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Jing Zhou - One of the best experts on this subject based on the ideXlab platform.
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retromer associates with the cytoplasmic amino terminus of polycystin 2
Journal of Cell Science, 2018Co-Authors: Frances C Tilley, Jing Zhou, Matthew Gallon, Chong Luo, Chris M Danson, Peter J CullenAbstract:Autosomal dominant polycystic kidney disease (ADPKD) is the most common monogenic human disease, with around 12.5 million people affected worldwide. ADPKD results from mutations in either PKD1 or PKD2, which encode the atypical G-protein coupled receptor polycystin-1 (PC1) and the transient receptor potential channel polycystin-2 (PC2), respectively. Although altered intracellular trafficking of PC1 and PC2 is an underlying feature of ADPKD, the mechanisms which govern vesicular transport of the Polycystins through the biosynthetic and endosomal membrane networks remain to be fully elucidated. Here, we describe an interaction between PC2 and retromer, a master controller for the sorting of integral membrane proteins through the endo-lysosomal network. We show that association of PC2 with retromer occurs via a region in the PC2 cytoplasmic amino-terminal domain, independently of the retromer-binding Wiskott-Aldrich syndrome and scar homologue (WASH) complex. Based on observations that retromer preferentially interacts with a trafficking population of PC2, and that ciliary levels of PC1 are reduced upon mutation of key residues required for retromer association in PC2, our data are consistent with the identification of PC2 as a retromer cargo protein.This article has an associated First Person interview with the first author of the paper.
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Loss of Polycystin-1 in Human Cyst-Lining Epithelia Leads
2015Co-Authors: To Ciliary Dysfunction, Jing ZhouAbstract:A “two-hit ” hypothesis predicts a second somatic hit, in addition to the germline mutation, as a prerequisite to cystogenesis and has been proposed to explain the focal nature for renal cyst formation in autosomal dominant polycystic kidney disease (ADPKD). It was reported previously that Pkd1null/nullmouse kidney epithelial cells are unresponsive to flow stimulation. This report shows that Pkd1/null cells are capable of responding to mechanical flow stimulation by changing their intracellular calcium concentration in a manner similar to that of wild-type cells. This paper reports that human renal epithelia require a higher level of shear stress to evoke a cytosolic calcium increase than do mouse renal epithelia. Both immortalized and primary cultured renal epithelial cells that originate from normal and nondilated ADPKD human kidney tubules display normal ciliary expression of the Polycystins and respond to fluid-flow shear stress with the typical change in cytosolic calcium. In contrast, immortalized and primary cultured cyst-lining epithelial cells from ADPKD patients with mutations in PKD1 or with abnormal ciliary expression of polycystin-1 or-2 were not responsive to fluid shear stress. These data support a two-hit hypothesis as a mechanism of cystogenesis. This report proposes that calcium response to fluid-flow shear stress can be used as a readout of polycystin function and that loss of mechanosensation in the renal tubular epithelia is a feature of PKD cysts. J Am Soc Nephrol 17: 1015–1025, 2006. doi: 10.1681/ASN.2005080830 A gene mutation can result in disease through direct orindirect mechanisms. For instance, in the gain-of-function mutation, a germline mutant allele confers new or enhanced protein activity with a pathologic function
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Polycystins and primary cilia primers for cell cycle progression
Annual Review of Physiology, 2009Co-Authors: Jing ZhouAbstract:Polycystins are a family of eight-transmembrane proteins united by sequence homology. The name stems from the identification of mutations in genes encoding polycystin-1 and -2 in polycystic kidney diseases. This review discusses recent topics in polycystin research, with a focus on the role of polycystin-1 and polycystin-2 in primary cilia and the cell cycle. Polycystins appear to play key roles during development, but a major question is their function in mature organs. Their roles in primary cilia, shear stress sensation, alteration of intracellular calcium, and planar cell polarity are examined. The third-hit hypothesis of polycystic kidney disease is discussed.
