Genomic anatomy of the Tyrp1 (brown) deletion complex

Abstract

Chromosome deletions in the mouse have proven invaluable in the dissection of gene function. The brown deletion complex comprises >28 independent genome rearrangements, which have been used to identify several functional loci on chromosome 4 required for normal embryonic and postnatal development. We have constructed a 172-bacterial artificial chromosome contig that spans this 22-megabase (Mb) interval and have produced a contiguous, finished, and manually annotated sequence from these clones. The deletion complex is strikingly gene-poor, containing only 52 protein-coding genes (of which only 39 are supported by human homologues) and has several further notable genomic features, including several segments of >1 Mb, apparently devoid of a coding sequence. We have used sequence polymorphisms to finely map the deletion breakpoints and identify strong candidate genes for the known phenotypes that map to this region, including three lethal loci (l4Rn1, l4Rn2, and l4Rn3) and the fitness mutant brown-associated fitness (baf). We have also characterized misexpression of the basonuclin homologue, Bnc2, associated with the inversion-mediated coat color mutant white-based brown (B w ).This study provides a molecular insight into the basis of several characterized mouse mutants, which will allow further dissection of this region by targeted or chemical mutagenesis.brown locus ͉ chromosome deletion ͉ mouse genome sequence T he mouse brown (b) mutation is one of the oldest known loci and was one of the first to be cloned. The mutation is in the tyrosinase-related protein 1 (Tyrp1) gene, encoding a melanocyte enzyme required for the production of dark eumelanin (1). Homozygous loss of Tyrp1 results in a brown coat; because it was a simply scored phenotype, brown was used as part the mousespecific locus test carried out at the Oak Ridge National Laboratories and elsewhere (reviewed in ref.2). Wild-type mice were exposed to chemical or radiological mutagens and crossed with a tester stock homozygous for seven recessive visible mutations. The efficacy of the mutagenic treatments is assessed in the resulting F 1 progeny. These experiments generated many chromosomal deletions that inactivated Tyrp1, particularly in the progeny of animals exposed to radiation or clastogenic chemical mutagens such as chlorambucil or melphalan.Deletions around the seven specific loci have provided a unique opportunity to study gene function within these intervals. Individual deletions are hemizygous viable but when homozygous or in combination with others give lethal or visible phenotypes, which indicate the presence of essential genes neighboring the specific loci. Intercrossing deletions of different extents provided an avenue to map the resulting phenotypes. The deletions also provided the means to produce physical maps of genetic markers. Studies of this kind have been published for several loci, including albino (Tyr), piebald (Ednrb), pink-eyed dilution (p), and the brown deletion complex (2-6).Studies of the brown deletions established genetic ...

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“…[27]) with regard to appearance and viability of homozygotes (namely, fully viable in the case of five of the loci, juvenile lethal in the case of Myo5a and Ednrb). An extensive literature on molecular characterization of the marker genes and their immediately flanking chromosomal regions (e.g., [8,29,30,6,7,4,31,32]) supports the conclusion that most, if not all, mutations classified OL in the present experiments did not involve lesions outside the marked loci. In past investigations, cases where such a conclusion did not always hold involved radiation-induced nulls, e.g., at Tyr [30] and Bmp5 [33].…”

Basonuclins 1 and 2, whose genes share a common origin, are proteins with widely different properties and functions

“…A chromosomal inversion, white-based brown (B w ), also occurs within the brown locus. Overexpression of bn2 placed under the control of the Tyrp-1 promoter by the inversion almost certainly is responsible for the melanocyte cell death that is presumed to cause the inversion phenotype (27). It appears probable that these two phenotypes are related to alterations in RNA processing caused by the absence or overexpression of bn2.…”

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“…The mutation occurred at the Oak Ridge National Laboratories (Oak Ridge, TN) in C3H male animals exposed to ionizing radiation. The defect has been characterized as an inversion of the first exon of the tyrp-1 gene, 52 resulting in complete absence of the TRP-1 protein expression, even in truncated or mutated form, thus B w mice represent a true immunologic knockout of TRP-1. 52 B w mice were backcrossed onto the C57BL/6n background for 8 generations using a speed congenic approach.…”