Targeted Disruption of the Meprin β Gene in Mice Leads to Underrepresentation of Knockout Mice and Changes in Renal Gene Expression Profiles

Abstract

Meprins are multidomain zinc metalloproteases that are highly expressed in mammalian kidney and intestinal brush border membranes and in leukocytes and certain cancer cells. Mature meprins are oligomers of evolutionarily related, separately encoded ␣ and/or ␤ subunits. Homooligomers of meprin ␣ are secreted; oligomers containing meprin ␤ are plasma membrane associated. Meprin substrates include bioactive peptides and extracellular matrix proteins. Meprins have been implicated in cancer and intestinal inflammation. Additionally, meprin ␤ is a candidate gene for diabetic nephropathy. To elucidate in vivo functions of these metalloproteases, meprin ␤ null mice were generated by targeted disruption of the meprin ␤ gene on mouse chromosome 18q12. Analyses of meprin ␤ knockout mice indicated that (i) 50% fewer null mice are born than the Mendelian distribution predicts, (ii) null mice that survive develop normally and are viable and fertile, (iii) meprin ␤ knockout mice lack membrane-associated meprin ␣ in kidney and intestine, and (iv) null mice have changes in renal gene expression profiles compared to wild-type mice as assessed by microarray analyses. Thus, disruption of the meprin ␤ allele in mice affects embryonic viability, birth weight, renal gene expression profiles, and the distribution of meprin ␣ in kidney and intestine.Meprins (meprin A [EC 3.4.24.18] and meprin B [EC 3.4.24.63]), metalloproteases of the astacin family, comprise approximately 5% of kidney brush border membrane protein in mice (2, 3). They are also expressed in intestinal brush border membranes, in leukocytes, and in certain epithelial cancer cells (17,21). Various growth factors, cytokines, and extracellular matrix proteins are substrates for meprin in vitro; among the best substrates are gastrin, gastrin-releasing peptide, monocyte chemoattractant protein-1, osteopontin, and laminin (1, 15). The nature of these substrates and the expression patterns of the subunits in acute renal failure, intestinal disease, and cancerous cells implicate meprins in the regulation of growth, inflammation, cancer cell metastasis, and matrix remodeling (5,15,17,18,21,33).The meprin subunits, ␣ and ␤, associate to form homo-or heterooligomers (19). Although the subunits are encoded by distinct genes on different chromosomes (Mep1␣ on chromosome 6 in humans and 17 in mice and Mep1␤ on chromosome 18 in both species), under some circumstances there is coordinate regulation of the expression of the genes that results in heterooligomeric protein complexes (4). Meprin ␣ and ␤ subunits are 42% identical at the amino acid level and share the same domain structure, except that meprin ␣ contains an inserted (I) domain which is not present in meprin ␤ (20). The I domain directs the proteolytic cleavage of meprin ␣ during maturation in the endoplasmic reticulum so that the bulk of the subunit is released from the membrane. The consequence of the cleavage is that meprin ␣ is secreted unless it interacts with membrane-associated meprin ␤. In vivo, meprin ␣ is secreted as ...

Deletion of the Mouse Meprin β Metalloprotease Gene Diminishes the Ability of Leukocytes to Disseminate through Extracellular Matrix

“…In fact, meprin ␣ is implicated in the cellular transmigration events of metastatic colon cancer cells, because these cell lines selectively express meprin ␣, and nonmetastatic colon cancer cells are meprin ␣ Ϫ (G. Matters, unpublished observations). Interestingly, in mice, active meprin ␣ is localized with meprin ␤ to the apical surface of the proximal tubular epithelium, while the secreted form of meprin ␣ is inactive and forms meprin ␣ homo-oligomers (5,19). Whatever the mechanism, it can be concluded that meprin ␤ is essential to leukocyte infiltration, at least in vitro in the mesenteric lymph node.…”

“…Meprin ␤ null and wild-type mice were generated on a C57BL/6:129/SvJ background and characterized (19). Leukocytes from the mesenteric lymph nodes from 3-mo-old meprin ␤ null mice and strain/age-matched wild-type mice were isolated, as described above.…”

Long-term survival following a single treatment of kidney tumors with multiwalled carbon nanotubes and near-infrared radiation

“…Along with the phenotype of Mep1a −/− and Mep1b −/− mice at the histological level, the observed reduced levels of procollagen I conversion in skin and primary fibroblasts isolated from meprin KO mice provide further evidence that meprin α and meprin β exhibit PCP activity in vivo. Interestingly, significantly fewer Mep1b −/− mice (∼50%) are born compared with WT mice, indicating essential functions of this protease already during early embryonic development (43). Moreover, morpholino-induced meprin knockdown in zebrafish embryos revealed severe defects in organogenesis and tail morphology, which also might be linked to reduced collagen fibril assembly in these animals (15).…”

Ectodomain shedding of CD99 within highly conserved regions is mediated by the metalloprotease meprin β and promotes transendothelial cell migration

“…The generation of Mep1b 2/2 mice has been described in Norman et al (33). Mice were kept in specific pathogen-free conditions in isolated ventilated cages, with a 12-h light-dark cycle and food and water provided ad libitum.…”