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Nek8 Regulates the Expression and Localization of Polycystin-1 and Polycystin-2
Journal of the American Society of Nephrology : JASN, 2008Co-Authors: Eisei Sohara, Ying Luo, Jingjing Zhang, Danielle K. Manning, David R. Beier, Jing ZhouAbstract:Nek8 is a serine/threonine kinase that is mutated in the jck (juvenile cystic kidneys) mouse, a model of autosomal recessive juvenile polycystic kidney disease, but its function is poorly understood. We used the jck mouse to study the functional relationship between Nek8 and other proteins that have been implicated in polycystic kidney diseases. In the collecting tubules and collecting ducts of wild-type mice, we found that Nek8 was localized to the proximal portion of primary cilia and was weakly detected in the cytosol. In the jck mutant, however, Nek8 was found along the entire length of cilia. Coimmunoprecipitation experiments demonstrated that Nek8 interacted with polycystin-2, but not with polycystin-1, and that the jck mutation did not affect this interaction. Western blot analysis and real-time reverse transcriptase PCR revealed that the protein and mRNA expression of polycystin-1 (PC1) and polycystin-2 (PC2) were increased in jck mouse kidneys. The jck mutation also led to abnormal phosphorylatin of PC2, and this was associated with longer cilia and ciliary accumulation of PC1 and PC2. Our data suggests that Nek8 interacts with the signal transduction pathways of the Polycystins and may control the targeting of these ciliary proteins. Dysfunction Nek8 may lead to cystogenesis by altering the structure and function of cilia in the distal nephron.
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fibrocystin polyductin found in the same protein complex with polycystin 2 regulates calcium responses in kidney epithelia
Molecular and Cellular Biology, 2007Co-Authors: Shixuan Wang, Ying Luo, Jingjing Zhang, Surya M Nauli, Patrick G Starremans, Kristina A Roberts, Jing ZhouAbstract:Recent evidence suggests that fibrocystin/polyductin (FPC), polycystin-1 (PC1), and polycystin-2 (PC2) are all localized at the plasma membrane and the primary cilium, where PC1 and PC2 contribute to fluid flow sensation and may function in the same mechanotransduction pathways. To further define the exact subcellular localization of FPC, the protein product encoded by the PKHD1 gene responsible for autosomal recessive polycystic kidney disease (PKD) in humans, and whether FPC has direct and/or indirect cross talk with PC2, which, in turn, is pivotal for the pathogenesis of autosomal dominant PKD, we performed double immunostaining and coimmunoprecipitation as well as a microfluorimetry study of kidney tubular epithelial cells. FPC and PC2 are found to completely or partially colocalize at the plasma membrane and the primary cilium and can be reciprocally coimmunoprecipitated. Although incomplete removal of FPC by small interfering RNA and antibody 803 to intracellular epitopes of FPC did not abolish flow-induced intracellular calcium responses, antibody 804 to extracellular epitopes of FPC blocked cellular calcium responses to flow stimulation. These findings suggest that FPC and Polycystins share, at least in part, a common mechanotransduction pathway. Inherited polycystic kidney diseases (PKD) are a large group of diseases characterized by the development of multiple fluid-filled cysts in the kidney, starting from infancy or adulthood, which gradually lead to end-stage renal disease (ESRD). About 10 percent of ESRD cases are caused by PKD. In humans, there are two major forms of PKD, i.e., autosomal dominant PKD (ADPKD)
Peter C Harris - One of the best experts on this subject based on the ideXlab platform.
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Regulation of polycystin expression, maturation and trafficking.
Cellular signalling, 2020Co-Authors: Peter C HarrisAbstract:The major autosomal dominant polycystic kidney disease (ADPKD) genes, PKD1 and PKD2, are wildly expressed at the organ and tissue level. PKD1 encodes polycystin 1 (PC1), a large membrane associated receptor-like protein that can complex with the PKD2 product, PC2. Various cellular locations have been described for both PC1, including the plasma membrane and extracellular vesicles, and PC2, especially the endoplasmic reticulum (ER), but compelling evidence indicates that the primary cilium, a sensory organelle, is the key site for the polycystin complex to prevent PKD. As with other membrane proteins, the ER biogenesis pathway is key to appropriately folding, performing quality control, and exporting fully folded PC1 to the Golgi apparatus. There is a requirement for binding with PC2 and cleavage of PC1 at the GPS for this folding and export to occur. Six different monogenic defects in this pathway lead to cystic disease development, with PC1 apparently particularly sensitive to defects in this general protein processing pathway. Trafficking of membrane proteins, and the Polycystins in particular, through the Golgi to the primary cilium have been analyzed in detail, but at this time, there is no clear consensus on a ciliary targeting sequence required to export proteins to the cilium. After transitioning though the trans-Golgi network, polycystin-bearing vesicles are likely sorted to early or recycling endosomes and then transported to the ciliary base, possibly via docking to transition fibers (TF). The membrane-bound polycystin complex then undergoes facilitated trafficking through the transition zone, the diffusion barrier at the base of the cilium, before entering the cilium. Intraflagellar transport (IFT) may be involved in moving the Polycystins along the cilia, but data also indicates other mechanisms. The ciliary polycystin complex can be ubiquitinated and removed from cilia by internalization at the ciliary base and may be sent back to the plasma membrane for recycling or to lysosomes for degradation. Monogenic defects in processes regulating the protein composition of cilia are associated with syndromic disorders involving many organ systems, reflecting the pleotropic role of cilia during development and for tissue maintenance. Many of these ciliopathies have renal involvement, likely because of faulty polycystin signaling from cilia. Understanding the expression, maturation and trafficking of the Polycystins helps understand PKD pathogenesis and suggests opportunities for therapeutic intervention.
