAA(glutamine) at codon 142 in murine ADA, suggest selection forreplacement of this CpG hotspot by CpA during ADA evolution.R142X, located within a purine-rich segment at nt 62/116 ofexon 5, caused skipping of the exon, possibly by disruptinga splicing enhancer. Absence of exon 5 in T cell ADA mRNA andlow ADA activity in T cells and erythrocytes obtained at age18–22 months from one of the Mennonite children, indicatelimited expression of a normal ADA cDNA from retrovirally transducedCD34+ umbilical cordleukocytes infused shortly after birth inan attempt at stem cell gene therapy.