PLP Splicing In Vivo Analysis in a Mouse Model

The long-term goals of this project are to elucidate the function of a G-rich 19-bp sequence in intron 3 of the PLP gene in PLP-DM20 splice site selection in vivo. We have shown that deletion of this sequence causes a neurological disease in humans and that this sequence regulates selection of PLP splicing in oligodendrocytes in vitro. G-rich elements have been shown to regulate splice site selection in chimeric genes in vitro. We propose to generate a line of knock-in mice carrying deletion of the G-rich 19-bp sequence and to utilize these mice to investigate PLP specific splicing in brain and peripheral nerves during development. In these studies, we will characterize splicing at the early stages of myelination, peak of myelination and adulthood. The production of PLP and other myelin proteins and myelin formation will be studied. The mouse phenotype will be characterized by motor testing. To our knowledge, mouse models designed to investigate in vivo regulation of alternative splicing are not available and none of the PLP mouse mutants allows studies of PLP splicing. The mouse model we propose to make will allow investigating the regulation of alternative splicing in the PLP gene in the context of the entire gene and in the presence of external cues. In addition, it has the potential to generate data of broader biological relevance about splicing in many other genes.