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A polycystin-centric view of cyst formation and disease: the Polycystins revisited
Kidney international, 2015Co-Authors: Albert C.m. Ong, Peter C HarrisAbstract:It is 20 years since the identification of PKD1, the major gene mutated in autosomal dominant polycystic kidney disease (ADPKD), followed closely by the cloning of PKD2. These major breakthroughs have led in turn to a period of intense investigation into the function of the two proteins encoded, polycystin-1 and polycystin-2, and how defects in either protein lead to cyst formation and nonrenal phenotypes. In this review, we summarize the major findings in this area and present a current model of how the polycystin proteins function in health and disease.
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loss of polycystin 1 in human cyst lining epithelia leads to ciliary dysfunction
Journal of The American Society of Nephrology, 2006Co-Authors: Surya M Nauli, Peter C Harris, Sandro Rossetti, Robert J Kolb, Francis J Alenghat, Mark B Consugar, Donald E Ingber, Mahmoud Loghmanadham, Jing ZhouAbstract:A “two-hit” hypothesis predicts a second somatic hit, in addition to the germline mutation, as a prerequisite to cystogenesis and has been proposed to explain the focal nature for renal cyst formation in autosomal dominant polycystic kidney disease (ADPKD). It was reported previously that Pkd1 null/null mouse kidney epithelial cells are unresponsive to flow stimulation. This report shows that Pkd1 +/null cells are capable of responding to mechanical flow stimulation by changing their intracellular calcium concentration in a manner similar to that of wild-type cells. This paper reports that human renal epithelia require a higher level of shear stress to evoke a cytosolic calcium increase than do mouse renal epithelia. Both immortalized and primary cultured renal epithelial cells that originate from normal and nondilated ADPKD human kidney tubules display normal ciliary expression of the Polycystins and respond to fluid-flow shear stress with the typical change in cytosolic calcium. In contrast, immortalized and primary cultured cyst-lining epithelial cells from ADPKD patients with mutations in PKD1 or with abnormal ciliary expression of polycystin-1 or -2 were not responsive to fluid shear stress. These data support a two-hit hypothesis as a mechanism of cystogenesis. This report proposes that calcium response to fluid-flow shear stress can be used as a readout of polycystin function and that loss of mechanosensation in the renal tubular epithelia is a feature of PKD cysts.
