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Peter Igarashi

Publications and source records attributed to Peter Igarashi.

At least 19 recordsLinked to original sources
PubMed2006

Proteolytic cleavage and nuclear translocation of fibrocystin is regulated by intracellular Ca2+ and activation of protein kinase C.

Fibrocystin, a type I membrane protein of unknown function, is the protein affected in the autosomal recessive form of polycystic kidney disease. Here we show that fibrocystin undergoes regulated proteolysis. Several proteolytic cleavages occur within the predicted ectodomain, whereas at least one cleavage occurs within the cytoplasmic portion. The latter generates a C-terminal intracellular fragment that harbors the nuclear localization signal KRKVSRLAVTGERTATPAPKIPRIT and translocates to the nucleus. Proteolytic cleavage of fibrocystin occurs constitutively in long term cultures of polarized inner medullary collecting duct cells (mIMCD-3). Activation of protein kinase C and release of intracellular Ca2+ are required for proteolysis under these conditions. In short term cultures of human embryonic kidney 293 cells (HEK-293), proteolytic cleavage of fibrocystin can be elicited by stimulation of intracellular Ca2+ release or activation of protein kinase C. These results identify a novel Ca2+-dependent pathway that signals from fibrocystin located in the cell membrane to the nucleus.

Amino Acid Sequence↗
PubMed2006

Expression of the basolateral Na-K-Cl cotransporter during mouse nephrogenesis and embryonic development.

We examined the expression of Slc12a2 (NKCC1) transcripts in the developing mouse by Northern blot analysis and in situ hybridization (ISH) using riboprobes transcribed from a cDNA encoding the transmembrane domain of human Slc12a2. In developing kidney, the 7.5-kb Slc12a2 transcript was expressed at all stages examined (13.5 d.p.c. to adult) but was more abundant in immature metanephroi. ISH revealed that NKCC1 was expressed in both mesenchymal cells and early nephric structures, but not branching ureteric buds, of developing metanephroi. A marked increase in expression was observed in the endocapillary cells of capillary loop stage glomeruli, and high expression was observed in the glomeruli of more mature nephrons. This was in contrast to Slc12a1 (NKCC2), where expression was excluded from the glomerulus. Extra-renal expression of Slc12a2 was examined in 13.5, 15.5, and 16.5 d.p.c. mouse embryos. Slc12a2 was highly expressed in the developing lung, gut, submandibular gland, tooth bud, and nasal epithelium. Slc12a2 expression was also observed in the developing central and peripheral nervous systems, including choroid plexus and trigeminal and dorsal root ganglia.

Animals↗
PubMed2005

Role of the hepatocyte nuclear factor-1beta (HNF-1beta) C-terminal domain in Pkhd1 (ARPKD) gene transcription and renal cystogenesis.

Hepatocyte nuclear factor-1beta (HNF-1beta) is a homeodomain-containing transcription factor that regulates tissue-specific gene expression in the kidney and other epithelial organs. Mutations of HNF-1beta produce congenital cystic abnormalities of the kidney, and previous studies showed that HNF-1beta regulates the expression of the autosomal recessive polycystic kidney disease (ARPKD) gene, Pkhd1. Here we show that the C-terminal region of HNF-1beta contains an activation domain that is functional when fused to a heterologous DNA-binding domain. An HNF-1beta deletion mutant lacking the C-terminal domain interacts with wild-type HNF-1beta, binds DNA, and functions as a dominant-negative inhibitor of a chromosomally integrated Pkhd1 promoter. The activation of the Pkhd1 promoter by wild-type HNF-1beta is stimulated by sodium butyrate or coactivators CREB (cAMP-response element)-binding protein (CBP) and P/CAF. The interaction with CBP and P/CAF requires the C-terminal domain. Expression of an HNF-1beta C-terminal deletion mutant in transgenic mice produces renal cysts, increased cell proliferation, and dilatation of the ureter similar to mice with kidney-specific inactivation of HNF-1beta. Pkhd1 expression is inhibited in cystic collecting ducts but not in non-cystic proximal tubules, despite transgene expression in this nephron segment. We conclude that the C-terminal domain of HNF-1beta is required for the activation of the Pkhd1 promoter. Deletion mutants lacking the C-terminal domain function as dominant-negative mutants, possibly by preventing the recruitment of histone acetylases to the promoter. Cyst formation correlates with inhibition of Pkhd1 expression, which argues that mutations of HNF-1beta produce kidney cysts by down-regulating the ARPKD gene, Pkhd1. Expression of HNF-1alpha in proximal tubules may protect against cystogenesis.

