Lissencephaly (smooth brain, LIS) syndromes are a group of presumed neuronal migration disorders producing brain malformations and epilepsy. The LISI gene in 17pl3.3 encodes the subunit of platelet activating factor acetylhydrolase (PAFAHlb) and its mutation results in Miller-Dieker syndrome (MDS) or isolated lissencephaly sequence (ILS). Mutations of the X-linked locus, XLIS, produce ILS in hemizygous males and less severe, subcortical band heterotopia (SBH or double cortex, DC), in heterozygous females. Investigators in this project and collaborators have identified a putative XLIS gene in Xq22.3, whose encoded protein is designated doublecortin (Dbcn). A novel protein, Dbcn contains a likely site for tyrosine phosphorylation by the cAbl kinase, a region of homology to a novel member of the CAM kinase family, and a Ser/Pro rich domain which is a potential site of protein-protein interactions. Experiments in the competing renewal will produce several mouse models to investigate the role of Dbcn in neuronal migration. The Co-principal investigator has already produced a mouse model of Lisl inactivation that will be compared with Xlis models.
Aim 1 will inactivate Xlis by homologous recombination to test the hypothesis that loss of Dbcn function in mice will closely mimic the human malformations.
Aim 2 will examine the development of Xlis null, Lisl and Xlis heterozygous cortices to test the hypothesis that loss of either Lisl or Dbcn will a) impair neuronal migration and b) affect tangential patterns of neuronal migration more than radial. Classical static histological and BrdU labeling techniques will be used to study the models created in Aim 1. In addition, the migration patterns of neurons in both the LISI heterozygotes and XLIS nulls will be compared in embryonic cortical slice cultures using time lapse confocal microscopy in which neurons are labeled by DiI or by GFP expressed under the direction of the Xlis promoter.
In Aim 3, genetic crosses will test the hypothesis that Dbcn is part of an intracellular signaling cascade and is activated by the cAbl tyrosine kinase in response to adhesion, serving to link signals at the cell surface with neuronal migration. Possible interactions between Dbcn and BLisl or Dbcn and the mDabl migration protein will be examined in double mutants created by crossbreeding mutant mouse strains.
Aim 4 will examine the hypothesis that point mutations in XLIS families produce milder phenotypes by virtue of partial loss of function or disregulation of Dbcn. A transgenic model will test whether over-expression of Dbcn activity can also disrupt migration. Mouse models of XLIS will help to reveal the pathogenesis of the X-linked form of LIS and SBHDC, will definitively determine whether XLIS is a neuronal migration gene, and will help to define genetic mechanisms leading to a major class of human developmental disorders and epilepsy.
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