Highly conserved upstream regions of the alpha 1-antitrypsin gene in two mouse species govern liver-specific expression by different mechanisms.

Abstract

ax-Antitrypsin (AT), the major elastase inhibitor in mammalian serum, is produced primarily in the liver. We have characterized AT gene structure and expression in the mouse species Mus caroli, which expresses high levels of AT in the kidneys as well as in the liver. Analysis of cDNA and genomic clones showed that the AT gene in M. caroli exhibits high sequence homology (>90%) to the gene in laboratory mice (M. domesticus) throughout the coding and 5'-flanking regions. Despite this extensive sequence conservation, the functional organization of cis-acting regulatory elements governing liver-specific expression is strikingly different between these species. Transient-transfection assays showed that the proximal region of the M. caroli promoter (i.e., between -120 and -2 relative to the transcriptional start site) is 10-fold more active than the analogous region of M. domesticus in driving the expression of an indicator gene in cultured liver cells. The increased activity of the proximal region of the M. caroli AT promoter appears to be the result of one or both of the two base substitutions at positions -46 and -48. The weak proximal promoter in M. domesticus is compensated for by the presence of upstream, liver-specific enhancers between -199 and -520; the analogous region in M. caroli is inactive. Thus, during the course of evolution, the modest 7% sequence divergence that has occurred between the 5'-flanking regions of the AT genes in these two species has generated distinct, yet equally effective, modes of hepatocyte-specific expression.The molecular basis for the liver-specific expression of atl-antitrypsin (AT), a major seine protease inhibitor that functions in the control of neutrophil elastase activity (4, 29), has been intensely investigated in humans and mice. For the human gene, maximal liver-specific expression requires 261 nucleotides of the 5'-flanking region, which can be subdivided into a distal element located between -261 and -210 and two proximal elements located between -137 and -37 (6,8,26). Although the proximal elements function only in hepatic cells, the distal element acts as a nonspecific enhancer (8). In striking contrast, the mouse gene, which is structurally similar to the human gene, requires 500 bases of the upstream region for full activity (12); several domains that function specifically in hepatic cells and that interact with hepatocyte-enriched DNA binding proteins have been identified within this region (7, 12). Thus, liver-specific expression of the AT gene is maintained by different mechanisms in the two species, reflecting the accumulation of regulatory mutations since the two species diverged.With one known exception, all mammals express AT predominantly in the liver. The wild-derived mouse species Mus caroli, in sharp contrast to laboratory mice (M. domesticus), transcribes the AT gene in the kidneys as well as the liver (2,19,24). Renal AT expression in M. caroli is specific to tubule cells, where it is regulated by androgens during development (19). This species differe...

Multiple murine alpha 1-protease inhibitor genes show unusual evolutionary divergence.

“…Although it is difficult to imagine what selective advantage diversity confers, it has been proposed that mice may have evolved multiple protease inhibitor genes in response to pathogen-produced proteases (16). However, clearly not all species are subjected to the same selective pressures since at least one species of mouse has been shown to have only a single al-PI gene (15). An alternative explanation may be that the apparently recent duplication of the a1-PI genes in laboratory mice was disadvantageous because it effectively increased the level of antiprotease activity.…”

“…However, all Mus musculus strains contain multiple a,-PI genes (8,14,15). Hill et al (13,16) have proposed that the serine protease inhibitor genes of mice and humans have undergone an unusual evolutionary selection process at their reactive sites to produce antiprotease activities at higher rates than would be expected based on random mutation and selection of the a,-PI genes.…”

Characterization of a thyroid-specific enhancer located 5.5 kilobase pairs upstream of the human thyroid peroxidase gene.

“…Mechanisms governing tissue-or developmental stage-specific gene regulation have been extensively studied in liver-or muscle-derived cell lines (4,29,34). In the case of thyroid tissue, however, little is known about the mechanisms by which thyroid-specific genes are controlled.…”

Evolution of murine ?1-proteinase inhibitors: Gene amplification and reactive center divergence

“…To obtain a rough estimate of cqPI gene copy number in the species M. saxicola, which is derived from a lineage that separated early in Mus evolution (Tseng-Crank and Berger 1987;Jouvin-Marche et al 1988), genomic DNAs were digested with EcoRI and analyzed by Southern blotting. The probe, a 503-bp PstI cDNA fragment (see Materials and Methods), spans a single EcoRI site within the M. domesticus and M. caroli alPI genes and should hybridize to two DNA fragments totaling a length of about 9 kb (Rheaume et al 1988;Latimer et al 1990). As seen in Fig.…”

“…In contrast, M. caroli contains two lightintensity bands that represent a total of about 9 kb of DNA. Since it is known that M. domesticus contains five alPI genes (Hill et al 1985;Krauter et al 1986;Boriello andKrauter 1990, 1991), and M. caroli contains only one (Latimer et al 1990), we estimate that there are approximately three to five genes in M. saxicola. The presence of multiple OtlPI genes in both M. domesticus and M. saxicola, which separated from each other 9-10 Mya (TsengCrank and Berger 1987;Jouvin-Marche et al 1988), suggests that amplification of the gene may have occurred early in the evolution of the Mus genus, leading to the fixation of multigene families in separate species.…”