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Analysis of the Polycystins in Aortic Vascular Smooth Muscle Cells
Journal of the American Society of Nephrology : JASN, 2003Co-Authors: Qi Qian, Stefan Somlo, Peter C Harris, Yiqiang Cai, Christopher J. Ward, Vicente E. TorresAbstract:ABSTRACT. The leading cause of death in autosomal dominant polycystic kidney disease (ADPKD) is cardiovascular. However, little is known about the pathogenesis of these manifestations. The present study was undertaken to characterize the ADPKD proteins, the Polycystins, in vascular smooth muscle cells. It was demonstrated that the expression of polycystin-1 is developmentally regulated, whereas polycystin-2 has a more constant level of expression. A polycystin-1 subpopulation was immunoprecipitated by polycystin-2, indicating an in vivo interaction of these two proteins. Analysis with glycosidase and cell surface biotinylation indicates that some polycystin-1 products, but not polycystin-2, are located on the plasma membrane. Immunofluorescence showed that most of the polycystin-1 and polycystin-2 was cytoplasmic but that persistent polycystin-1 staining was located in proximity to the cell surface after a Triton-X extraction, whereas no clear surface localization of polycystin-2 was detected. Immuno-gold electron microscopy revealed that polycystin-1 was localized at the plasma membrane and sarcoplasmic reticulum, whereas polycystin-2 was mainly located in the sarcoplasmic reticulum. Both Polycystins were found to be associated with dense plaques. These observations are consistent with an important role of the Polycystins in the development, maintenance, and function of the myoelastic arterial organization and with the vascular phenotype associated with ADPKD. E-mail: Qian.qi@mayo.edu
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Analysis of the Polycystins in Aortic Vascular Smooth Muscle Cells
Journal of the American Society of Nephrology : JASN, 2003Co-Authors: Qi Qian, Stefan Somlo, Peter C Harris, Yiqiang Cai, Christopher J. Ward, Vicente E. TorresAbstract:The leading cause of death in autosomal dominant polycystic kidney disease (ADPKD) is cardiovascular. However, little is known about the pathogenesis of these manifestations. The present study was undertaken to characterize the ADPKD proteins, the Polycystins, in vascular smooth muscle cells. It was demonstrated that the expression of polycystin-1 is developmentally regulated, whereas polycystin-2 has a more constant level of expression. A polycystin-1 subpopulation was immunoprecipitated by polycystin-2, indicating an in vivo interaction of these two proteins. Analysis with glycosidase and cell surface biotinylation indicates that some polycystin-1 products, but not polycystin-2, are located on the plasma membrane. Immunofluorescence showed that most of the polycystin-1 and polycystin-2 was cytoplasmic but that persistent polycystin-1 staining was located in proximity to the cell surface after a Triton-X extraction, whereas no clear surface localization of polycystin-2 was detected. Immuno-gold electron microscopy revealed that polycystin-1 was localized at the plasma membrane and sarcoplasmic reticulum, whereas polycystin-2 was mainly located in the sarcoplasmic reticulum. Both Polycystins were found to be associated with dense plaques. These observations are consistent with an important role of the Polycystins in the development, maintenance, and function of the myoelastic arterial organization and with the vascular phenotype associated with ADPKD.
Stefan Somlo - One of the best experts on this subject based on the ideXlab platform.
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the Polycystins are modulated by cellular oxygen sensing pathways and regulate mitochondrial function
Molecular Biology of the Cell, 2017Co-Authors: Valeria Padovano, Stefan Somlo, Ivana Y Kuo, Lindsey K Stavola, Hansrudolf Aerni, Benjamin J Flaherty, Hannah C Chapin, Alessandra Boletta, Barbara E. EhrlichAbstract:Autosomal dominant polycystic kidney disease is caused by mutations in the genes encoding polycystin-1 (PC1) and polycystin-2 (PC2), which form an ion channel complex that may mediate ciliary sensory processes and regulate endoplasmic reticulum (ER) Ca2+ release. Loss of PC1 expression profoundly alters cellular energy metabolism. The mechanisms that control the trafficking of PC1 and PC2, as well as their broader physiological roles, are poorly understood. We found that O2 levels regulate the subcellular localization and channel activity of the polycystin complex through its interaction with the O2-sensing prolyl hydroxylase domain containing protein EGLN3 (or PHD3), which hydroxylates PC1. Moreover, cells lacking PC1 expression use less O2 and show less mitochondrial Ca2+ uptake in response to bradykinin-induced ER Ca2+ release, indicating that PC1 can modulate mitochondrial function. These data suggest a novel role for the Polycystins in sensing and responding to cellular O2 levels.
