Role of Neuron-Specific Splicing Regulators as Modifiers of Neurofibromatosis Type 1

摘要

We hypothesized that the tightly regulated ratio of NF1 isoforms is critical in maintaining the homeostasis of cell growth and differentiation, thereby determining the functional output of NF1 gene. A corollary is that splicing regulators may modify the function of neurofibromin through altering alternative splicing of the NF1 pre-mRNA. We propose two specific aims to study the role of regulated alternative splicing in the function of neurofibromin. In Aim I, we will determine how changes in alternative splicing affects NF1 function. In Aim II, we will determine the biological consequence of altering the ratio of neurofibromin isoforms in cells with natural NF1 expression such as neuronal and glial cells. To this end, we have identified the cis-acting elements located to the vicinity of exon 23a that play key roles in regulating inclusion of this exon and tested the effects of mutations of these elements in primary neurons. We also generated the gene-targeting vector to create a knock- in allele at the NF1 locus in mouse ES cells and are in the process of screening ES cell clones that have the correctly targeted allele. In addition, we successfully established the ES cell differentiation procedure. Results of the proposed studies will not only provide important novel insights into the etiology of NF1 disease, but also shed light on how genetic variations in splicing regulators affect the progression of other diseases.