Acetyltransferases↗
PubMed2005

Elucidating the function of primary cilia by conditional gene inactivation.

PURPOSE OF REVIEW: This review discusses recent experimental approaches to determine the function of primary cilia by conditional inactivation of genes crucial for cilia formation. RECENT FINDINGS: A functional role in the sensing of fluid flow was recently assigned to the primary cilia. This discovery shed light onto how cells sense dynamic fluid movements. Conditional inactivation of primary cilia formation in later ontogenic stages demonstrated the crucial role renal primary cilia play in the control of cell proliferation. SUMMARY: Primary cilia can act as flow sensors, transmitting signals by means of calcium influx into the cells. Structures based on primary cilia are also crucial for the function of photoreceptor cells and it can be expected that additional functions of these organelles will be determined in the future. An important experimental approach to elucidate the involvement of primary cilia in other physiological processes is to specifically inactivate genes crucial for formation of primary cilia. Morphological and physiological changes induced by the loss of primary cilia will help determine additional roles primary cilia play in physiology and organ development.

Animals↗
PubMed2005

Roles of HNF-1beta in kidney development and congenital cystic diseases.

Hepatocyte nuclear factor-1beta (HNF-1beta) is a Pit-1/Oct-1/Unc-86 (POU)/homeodomain-containing transcription factor that regulates tissue-specific gene expression in the kidney, liver, pancreas, and other epithelial organs. Mutations of HNF-1beta produce maturity-onset diabetes of the young type 5 (MODY5) and are associated with congenital cystic abnormalities of the kidney. Transgenic mice expressing mutant HNF-1beta under the control of a kidney-specific promoter develop kidney cysts and renal failure, which is similar to the phenotype of humans with MODY5. Similarly, kidney-specific deletion of HNF-1beta using Cre/loxP recombination results in renal cyst formation. HNF-1beta directly regulates the Pkhd1 promoter. HNF-1beta mutant mice show decreased expression of Pkhd1, the gene that is mutated in humans with autosomal-recessive polycystic kidney disease (ARPKD). These studies demonstrate that HNF-1beta is required for the development of the mammalian kidney. They establish a previously unrecognized link between two renal cystic diseases, MODY5 and ARPKD, and suggest that the mechanism of cyst formation in humans with mutations of HNF-1beta involves down-regulation of PKHD1 gene transcription.

Animals↗
PubMed2005

Intrarenal cells, not bone marrow-derived cells, are the major source for regeneration in postischemic kidney.

Ischemic injury to the kidney produces acute tubular necrosis and apoptosis followed by tubular regeneration and recovery of renal function. Although mitotic cells are present in the tubules of postischemic kidneys, the origins of the proliferating cells are not known. Bone marrow cells (BMCs) can differentiate across lineages to repair injured organs, including the kidney. However, the relative contribution of intrarenal cells and extrarenal cells to kidney regeneration is not clear. We produced transgenic mice that expressed enhanced GFP (EGFP) specifically and permanently in mature renal tubular epithelial cells. Following ischemia/reperfusion injury (IRI), EGFP-positive cells showed incorporation of BrdU and expression of vimentin, which provides direct evidence that the cells composing regenerating tubules are derived from renal tubular epithelial cells. In BMC-transplanted mice, 89% of proliferating epithelial cells originated from host cells, and 11% originated from donor BMCs. Twenty-eight days after IRI, the kidneys contained 8% donor-derived cells, of which 8.4% were epithelial cells, 10.6% were glomerular cells, and 81% were interstitial cells. No renal functional improvement was observed in mice that were transplanted with exogenous BMCs. These results show that intrarenal cells are the main source of renal repair, and a single injection of BMCs does not make a significant contribution to renal functional or structural recovery.

Animals↗
PubMed2004

Cystic renal neoplasia following conditional inactivation of apc in mouse renal tubular epithelium.

Alterations in Wnt/beta-catenin signaling have been linked to abnormal kidney development and tumorigenesis. To gain more insights into the effects of these alterations, we created mice carrying a conditional deletion of the Apc tumor suppressor gene specifically in the renal epithelium. As expected, the loss of Apc leads to increased levels of beta-catenin protein in renal epithelium. Most of these mice die shortly after birth, and multiple kidney cysts were found upon histological examination. Only rarely did these animals survive to adulthood. Analysis of these adults revealed severely cystic kidneys associated with the presence of renal adenomas. Our results confirm an important role for proper regulation of Wnt/beta-catenin signaling in renal development and provide evidence that dysregulation of the pathway can initiate tumorigenesis in the kidney.

