Assessment of individual functions of Myc family genes

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To check the possibility of functional redundancy between the N-myc and L-myc genes, we generated N^<+/->L^<-/-> as well as N-myc and L-myc double mutant mice (N^<-/->L^<-/->). N-myc and L-myc double mutant embryos showed an overall phenotype similar to ablating N-myc only in mice (N^<-/->L^<+/+> or N^<-/->L^<+/>). The results indicate that L-myc is not essential for embryonic development, and negates the notion of redundant functions between the N-myc and L-myc genes.To identify N-myc regulatory target genes we subtracted total cDNAs of mutant embryo from wild type cDNAs at 10.5 dpc and vice versa, and isolated cDNA of down-regulated mRNA in the mutants or of up-regulated mRNA species in the opposite combination. One of the N-myc downstream genes, Ndrl was studied in detail. The promoter of Ndrl gene was directly repressed by N-myc and Max heterodimer complex. In various embryonic organ rudiments an inverse relationship was found between the expression of N-myc and Ndrl, indicating that repression of the latter by the former is a regulation operating in embryogenesis.To address whether N-myc and c-myc have overlapping functions, we have introduced the c-myc coding region into the N-myc locus so that it comes under the regulation of N-myc. No heterozygous offspring were observed among 32 ES cell "fathered" pups. This is a strong indication that the embryos expressing c-myc from the N-myc locus die in utero or may not even be generated.