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cyst growth Polycystins and primary cilia in autosomal dominant polycystic kidney disease
Kidney research and clinical practice, 2014Co-Authors: Seung Hun Lee, Stefan SomloAbstract:The primary cilium of renal epithelia acts as a transducer of extracellular stimuli. Polycystin (PC)1 is the protein encoded by the PKD1 gene that is responsible for the most common and severe form of autosomal dominant polycystic kidney disease (ADPKD). PC1 forms a complex with PC2 via their respective carboxy-terminal tails. Both proteins are expressed in the primary cilia. Mutations in either gene affect the normal architecture of renal tubules, giving rise to ADPKD. PC1 has been proposed as a receptor that modulates calcium signals via the PC2 channel protein. The effect of PC1 dosage has been described as the rate-limiting modulator of cystic disease. Reduced levels of PC1 or disruption of the balance in PC1/PC2 level can lead to the clinical features of ADPKD, without complete inactivation. Recent data show that ADPKD resulting from inactivation of Polycystins can be markedly slowed if structurally intact cilia are also disrupted at the same time. Despite the fact that no single model or mechanism from these has been able to describe exclusively the pathogenesis of cystic kidney disease, these findings suggest the existence of a novel cilia-dependent, cyst-promoting pathway that is normally repressed by polycystin function. The results enable us to rethink our current understanding of genetics and cilia signaling pathways of ADPKD.
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Polycystin-2 Regulates Proliferation and Branching Morphogenesis in Kidney Epithelial Cells
The Journal of biological chemistry, 2005Co-Authors: David H. Grimm, Stefan Somlo, Yiqiang Cai, Anil Karihaloo, Lloyd G. Cantley, Michael J. CaplanAbstract:Abstract Autosomal dominant polycystic kidney disease (ADPKD) is characterized by the formation of multiple fluid-filled cysts that expand over time and destroy the renal architecture. Loss or mutation of polycystin-1 or polycystin-2, the respective proteins encoded by the ADPKD genes PKD1 and PKD2, is associated with most cases of ADPKD. Thus, the polycystin proteins likely play a role in cell proliferation and morphogenesis. Recent studies indicate that polycystin-1 is involved in these processes, but little is known about the role played by polycystin-2. To address this question, we created a number of related cell lines variable in their expression of polycystin-2. We show that the basal and epidermal growth factor-stimulated rate of cell proliferation is higher in cells that do not express polycystin-2 versus those that do, indicating that polycystin-2 acts as a negative regulator of cell growth. In addition, cells not expressing polycystin-2 exhibit significantly more branching morphogenesis and multicellular tubule formation under basal and hepatocyte growth factor-stimulated conditions than their polycystin-2-expressing counterparts, suggesting that polycystin-2 may also play an important role in the regulation of tubulogenesis. Cells expressing a channel mutant of polycystin-2 proliferated faster than those expressing the wild-type protein, but exhibited blunted tubule formation. Thus, the channel activity of polycystin-2 may be an important component of its regulatory machinery. Finally, we show that polycystin-2 regulation of cell proliferation appears to be dependent on its ability to prevent phosphorylated extracellular-related kinase from entering the nucleus. Our results indicate that polycystin-2 is necessary for the proper growth and differentiation of kidney epithelial cells and suggest a possible mechanism for the cyst formation seen in ADPKD2.
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The N-terminal Extracellular Domain Is Required for Polycystin-1-dependent Channel Activity
The Journal of biological chemistry, 2004Co-Authors: Victor Babich, Stefan Somlo, Yiqiang Cai, Oxana Ibraghimov-beskrovnaya, Wei Zhong Zeng, Byung Il Yeh, Chou Long HuangAbstract:Abstract Autosomal dominant polycystic kidney disease (PKD) is caused by mutation of polycystin-1 or polycystin-2. Polycystin-2 is a Ca2+-permeable cation channel. Polycystin-1 is an integral membrane protein of less defined function. The N-terminal extracellular region of polycystin-1 contains potential motifs for protein and carbohydrate interaction. We now report that expression of polycystin-1 alone in Chinese hamster ovary (CHO) cells and in PKD2-null cells can confer Ca2+-permeable non-selective cation currents. Co-expression of a loss-of-function mutant of polycystin-2 in CHO cells does not reduce polycystin-1-dependent channel activity. A polycystin-1 mutant lacking ∼2900 amino acids of the extracellular region is targeted to the cell surface but does not produce current. Extracellular application of antibodies against the immunoglobulin-like PKD domains reduces polycystin-1-dependent current. These results support the hypothesis that polycystin-1 is a surface membrane receptor that transduces the signal via changes in ionic currents.
