ANALYSIS OF INSERTIONAL MUTATIONS IN TRANSGENIC MICE

许多感觉器官的先天性异常，如眼和耳

Many congenital abnormalities of sensory organs such as eye and ear malformations are the result of derailed development due to malfunctioning genes. We have recently discovered a basic-helix-loop- heli-zipper transcription factor gene, called microphthalmia (mi), whose mutations in mice are associated with smaller-than-normal eyes, inner ear deafness, and loss of coat pigmentation. We have also isolated the human homolog of this mouse gene, and it turns out that a particular form of human syndromic hearing impairment called Waardenburg syndrome IIa is in fact due to mutations in this gene. In mice, the common denominator of the abnormalities is the aberrant development or plain absence of pigment cells in eye, inner ear and skin, and the same may be true in humans. We have now determined the developmental expression profile of mi in wild type and mutant mouse embryos as well as their cultured melanocytes. In wild type mouse embryos, expression starts in cells in the outer layer of the developing optic cup and soon thereafter in a very small number of cells derived from the neural crest. A few hours after expressing mi, the majority of these cells start to express Trp2, a melanoblast marker, and thus they seem to be committed to the melanocyte lineage. Since in some locations mi expression is found in cells located still in the dorsal wall of the neural tube, commitment may start prior to emigration. In embryos homozygous for certain mutant alleles, neural crest-derived mi-expressing cells are hardly detectable, and staining for other melanoblast markers such as Trp2 remains negative. However, the retinal pigment layer cells stay in place and continue to express Trp2 but not Trp1 and tyrosinase, two other melanocyte markers that based on in vitro analyses are direct target genes of mi. Since after birth, mi is no longer expressed in pigment cells except in those of the hair bulbs, it appears that all abnormalities in eyes and ears may be determined prior to birth, whereas abnormalities in skin pigmentation may continue to develop during life. In a collaborative effort, we have also started to determine the precise molecular defects associated with different mutant alleles in mice and men to analyze the biochemical and biological consequences of these defects in vitro and in tissue culture cells. Interestingly, coexpression of different mutant forms of the Mi protein in compound heterozygotes may lead to milder or more severe abnormalities when compared to the corresponding homozygous mice. Future studies will show whether this observation can be explained solely on the basis of formation of dimers between different mutant forms of the Mi protein or whether it implies involvement of other, related basic helix- loop-helix-zipper transcription factors.