Cloning And Functional Characterization Of The Human The Numb Ligand Protein Genes Lnx

Asymmetric cell division, in which a cell divides into two cells of different developmental potentials, is a fundamental means of generating cell diversity. Intrinsic cell fate determinant Numb is the first protein found to be asymmetrically distributed during neural development. From the human fetal brain cDNA library we cloned, we obtain a novel cDNA highly homologous to the mouse Lnx (Ligand of Numb Protein-X) gene. So we name this gene human LNX in agreement of the HUGO nomenclature committee. The 3737 bp cDNA has an open reading frame from nucleotide 236 to nucleotide 2134, encoding a protein with a LDNPAY motif for the binding of Numb PTB domain and 4 PDZ domains. Northern hybridization, RT-PCR and in situ hybridization indicate that LNX mRNA has a specific distribution in brain, and is highly expressed in glioma. In situ hybridization of different developmental stages of mouse kidney shows LNX expression in renal medulla, and its expression decreases as kidney develops. Cluster analysis of microarray data reveals some LNX-related genes, including FE65 and Frizzled involved in signaling pathways. Using the PDZ domain in LNX protein as bait, we screened for LNX-interacting proteins by yeast-2-hybrid system. We found that LNX can interact with Ski-interacting protein (SKIP) and PDZ-binding kinase (PBK). Yeast mating assay, in vitro pulldown assay and co-immunoprecipitation assay verified their binding. LNX can also colocalize with Numb, SKIP and PBK, respectively, in nucleus. We speculate that LNX protein may recruit related proteins to certain site in the cell via its PDZ domains, and thus involved in some signaling pathways including Notch pathway.