Adenoma↗
PubMed2004

A transcriptional network in polycystic kidney disease.

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.

Animals↗
PubMed2004

Loss of NFAT5 results in renal atrophy and lack of tonicity-responsive gene expression.

The transcription factor NFAT5/TonEBP, a member of the NFAT/Rel family of transcription factors, has been implicated in diverse cellular responses, including the response to osmotic stress, integrin-dependent cell migration, T cell activation, and the Ras pathway in Drosophila. To clarify the in vivo role of NFAT5, we generated NFAT5-null mice. Homozygous mutants were genetically underrepresented after embryonic day 14.5. Surviving mice manifested a progressive and profound atrophy of the kidney medulla with impaired activation of several osmoprotective genes, including those encoding aldose reductase, Na+/Cl--coupled betaine/gamma-aminobutyric acid transporter, and the Na+/myo-inositol cotransporter. The aldose reductase gene is controlled by a tonicity-responsive enhancer, which was refractory to hypertonic stress in fibroblasts lacking NFAT5, establishing this enhancer as a direct transcriptional target of NFAT5. Our findings demonstrate a central role for NFAT5 as a tonicity-responsive transcription factor required for kidney homeostasis and function.

Animals↗
PubMed2004

Mutation of hepatocyte nuclear factor-1beta inhibits Pkhd1 gene expression and produces renal cysts in mice.

Hepatocyte nuclear factor-1beta (HNF-1beta) is a Pit-1, Oct-1/2, UNC-86 (POU)/homeodomain-containing transcription factor that regulates tissue-specific gene expression in the liver, kidney, and other organs. Humans with autosomal dominant mutations of HNF-1beta develop maturity-onset diabetes of the young type 5 (MODY5) and congenital cystic abnormalities of the kidney. Autosomal recessive polycystic kidney disease (ARPKD) is an inherited cystic disorder that produces renal failure in infants and children and is caused by mutations of PKHD1. The proximal promoter of the mouse Pkhd1 gene contains an evolutionarily conserved HNF-1-binding site that is located near a region of deoxyribonuclease hypersensitivity. HNF-1beta and the structurally related HNF-1alpha bind specifically to the Pkhd1 promoter and stimulate gene transcription. Mutations of the HNF-1 site or expression of a dominant-negative HNF-1beta mutant inhibit Pkhd1 promoter activity in transfected cells. Transgenic mice expressing a dominant-negative HNF-1beta mutant under the control of a kidney-specific promoter develop renal cysts, similarly to humans with MODY5. Pkhd1 transcripts are absent in the cells lining the cysts but are present in morphologically normal surrounding tubules. These studies identify a link between two cystic disease genes, HNF1beta (MODY5) and PKHD1 (ARPKD). HNF-1beta directly regulates the transcription of Pkhd1, and inhibition of PKHD1 gene expression may contribute to the formation of renal cysts in humans with MODY5.

Animals↗
PubMed2004

Mechanical stimuli induce cleavage and nuclear translocation of the polycystin-1 C terminus.

Polycystin-1, which is encoded by a gene that is mutated in autosomal dominant polycystic kidney disease (ADPKD), is involved in cell-matrix interactions as well as in ciliary signaling. The precise mechanisms by which it functions, however, remain unclear. Here we find that polycystin-1 undergoes a proteolytic cleavage that releases its C-terminal tail (CTT), which enters the nucleus and initiates signaling processes. The cleavage occurs in vivo in association with alterations in mechanical stimuli. Polycystin-2, the product of the second gene mutated in ADPKD, modulates the signaling properties of the polycystin-1 CTT. These data reveal a novel pathway by which polycystin-1 transmits messages directly to the nucleus.

Amino Acid Sequence↗
PubMed2003

Kidney-specific inactivation of the KIF3A subunit of kinesin-II inhibits renal ciliogenesis and produces polycystic kidney disease.