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Analysis of the Polycystins in Aortic Vascular Smooth Muscle Cells
Journal of the American Society of Nephrology : JASN, 2003Co-Authors: Qi Qian, Stefan Somlo, Peter C Harris, Yiqiang Cai, Christopher J. Ward, Vicente E. TorresAbstract:ABSTRACT. The leading cause of death in autosomal dominant polycystic kidney disease (ADPKD) is cardiovascular. However, little is known about the pathogenesis of these manifestations. The present study was undertaken to characterize the ADPKD proteins, the Polycystins, in vascular smooth muscle cells. It was demonstrated that the expression of polycystin-1 is developmentally regulated, whereas polycystin-2 has a more constant level of expression. A polycystin-1 subpopulation was immunoprecipitated by polycystin-2, indicating an in vivo interaction of these two proteins. Analysis with glycosidase and cell surface biotinylation indicates that some polycystin-1 products, but not polycystin-2, are located on the plasma membrane. Immunofluorescence showed that most of the polycystin-1 and polycystin-2 was cytoplasmic but that persistent polycystin-1 staining was located in proximity to the cell surface after a Triton-X extraction, whereas no clear surface localization of polycystin-2 was detected. Immuno-gold electron microscopy revealed that polycystin-1 was localized at the plasma membrane and sarcoplasmic reticulum, whereas polycystin-2 was mainly located in the sarcoplasmic reticulum. Both Polycystins were found to be associated with dense plaques. These observations are consistent with an important role of the Polycystins in the development, maintenance, and function of the myoelastic arterial organization and with the vascular phenotype associated with ADPKD. E-mail: Qian.qi@mayo.edu
Yiqiang Cai - One of the best experts on this subject based on the ideXlab platform.
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altered trafficking and stability of Polycystins underlie polycystic kidney disease
Journal of Clinical Investigation, 2014Co-Authors: Yiqiang Cai, Sorin V Fedeles, Ke Dong, Georgia I Anyatonwu, Tamehito Onoe, Michihiro Mitobe, Jiandong Gao, Dayne Okuhara, Xin Tian, Annarachel GallagherAbstract:The most severe form of autosomal dominant polycystic kidney disease occurs in patients with mutations in the gene (PKD1) encoding polycystin-1 (PC1). PC1 is a complex polytopic membrane protein expressed in cilia that undergoes autoproteolytic cleavage at a G protein–coupled receptor proteolytic site (GPS). A quarter of PKD1 mutations are missense variants, though it is not clear how these mutations promote disease. Here, we established a cell-based system to evaluate these mutations and determined that GPS cleavage is required for PC1 trafficking to cilia. A common feature among a subset of pathogenic missense mutations is a resulting failure of PC1 to traffic to cilia regardless of GPS cleavage. The application of our system also identified a missense mutation in the gene encoding polycystin-2 (PC2) that prevented this protein from properly trafficking to cilia. Using a Pkd1-BAC recombineering approach, we developed murine models to study the effects of these mutations and confirmed that only the cleaved form of PC1 exits the ER and can rescue the embryonically lethal Pkd1-null mutation. Additionally, steady-state expression levels of the intramembranous COOH-terminal fragment of cleaved PC1 required an intact interaction with PC2. The results of this study demonstrate that PC1 trafficking and expression require GPS cleavage and PC2 interaction, respectively, and provide a framework for functional assays to categorize the effects of missense mutations in Polycystins.
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Polycystin-2 Regulates Proliferation and Branching Morphogenesis in Kidney Epithelial Cells
The Journal of biological chemistry, 2005Co-Authors: David H. Grimm, Stefan Somlo, Yiqiang Cai, Anil Karihaloo, Lloyd G. Cantley, Michael J. CaplanAbstract:Abstract Autosomal dominant polycystic kidney disease (ADPKD) is characterized by the formation of multiple fluid-filled cysts that expand over time and destroy the renal architecture. Loss or mutation of polycystin-1 or polycystin-2, the respective proteins encoded by the ADPKD genes PKD1 and PKD2, is associated with most cases of ADPKD. Thus, the polycystin proteins likely play a role in cell proliferation and morphogenesis. Recent studies indicate that polycystin-1 is involved in these processes, but little is known about the role played by polycystin-2. To address this question, we created a number of related cell lines variable in their expression of polycystin-2. We show that the basal and epidermal growth factor-stimulated rate of cell proliferation is higher in cells that do not express polycystin-2 versus those that do, indicating that polycystin-2 acts as a negative regulator of cell growth. In addition, cells not expressing polycystin-2 exhibit significantly more branching morphogenesis and multicellular tubule formation under basal and hepatocyte growth factor-stimulated conditions than their polycystin-2-expressing counterparts, suggesting that polycystin-2 may also play an important role in the regulation of tubulogenesis. Cells expressing a channel mutant of polycystin-2 proliferated faster than those expressing the wild-type protein, but exhibited blunted tubule formation. Thus, the channel activity of polycystin-2 may be an important component of its regulatory machinery. Finally, we show that polycystin-2 regulation of cell proliferation appears to be dependent on its ability to prevent phosphorylated extracellular-related kinase from entering the nucleus. Our results indicate that polycystin-2 is necessary for the proper growth and differentiation of kidney epithelial cells and suggest a possible mechanism for the cyst formation seen in ADPKD2.
