Effect of L1ECD on mouse primarily cultured neurons and construction of transgenic mice specifically expressing L1ECD in brain]

Neural cell adhesion molecule L1 is an important molecule mediating cell-cell interactions during the development of nervous system. L1 can promote axonal outgrowth and is related with nerve cell migration, and therefore L1 plays an important role both in the development and maintaince of the nervous system. In humans, mutations in the L1 gene can lead to mental retardation, spastic paraplegia, hydrocephalus, and other developmental abnormalities. The molecular mechanisms of mutations in L1 gene to induce inherited neurological diseases are not clear. In present investigation, a transgenic DNA of mouse L1 extracellular domain (L1ECD) was constructed by adding a stop codon to the end of L1ECD cDNA and then putting it under the control of CAMK II promoter, which is active specifically in the brain. To verify this construct, L1ECD cDNA was subcloned into an expression vector pCEP4 and then transfected the C6 cells. The expression of L1ECD cDNA in C6 cells was confirmed by Northern blotting and the effects of L1ECD on the growth rate and morphology of C6 cells in vitro as well as primarily cultured neurons were observed. The L1ECD constructs were microinjected into the fertilized zygotes of C57BL/6 mice. The transgenic mice thus produced were identified by Southern and Northern hybridization analysis. The results demonstrated that the L1ECD was integrated in the genome of transgenic mice and expressed specifically in the brain.