Polycystic kidney disease (PKD) is the most common genetic cause of renal failure in humans. Several proteins that are encoded by genes associated with PKD have recently been identified in primary cilia in renal tubular epithelia. These findings have suggested that abnormalities in cilia formation and function may play a role in the pathogenesis of PKD. To directly determine whether cilia are essential to maintain tubular integrity, we conditionally inactivated KIF3A, a subunit of kinesin-II that is essential for cilia formation, in renal epithelia. Constitutive inactivation of KIF3A produces abnormalities of left-right axis determination and embryonic lethality. Here we show that tissue-specific inactivation of KIF3A in renal tubular epithelial cells results in viable offspring with normal-appearing kidneys at birth. Cysts begin to develop in the kidney at postnatal day 5 and cause renal failure by postnatal day 21. The cyst epithelial cells lack primary cilia and exhibit increased proliferation and apoptosis, apical mislocalization of the epidermal growth factor receptor, increased expression of beta-catenin and c-Myc, and inhibition of p21(CIP1). These results demonstrate that the absence of renal cilia produces both the clinical and cell biological findings associated with PKD. Most generally, the phenotype of Kif3a mutant mice suggests a role for primary cilia in the maintenance of lumen-forming epithelial differentiation.

Animals↗
PubMed2003

Hematopoietic stem cells contribute to the regeneration of renal tubules after renal ischemia-reperfusion injury in mice.

Ischemia-reperfusion injury (I/R injury) is a common cause of acute renal failure. Recovery from I/R injury requires renal tubular regeneration. Hematopoietic stem cells (HSC) have been shown to be capable of differentiating into hepatocytes, cardiac myocytes, gastrointestinal epithelial cells, and vascular endothelial cells during tissue repair. The current study tested the hypothesis that murine HSC can contribute to the regeneration of renal tubular epithelial cells after I/R injury. HSC isolated from male Rosa26 mice that express beta-galactosidase constitutively were transplanted into female nontransgenic mice after unilateral renal I/R injury. Four weeks after HSC transplantation, beta-galactosidase-positive cells were detected in renal tubules of the recipients by X-Gal staining. PCR analysis of the male-specific Sry gene and Y chromosome fluorescence in situ hybridization confirmed the presence of male-derived cells in the kidneys of female recipients. Antibody co-staining showed that beta-galactosidase was primarily expressed in renal proximal tubules. This is the first report to show that HSC can differentiate into renal tubular cells after I/R injury. Because of their availability, HSC may be useful for cell replacement therapy of acute renal failure.

Animals↗
PubMed2003

Following the expression of a kidney-specific gene from early development to adulthood.

Ksp-cadherin (Cadherin 16) is a unique, tissue-specific member of the cadherin family of cell adhesion proteins that is expressed exclusively in tubular epithelial cells in the adult kidney and developing genitourinary (GU) tract. Since its original identification in 1995, Ksp-cadherin has proven to be a useful experimental model for studies of kidney-specific and developmental gene regulation. These investigations have led to the creation of new strains of transgenic mice that will be useful for studies of renal cell lineage and tissue-specific gene targeting. Recent studies have also identified Ksp-cadherin as the first kidney-specific promoter that it is regulated by HNF-1beta (hepatocyte nuclear factor-1beta), a transcription factor that has been implicated in human renal cystic disease.

Animals↗
PubMed2002

Deregulated expression of the homeobox gene Cux-1 in transgenic mice results in downregulation of p27(kip1) expression during nephrogenesis, glomerular abnormalities, and multiorgan hyperplasia.

Cux-1 is a murine homeobox gene that is highly expressed in the developing kidney with expression restricted to the nephrogenic zone. Cux-1 is highly expressed in cyst epithelium of polycystic kidneys from C57BL/6J-cpk/cpk mice, but not in kidneys isolated from age-matched phenotypically normal littermates. To further elucidate the role of Cux-1 in renal development, we generated transgenic mice expressing Cux-1 under the control of the CMV immediate early gene promoter. Mice constitutively expressing Cux-1 developed multiorgan hyperplasia and organomegaly, but not an overall increase in body size. Transgenic kidneys were enlarged 50% by 6 weeks of age, with the increased growth primarily restricted to the cortex. Proliferating cells were found in proximal and distal tubule epithelium throughout the cortex, and the squamous epithelium that normally lines Bowman's capsule was replaced with proximal tubule epithelium. However, the total number of nephrons was not increased. In the developing kidneys of transgenic mice, Cux-1 was ectopically expressed in more highly differentiated tubules and glomeruli, and this was associated with reduced expression of the cyclin kinase inhibitor, p27. Transient transfection experiments revealed that Cux-1 is an inhibitor of p27 promoter activity. These results suggest that Cux-1 regulates cell proliferation during early nephrogenesis by inhibiting expression of p27.

Animals↗