Calcium Dependence of Polycystin-2 Channel Activity Is Modulated by Phosphorylation at Ser812

Cai, Yiqiang; Anyatonwu, Georgia I.; Okuhara, Dayne; Lee, Kyu Beck; Yu, Zhiheng; Onoe, Tamehito; Mei, Chang Lin; Qian, Qi; Geng, Lin; Wiztgall, Ralph; Ehrlich, Barbara E.; Somlo, Stefan

doi:10.1074/jbc.m312031200

Cited by 140 publications

(200 citation statements)

References 35 publications

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“…A number of studies have suggested that disrupted Ca 2+ signaling as a consequence of PC2 mutations is associated with cyst development (24,27,30); however, a clear link between Ca 2+ channel disruption and cyst development has not been recapitulated in 3D tissues. The majority of PKD studies using 3D tissue culture have used MDCK cells as the cell line of choice (6)(7)(8)(9)(10)(11)(12), but PC2-dependent Ca 2+ signaling has not been studied extensively in this cell line.…”

Section: Discussionmentioning

confidence: 99%

“…Over 99% of PC2 resides on the endoplasmic reticulum (22), where it is known to act as a modulator of the InsP3R and the ryanodine receptor (RyR) (23), with the remainder on the primary cilia. PC2 itself can function as a Ca 2+ -activated Ca 2+ release channel (22,24).Although it was demonstrated in 3D cultures that the knockdown of PC1 leads to cyst development (25), the effect of knocking down PC2 or other Ca 2+ -signaling proteins has not been shown. It has been hypothesized that the disruption of PC2, or the proteins that it interacts with, will result in cyst growth, as Ca 2+ is a major signaling molecule (26,27).…”

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confidence: 99%

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Cyst formation following disruption of intracellular calcium signaling

Kuo¹,

DesRochers

Kimmerling

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Mutations in polycystin 1 and 2 (PC1 and PC2) cause the common genetic kidney disorder autosomal dominant polycystic kidney disease (ADPKD). It is unknown how these mutations result in renal cysts, but dysregulation of calcium (Ca 2+ ) signaling is a known consequence of PC2 mutations. PC2 functions as a Ca 2+ -activated Ca 2+ channel of the endoplasmic reticulum. We hypothesize that Ca 2+ signaling through PC2, or other intracellular Ca 2+ channels such as the inositol 1,4,5-trisphosphate receptor (InsP3R), is necessary to maintain renal epithelial cell function and that disruption of the Ca 2+ signaling leads to renal cyst development. The cell line LLC-PK1 has traditionally been used for studying PKD-causing mutations and Ca 2+ signaling in 2D culture systems. We demonstrate that this cell line can be used in long-term (8 wk) 3D tissue culture systems. In 2D systems, knockdown of InsP3R results in decreased Ca 2+ transient signals that are rescued by overexpression of PC2. In 3D systems, knockdown of either PC2 or InsP3R leads to cyst formation, but knockdown of InsP3R type 1 (InsP3R1) generated the largest cysts. InsP3R1 and InsP3R3 are differentially localized in both mouse and human kidney, suggesting that regional disruption of Ca 2+ signaling contributes to cystogenesis. All cysts had intact cilia 2 wk after starting 3D culture, but the cells with InsP3R1 knockdown lost cilia as the cysts grew. Studies combining 2D and 3D cell culture systems will assist in understanding how mutations in PC2 that confer altered Ca 2+ signaling lead to ADPKD cysts. primary cilia | polycysin 2 | calcium release T he commonly occurring genetic kidney disorder, autosomal dominant polycystic kidney disease (ADPKD), is the result of mutations in polycystin 1 or 2 (PC1 or PC2). The progressive cyst formation within all segments of the nephron that defines the disorder leads to renal failure requiring treatment by dialysis and/or organ transplantation (1-3). Altered Ca 2+ signaling is one of several pathways that have been implicated in the disease (4, 5). A major limitation toward elucidating the role of Ca 2+ signaling in cyst formation has been the lack of easily manipulated, physiologically relevant experimental methodologies.In the past, ADPKD research has relied largely upon data from mouse models and cells maintained in 2D cell culture. Mouse models have played a significant role in understanding the biology of cyst formation but are unable to fully recapitulate the physiology of disease progression in humans due to the inherent differences between the species including life span, genetics, and environment. Two-dimensional cell culture has the ability to provide information on signaling pathways and response to therapies in a fast, high-throughput manner, but is incapable of replicating the inherent 3D nature of cyst formation. Advances in 3D tissue culture over the past 2 decades have improved the ability to model cyst development in vitro. However, previously published 3D tissue models of ADPKD have relied upon shortter...