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The N-terminal Extracellular Domain Is Required for Polycystin-1-dependent Channel Activity
The Journal of biological chemistry, 2004Co-Authors: Victor Babich, Stefan Somlo, Yiqiang Cai, Oxana Ibraghimov-beskrovnaya, Wei Zhong Zeng, Byung Il Yeh, Chou Long HuangAbstract:Abstract Autosomal dominant polycystic kidney disease (PKD) is caused by mutation of polycystin-1 or polycystin-2. Polycystin-2 is a Ca2+-permeable cation channel. Polycystin-1 is an integral membrane protein of less defined function. The N-terminal extracellular region of polycystin-1 contains potential motifs for protein and carbohydrate interaction. We now report that expression of polycystin-1 alone in Chinese hamster ovary (CHO) cells and in PKD2-null cells can confer Ca2+-permeable non-selective cation currents. Co-expression of a loss-of-function mutant of polycystin-2 in CHO cells does not reduce polycystin-1-dependent channel activity. A polycystin-1 mutant lacking ∼2900 amino acids of the extracellular region is targeted to the cell surface but does not produce current. Extracellular application of antibodies against the immunoglobulin-like PKD domains reduces polycystin-1-dependent current. These results support the hypothesis that polycystin-1 is a surface membrane receptor that transduces the signal via changes in ionic currents.
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Analysis of the Polycystins in Aortic Vascular Smooth Muscle Cells
Journal of the American Society of Nephrology : JASN, 2003Co-Authors: Qi Qian, Stefan Somlo, Peter C Harris, Yiqiang Cai, Christopher J. Ward, Vicente E. TorresAbstract:ABSTRACT. The leading cause of death in autosomal dominant polycystic kidney disease (ADPKD) is cardiovascular. However, little is known about the pathogenesis of these manifestations. The present study was undertaken to characterize the ADPKD proteins, the Polycystins, in vascular smooth muscle cells. It was demonstrated that the expression of polycystin-1 is developmentally regulated, whereas polycystin-2 has a more constant level of expression. A polycystin-1 subpopulation was immunoprecipitated by polycystin-2, indicating an in vivo interaction of these two proteins. Analysis with glycosidase and cell surface biotinylation indicates that some polycystin-1 products, but not polycystin-2, are located on the plasma membrane. Immunofluorescence showed that most of the polycystin-1 and polycystin-2 was cytoplasmic but that persistent polycystin-1 staining was located in proximity to the cell surface after a Triton-X extraction, whereas no clear surface localization of polycystin-2 was detected. Immuno-gold electron microscopy revealed that polycystin-1 was localized at the plasma membrane and sarcoplasmic reticulum, whereas polycystin-2 was mainly located in the sarcoplasmic reticulum. Both Polycystins were found to be associated with dense plaques. These observations are consistent with an important role of the Polycystins in the development, maintenance, and function of the myoelastic arterial organization and with the vascular phenotype associated with ADPKD. E-mail: Qian.qi@mayo.edu
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Analysis of the Polycystins in Aortic Vascular Smooth Muscle Cells
Journal of the American Society of Nephrology : JASN, 2003Co-Authors: Qi Qian, Stefan Somlo, Peter C Harris, Yiqiang Cai, Christopher J. Ward, Vicente E. TorresAbstract:The leading cause of death in autosomal dominant polycystic kidney disease (ADPKD) is cardiovascular. However, little is known about the pathogenesis of these manifestations. The present study was undertaken to characterize the ADPKD proteins, the Polycystins, in vascular smooth muscle cells. It was demonstrated that the expression of polycystin-1 is developmentally regulated, whereas polycystin-2 has a more constant level of expression. A polycystin-1 subpopulation was immunoprecipitated by polycystin-2, indicating an in vivo interaction of these two proteins. Analysis with glycosidase and cell surface biotinylation indicates that some polycystin-1 products, but not polycystin-2, are located on the plasma membrane. Immunofluorescence showed that most of the polycystin-1 and polycystin-2 was cytoplasmic but that persistent polycystin-1 staining was located in proximity to the cell surface after a Triton-X extraction, whereas no clear surface localization of polycystin-2 was detected. Immuno-gold electron microscopy revealed that polycystin-1 was localized at the plasma membrane and sarcoplasmic reticulum, whereas polycystin-2 was mainly located in the sarcoplasmic reticulum. Both Polycystins were found to be associated with dense plaques. These observations are consistent with an important role of the Polycystins in the development, maintenance, and function of the myoelastic arterial organization and with the vascular phenotype associated with ADPKD.
