NF1mutation rather than individual genetic variability is the main determinant of theNF1-transcriptional profile of mutations affecting splicing

Abstract

A significant number of neurofibromatosis type 1 (NF1) mutations result in exon skipping. The majority of these mutations do not occur in the canonical splice sites and can produce different aberrant transcripts whose proportions have not been well studied. It has been hypothesized that differences in the mutation-determined NF1-transcriptional profile could partially explain disease variability among patients bearing the same NF1 splice defect. In order to gain insight into these aspects, we analyzed the proportion of the different transcripts generated by nine NF1-splicing mutations in 30 patients. We assessed the influence of the mutation in the NF1-related transcriptional profiles and investigated the existence of individual differences in a global manner. We analyzed potential differences in tissue-specific transcriptional profiles and evaluated the influence of sample processing and mRNA nonsense-mediated decay (NMD). Small transcriptional differences were found in neurofibromas and neurofibroma-derived Schwann cells (SC) compared to blood. We also detected a higher cell culture-dependent NMD. We observed that mutation per se explains 93.5% of the profile variability among mutations studied. However, despite the importance of mutation in determining the proportion of NF1 transcripts generated, we found certain variability among patients with the same mutation. From our results, it seems that genetic factors influencing RNA processing play a minor role in determining the NF1-transcriptional profile. Nevertheless neurofibromin studies would clarify whether these small differences translate into significant functional changes that could explain the great clinical expressivity observed in the disease or any of the disease-related traits.

Nature and mRNA effect of 282 differentNF1point mutations: focus on splicing alterations

“…A total of 124 (44%) mutations of various types affected splicing (173/374 patients). We and others have shown that several of these mutations generate a NF1-transcriptional profile rather than a single mutated mRNA, where correctly spliced mRNAs containing the original DNA mutation coexist with mRNAs exhibiting a splicing alteration (Messiaen et al, 2000;Wimmer et al, 2000b;Zatkova et al, 2004;Pros et al, 2006). For example, we have already shown that mutations c.910C>T, c.5546G>A, c.6791dupA and c.6792C>A generate different proportions of mutated transcripts for each NF1 mutation (Pros et al, 2006) and that these proportions were similar among patients with the same mutations.…”

“…A previous study of the NF1-associated transcriptional profile in patients carriers of several Type V recurrent mutations detected always two types of aberrant transcripts (Pros et al, 2006). The proportion of each transcript varied according to the mutation, some mutations generated high proportions of transcripts including the mutated exon while others led to exon skipping in the majority of transcripts.…”

“…In a previous study we examined the transcriptional profile of one of these recurrent mutations, c.5546G>A. While most of the transcripts showed exon skipping, still a small proportion contained the exon with the missense mutation (Pros et al, 2006).…”

“…The proportion of each transcript varied according to the mutation, some mutations generated high proportions of transcripts including the mutated exon while others led to exon skipping in the majority of transcripts. The ratio of exon containing the original mutation and exon skipped transcripts was unique for each mutation and was well preserved among different patients carrying the same mutation (Pros et al, 2006). These mutations are difficult to classify due to the presence of two abnormal transcripts from the mutated allele and the variety of different expression patterns.…”

Modulation of aberrant NF1 pre-mRNA splicing by kinetin treatment

“…A previous study by our group demonstrated that mutations in exons affecting correct NF1 splicing produce two types of aberrant transcripts: transcripts with the original mutation and transcripts showing exon skipping. 2,12 Skipping of exons 7, 18 and 37 is in frame, but skipping of exon 36 is out of frame. It is well known that nonsense transcripts could be subjected to degradation by the NMD phenomena.…”