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

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Cyst formation following disruption of intracellular calcium signaling

Kuo¹,

DesRochers

Kimmerling

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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“…PC1 localization in plasma membrane was also detected in the kidneys of jck mice ( Figure 5W). We and others have previously shown that PC2 is localized to primary cilia in a variety of cultured cell lines [17][18][19][20][21] and in the kidney. 21 To our surprise, we did not detect PC2 in the cilia of tubular epithelia from wild-type mouse kidney, using either snap-frozen or perfusion-fixed kidney sections with nicely dilated lumens under our experimental conditions ( Figure 5, M to O), although Nek8 was easily detectable in cilia of kidney tubules ( Figure 5A).…”

Section: Increased Phosphorylation Of Pc2 In Jck Micementioning

confidence: 99%

“…PC2 has calcium channel functions in endoplasmic reticulum, plasma membrane, and cilia. [17][18][19][20][21] Proteins responsible for autosomal recessive PKD (ARPKD) are also found in cilia of renal epithelia, such as cystin for cpk (congenital polycystic kidney) mouse model, 22 polaris for orpk (Oak Ridge polycystic kidney) mouse model, 23,24 and fibrocystin in human ARPKD. 25,26 In this study, we report that the Nek8 protein is located in the proximal region of the primary cilia in mouse kidney tubules and in the same protein complex with PC2.…”

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Nek8 Regulates the Expression and Localization of Polycystin-1 and Polycystin-2

Sohara¹,

Luo²,

Zhang³

et al. 2008

Journal of the American Society of Nephrology

113

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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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Role of cilia in normal pancreas function and in diseased states

Diiorio

Rittenhouse

Bortell

et al. 2014

Birth Defects Research Pt C

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Primary cilia play an essential role in modulating signaling cascades that shape cellular responses to environmental cues to maintain proper tissue development. Mutations in primary cilium proteins have been linked to several rare developmental disorders, collectively known as ciliopathies. Together with other disorders associated with dysfunctional cilia/centrosomes, affected individuals have increased risk of developing metabolic syndrome, neurologic disorders, and diabetes. In pancreatic tissues, cilia are found exclusively in islet and ductal cells where they play an essential role in pancreatic tissue organization. Their absence or disorganization leads to pancreatic duct abnormalities, acinar cell loss, polarity defects, and dysregulated insulin secretion. Cilia in pancreatic tissues are hubs for cellular signaling. Many signaling components, such as Hh, Notch, and Wnt, localize to pancreatic primary cilia and are necessary for proper development of pancreatic epithelium and β-cell morphogenesis. Receptors for neuroendocrine hormones, such as Somatostatin Receptor 3, also localize to the cilium and may play a more direct role in controlling insulin secretion due to somatostatin's inhibitory function. Finally, unique calcium signaling, which is at the heart of β-cell function, also occurs in primary cilia. Whereas voltage-gated calcium channels trigger insulin secretion and serve a variety of homeostatic functions in β-cells, transient receptor potential channels regulate calcium levels within the cilium that may serve as a feedback mechanism, regulating insulin secretion. This review article summarizes our current understanding of the role of primary cilia in normal pancreas function and in the diseased state.

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Calcium Dependence of Polycystin-2 Channel Activity Is Modulated by Phosphorylation at Ser812

Cited by 140 publications

References 35 publications

Cyst formation following disruption of intracellular calcium signaling

Cyst formation following disruption of intracellular calcium signaling

Nek8 Regulates the Expression and Localization of Polycystin-1 and Polycystin-2

Role of cilia in normal pancreas function and in diseased states

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