Vicente E. Torres - One of the best experts on this subject based on the ideXlab platform.
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Analysis of the Polycystins in Aortic Vascular Smooth Muscle Cells
Journal of the American Society of Nephrology : JASN, 2003Co-Authors: Qi Qian, Stefan Somlo, Peter C Harris, Yiqiang Cai, Christopher J. Ward, Vicente E. TorresAbstract:ABSTRACT. The leading cause of death in autosomal dominant polycystic kidney disease (ADPKD) is cardiovascular. However, little is known about the pathogenesis of these manifestations. The present study was undertaken to characterize the ADPKD proteins, the Polycystins, in vascular smooth muscle cells. It was demonstrated that the expression of polycystin-1 is developmentally regulated, whereas polycystin-2 has a more constant level of expression. A polycystin-1 subpopulation was immunoprecipitated by polycystin-2, indicating an in vivo interaction of these two proteins. Analysis with glycosidase and cell surface biotinylation indicates that some polycystin-1 products, but not polycystin-2, are located on the plasma membrane. Immunofluorescence showed that most of the polycystin-1 and polycystin-2 was cytoplasmic but that persistent polycystin-1 staining was located in proximity to the cell surface after a Triton-X extraction, whereas no clear surface localization of polycystin-2 was detected. Immuno-gold electron microscopy revealed that polycystin-1 was localized at the plasma membrane and sarcoplasmic reticulum, whereas polycystin-2 was mainly located in the sarcoplasmic reticulum. Both Polycystins were found to be associated with dense plaques. These observations are consistent with an important role of the Polycystins in the development, maintenance, and function of the myoelastic arterial organization and with the vascular phenotype associated with ADPKD. E-mail: Qian.qi@mayo.edu
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Analysis of the Polycystins in Aortic Vascular Smooth Muscle Cells
Journal of the American Society of Nephrology : JASN, 2003Co-Authors: Qi Qian, Stefan Somlo, Peter C Harris, Yiqiang Cai, Christopher J. Ward, Vicente E. TorresAbstract:The leading cause of death in autosomal dominant polycystic kidney disease (ADPKD) is cardiovascular. However, little is known about the pathogenesis of these manifestations. The present study was undertaken to characterize the ADPKD proteins, the Polycystins, in vascular smooth muscle cells. It was demonstrated that the expression of polycystin-1 is developmentally regulated, whereas polycystin-2 has a more constant level of expression. A polycystin-1 subpopulation was immunoprecipitated by polycystin-2, indicating an in vivo interaction of these two proteins. Analysis with glycosidase and cell surface biotinylation indicates that some polycystin-1 products, but not polycystin-2, are located on the plasma membrane. Immunofluorescence showed that most of the polycystin-1 and polycystin-2 was cytoplasmic but that persistent polycystin-1 staining was located in proximity to the cell surface after a Triton-X extraction, whereas no clear surface localization of polycystin-2 was detected. Immuno-gold electron microscopy revealed that polycystin-1 was localized at the plasma membrane and sarcoplasmic reticulum, whereas polycystin-2 was mainly located in the sarcoplasmic reticulum. Both Polycystins were found to be associated with dense plaques. These observations are consistent with an important role of the Polycystins in the development, maintenance, and function of the myoelastic arterial organization and with the vascular phenotype associated with ADPKD.