Structural Genes of the Urinary Protein Are on Mouse Major Chromosome 4

The major urinary proteins (MUPs) of mouse are a family of at least three major proteins which are synthesized in the liver of all strains of mice. The relative levels of synthesis of these proteins with respect to each other in the presence of testosterone is regulated by the Mup-a locus located on chromosome 4. In an effort to determine the mechanism of this regulation in molecular terms, a cDNA clone containing most of the coding region of a MUP protein has been isolated and identified by partial DNA sequence analysis. Using a combination of hybridization analysis and somatic cell genetics, the structural gene family has been unambiguously mapped to mouse chromosome 4. These data suggest that Mup-a regulation operates in a cis fashion and that models proposing trans regulation of MUP protein synthesis are unlikely. The differential expression of members of a multigene family in the course of the life cycle of an organism is a well known phenomenon, and describing the regulation of this expression in molecular terms is a popular topic in eucaryotic developmental biology. One widely used model for such regulation proposes the existence of a constant family of structural genes, dependent upon a regulatory gene (or genes) that modulates expression of the various members of the family at the appropriate points in development. Polymorphisms in the regulatory gene would further cause heritable variations in the expression of the family even though the structural genes themselves were unchanged. Such a model has been discussed for the expression of the family even though the structural genes themselves were unchanged. Such a model has been discussed for the expression of H-2 and TL surface antigens on normal and neoplastic mouse tissues (1), and for the expression of immunoglobulin allotype markers in rabbits and mice (17). The model might also be applied to the regulation of synthesis of the group of major urinary proteins (MUPs) in mice (15, 16). However regulatory elements of gene families in mammalian cells have not been identified as independent genetic loci separable from the gene families they might control as have such loci in yeast, for example (2). The MUPs are a series of at least three closely related electrophoretically separable proteins, MUP1, 2, and 3 (6, 7, 12), synthesized in large amounts in the liver of malez and 414 testosterone-treated females, secreted into the blood, and excreted with the urine. All inbred strains of mice tested can produce all three MUP proteins (15). Thus, BALB/c mice produce 80% of MUP1, barely detectable amounts of MUP2, and 20% of MUP3, whereas C57BL/6 mice produce 20% MUPI, 40% MUP2, and 40% MUP3 (14, 16). The difference in the levels of the MUP1 and 2 proteins produced by these different strains of mice has been hypothesized to be the result of an element termed Mup-a that segregates with the markers known to be on chromosome 4. If an animal produces mostly MUP1 and little MUP2 it has been described as Mup-a E and if it produces more MUP2 than MUP1 it is described as Mupa 2. It however has not been possible in crosses of inbred strains to separate genetically the presumed alleles of the Mup-a locus from the "structural" MUPI and 2 proteins. This is because no genetic assay (e.g. polymorphism of MUP-proteins) for the structural genes exists. With the advent of recombinant DNA it is possible to determine chromosome locations of genes by molecular techniques. To determine the basis for MUP expression in different mouse strains we have begun an isolation and characterization of the mouse DNA that encodes the MUP gene family in the two strains. In this report we describe characterization of a recombinant clone containing double-stranded cDNA corresponding to the mRNA encoding one of the MUPs. Using this clone as a hybridization probe, we have investigated the MUP THE JOURNAL OE CELL BIOLOGY VOLUME 94 AUGUST 1982 414-417 © The Rockefel ler Univers i ty Press 0021-9525/82/08/0414/04 $1.00 on O cber 9, 2017 jcb.rress.org D ow nladed fom structural gene family and shown that the structural genes encoding the MUP proteins are located on the same chromosome as the putative regulatory locus. M A T E R I A L S A N D M E T H O D S Liver cDNA clones were isolated, characterized, and cloned DNA prepared as previusly described (5). DNA for sequence analysis was cleaved with appropriate restriction endonucleases and labeled at the 3' termini with cordycepin tripliosphate using a kit supplied by New England Nuclear Corp. (Boston, MA). DNA sequencing was performed by the chemical method of Maxam and Gilbert (10). Mouse hamster cell hybrids were prepared, characterized, and maintained as previously described (3, 13). DNA for mapping experiments was cleaved with EcoR l according to supplier's instructions. Gel electrophoresis, transfer of DNA from gels to nitrocellulose, and hybridization of t'dters has been previously described (3, 5).