Abstract. Ksp-cadherin is a unique, tissue-specific member of the cadherin family of cell adhesion molecules that is expressed in tubular epithelial cells in the kidney and developing genitourinary (GU) tract. It has recently been shown that a 1341-bp fragment of the 5' flanking region containing the Ksp-cadherin gene promoter can recapitulate the complete expression pattern of the gene in the developing kidney and GU tract. Similar to the endogenous Ksp-cadherin gene, transgenes containing 1341 bp of the 5' flanking region are expressed in developing nephrons, ureteric bud, mesonephric tubules, Wolffian duct, and Müllerian duct. In adult mice, the expression is restricted to renal tubules. In the current study, Ksp1.3/Cre transgenic mice carrying 1329 bp of the Kspcadherin 5' flanking region linked to the Cre recombinase gene were produced. Adult transgenic mice expressed Cre recombinase in renal tubules, especially collecting ducts and thick ascending limbs of Henle's loops. Transgenic embryos expressed Cre recombinase in the branching ureteric bud, developing renal tubules, and sex ducts. Ksp1.3/Cre transgenic mice were crossed with mice carrying a lacZ reporter gene that is activated by Cre/lox-mediated genetic recombination. Bitransgenic progeny expressed lacZ exclusively in renal tubules, mesonephric tubules, ureteric bud, developing ureter, and Wolffian duct. These results demonstrate that Ksp1.3/Cre transgenic mice express Cre recombinase exclusively in the kidney and developing GU tract and can mediate epithelialspecific Cre/lox recombination in these tissues. Ksp1.3/Cre transgenic mice should be useful for cell lineage studies and kidney-specific gene targeting.Cre recombinase (cyclization recombination) is a site-specific DNA recombinase from bacteriophage P1 that mediates genetic recombination at specific 34-bp recognition sites (called loxP) without any requirement for accessory proteins or highenergy cofactors (1). Cre-mediated intramolecular recombination between two directly repeated loxP sites flanking a DNA segment results in deletion of the intervening segment, leaving a single loxP site. If two loxP sites are introduced into the genomic DNA flanking an essential exon

Mutations in cystic kidney disease genes represent a major genetic cause of end-stage renal disease. However, the molecular cascades controlling the expression of these genes are still poorly understood. Hepatocyte Nuclear Factor 1beta (HNF1beta) is a homeoprotein predominantly expressed in renal, pancreatic and hepatic epithelia. We report here that mice with renal-specific inactivation of HNF1beta develop polycystic kidney disease. We show that renal cyst formation is accompanied by a drastic defect in the transcriptional activation of Umod, Pkhd1 and Pkd2 genes, whose mutations are responsible for distinct cystic kidney syndromes. In vivo chromatin immunoprecipitation experiments demonstrated that HNF1beta binds to several DNA elements in murine Umod, Pkhd1, Pkd2 and Tg737/Polaris genomic sequences. Our results uncover a direct transcriptional hierarchy between HNF1beta and cystic disease genes. Interestingly, most of the identified HNF1beta target gene products colocalize to the primary cilium, a crucial organelle that plays an important role in controlling the proliferation of tubular cells. This may explain the increased proliferation of cystic cells in MODY5 patients carrying autosomal dominant mutations in HNF1beta.

Smad2 and Smad3 interact and mediate TGF-␤ signaling. Although Smad3 promotes fibrosis, the role of Smad2 in fibrogenesis is largely unknown. In this study, conditional deletion of Smad2 from the kidney tubular epithelial cells markedly enhanced fibrosis in response to unilateral ureteral obstruction. In vitro, Smad2 knockdown in tubular epithelial cells increased expression of collagen I, collagen III, and TIMP-1 and decreased expression of the matrix-degrading enzyme MMP-2 in response to TGF-␤1 compared with similarly treated wild-type cells. We obtained similar results in Smad2-knockout fibroblasts. Mechanistically, Smad2 deletion promoted fibrosis through enhanced TGF-␤/Smad3 signaling, evidenced by greater Smad3 phosphorylation, nuclear translocation, promoter activity, and binding of Smad3 to a collagen promoter (COL1A2). Moreover, deletion of Smad2 increased autoinduction of TGF-␤1. Conversely, overexpression of Smad2 attenuated TGF-␤1-induced Smad3 phosphorylation and collagen I matrix expression in tubular epithelial cells. In conclusion, in contrast to Smad3, Smad2 protects against TGF-␤-mediated fibrosis by counteracting TGF-␤/Smad3 signaling.

Polycystic kidney disease (PKD) is an inherited disorder that is characterized by the accumulation of cysts in the renal parenchyma and progressive decline in renal function. Recent studies suggest that PKD arises from abnormalities of the primary cilium. We have previously shown that kidney-specific inactivation of the ciliogenic gene Kif3a during embryonic development produces kidney cysts and renal failure. Here, we used tamoxifen-inducible, kidney-specific gene targeting to inactivate Kif3a in the postnatal mouse kidney. Kidney-specific inactivation of Kif3a in newborn mice resulted in the loss of primary cilia and produced kidney cysts primarily in the loops of Henle, whereas inactivation in adult mice did not lead to the rapid development of cysts despite a comparable loss of primary cilia. The age-dependence and locations of the cysts suggested that cyst formation required increased rates of cell proliferation. To test this possibility, we stimulated cell proliferation in the adult kidney by inducing acute kidney injury and tubular regeneration. Acute kidney injury induced cyst formation in adult Kif3a mutant mice. Analysis of pre-cystic tubules in Kif3a mutant mice showed that the loss of cilia did not stimulate cell proliferation but instead resulted in aberrant planar cell polarity as manifested by abnormalities in the orientation of cell division. We conclude that primary cilia are required for the maintenance of planar cell polarity in the mammalian kidney and that acute kidney injury exacerbates cystic disease.

SUMMARY Mutations in 1-acylglycerol-3-phosphate-O-acyltransferase 2 (AGPAT2) cause congenital generalized lipodystrophy. To understand the molecular mechanisms underlying the metabolic complications associated with AGPAT2 deficiency, Agpat2 null mice were generated. Agpat2−/− mice develop severe lipodystrophy affecting both white and brown adipose tissue, severe insulin resistance, diabetes, and hepatic steatosis. The expression of lipogenic genes and rates of de novo fatty acid biosynthesis were increased ~4-fold in Agpat2−/− mouse livers. The mRNA and protein levels of monoacylglycerol acyltransferase isoform 1 were markedly increased in the livers of Agpat2−/− mice suggesting that the alternative monoacylglycerol pathway for triglyceride biosynthesis is activated in the absence of AGPAT2. Feeding a fat-free diet reduced liver triglycerides by ~50% in Agpat2−/− mice. These observations suggest that both dietary fat and hepatic triglyceride biosynthesis via a novel monoacylglycerol pathway may contribute to hepatic steatosis in Agpat2−/− mice.