The Experts below are selected from a list of 900 Experts worldwide ranked by ideXlab platform

Andrey S Shaw - One of the best experts on this subject based on the ideXlab platform.

  • effects of CD2 Associated Protein deficiency on amyloid β in neuroblastoma cells and in an app transgenic mouse model
    Molecular Neurodegeneration, 2015
    Co-Authors: Fan Liao, Andrey S Shaw, Hong Jiang, Subhashini Srivatsan, Qingli Xiao, Katheryn B Lefton, Kaoru Yamada, Thomas E Mahan, David M Holtzman
    Abstract:

    Background CD2-Associated Protein (CD2AP) is an SH3-containing scaffold adaptor Protein which regulates the actin cytoskeleton. Recently, CD2AP was identified as a genetic risk factor for Alzheimer’s disease (AD) by several genome-wide association studies. One of the hallmarks of AD is the accumulation of aggregated forms of Amyloid-β (Aβ) in the brain. In humans, CD2AP AD susceptibility locus (rs9349407) is Associated with an increased plaque burden. Aβ production is highly regulated by endocytosis and is influenced by lysosomal function. Lysosomal trafficking is influenced by CD2AP. In this study, we decreased CD2AP levels in N2a neuroblastoma cultures and PS1APP mice and analyzed Aβ levels and plaque burden.

  • CD2 Associated Protein regulates plasmacytoid dendritic cell migration but is dispensable for their development and cytokine production
    Journal of Immunology, 2013
    Co-Authors: Subhashini Srivatsan, Melissa Swiecki, Karel Otero, Marina Cella, Andrey S Shaw
    Abstract:

    Plasmacytoid dendritic cells (pDCs) are a dendritic cell subset that secrete type I IFNs in response to microbial stimuli. The scaffold Protein, CD2-Associated Protein (CD2AP), is a marker of human pDCs as it is highly expressed in this cell type. Recently, in human pDCs, decreased CD2AP expression appeared to enhance the production of type I IFNs via an inhibitory receptor-induced signaling cascade. In this study, we sought to determine the role of CD2AP in murine pDCs using CD2AP knockout (KO) mice. CD2AP was dispensable for the development of pDCs and for the upregulation of activation markers following stimulation. Loss of CD2AP expression did not affect the production of type I IFNs stimulated by TLR ligation, and only slightly impaired type I IFN production when inhibitory pathways were engaged in vitro. This was also confirmed by showing that CD2AP deficiency did not influence type I IFN production by pDCs in vivo. Because CD2AP plays a role in regulating actin dynamics, we examined the actin cytoskeleton in pDCs and found that activated CD2AP KO pDCs had significantly higher levels of actin polymerization than wild-type pDCs. Using two different inflammation models, we found that CD2AP KO pDCs have a defect in lymph node migration, correlating with the defects in actin dynamics. Our work excludes a role for CD2AP in the regulation of type I IFNs in pDCs, and suggests that the major function of CD2AP is on the actin cytoskeleton, affecting migration to local lymph nodes under conditions of inflammation.

  • septin 7 forms a complex with CD2ap and nephrin and regulates glucose transporter trafficking
    Molecular Biology of the Cell, 2012
    Co-Authors: Anita A Wasik, Andrey S Shaw, Zydrune Polianskyteprause, Mengqiu Dong, John R Yates, Marilyn G Farquhar, Sanna Lehtonen
    Abstract:

    Podocytes are insulin-sensitive and take up glucose in response to insulin. This requires nephrin, which interacts with vesicle-Associated membrane Protein 2 (VAMP2) on GLUT4 storage vesicles (GSVs) and facilitates their fusion with the plasma membrane. In this paper, we show that the filament-forming GTPase septin 7 is expressed in podocytes and associates with CD2-Associated Protein (CD2AP) and nephrin, both essential for glomerular ultrafiltration. In addition, septin 7 coimmunoprecipitates with VAMP2. Subcellular fractionation of cultured podocytes revealed that septin 7 is found in both cytoplasmic and membrane fractions, and immunofluorescence microscopy showed that septin 7 is expressed in a filamentous pattern and is also found on vesicles and the plasma membrane. The filamentous localization of septin 7 depends on CD2AP and intact actin organization. A 2-deoxy-d-glucose uptake assay indicates that depletion of septin 7 by small interfering RNA or alteration of septin assembly by forchlorfenuron facilitates glucose uptake into cells and further, knockdown of septin 7 increased the interaction of VAMP2 with nephrin and syntaxin 4. The data indicate that septin 7 hinders GSV trafficking and further, the interaction of septin 7 with nephrin in glomeruli suggests that septin 7 may participate in the regulation of glucose transport in podocytes.

  • focal segmental glomerulosclerosis in a patient homozygous for a CD2ap mutation commentary
    Kidney International, 2007
    Co-Authors: Shreeram Akilesh, Andrey S Shaw, A Koziell, M M Lowik, Patricia J T A Groenen, Inge Pronk, Marc R Lilien, Roel Goldschmeding, Henry B.p.m. Dijkman, Elena Levtchenko
    Abstract:

    Focal segmental glomerulosclerosis (FSGS) is a histologic diagnosis in several kidney diseases characterized by Proteinuria and a severe decrease in kidney function. Mutations in several genes were found in patients with primary FSGS, one of which is a CD2-Associated Protein CD2AP (originally referred to as CMS). This gene encodes an adaptor Protein that plays a role in endocytosis, cell motility, and cell survival. Mice deficient in CD2ap (the mouse homolog) die due to kidney failure, while heterozygous mice develop lesions similar to those of FSGS patients. In the kidney, CD2AP regulates the actin cytoskeleton. The only previously described patient with CD2AP mutation had a severely truncated Protein. In this study, we describe a patient with a novel mutation resulting in a premature stop codon yielding a Protein truncated by only 4%. This shortened CD2AP Protein displays a significantly decreased F-actin binding efficiency in vitro with no expression of the mutated allele in the patient's lymphocytes. Heterozygous expression of the CD2AP mutation in both parents did not lead to any kidney pathology, as both have normal glomerular filtration rates and no Proteinuria.

  • identification of a novel inhibitory actin capping Protein binding motif in CD2 Associated Protein
    Journal of Biological Chemistry, 2006
    Co-Authors: Serawit Bruck, Tobias B Huber, Robert J Ingham, Hanspeter Niederstrasser, Paul M Allen, Tony Pawson, John A Cooper, Andrey S Shaw
    Abstract:

    Abstract CD2-Associated Protein (CD2AP) is a scaffold molecule that plays a critical role in the maintenance of the kidney filtration barrier. Little, however, is understood about its mechanism of function. We used mass spectrometry to identify CD2AP-interacting Proteins. Many of the Proteins that we identified suggest a role for CD2AP in endocytosis and actin regulation. To address the role of CD2AP in regulation of the actin cytoskeleton, we focused on characterizing the interaction of CD2AP with actin-capping Protein CP. We identified a novel binding motif LXHXTXXRPK(X)6P present in CD2AP that is also found in its homolog Cin85 and other capping Protein-Associated Proteins such as CARMIL and CKIP-1. CD2AP inhibits the function of capping Protein in vitro. Therefore, our results support a role of CD2AP in the regulation of the actin cytoskeleton.

Sanna Lehtonen - One of the best experts on this subject based on the ideXlab platform.

  • inhibition of ship2 in CD2ap deficient podocytes ameliorates reactive oxygen species generation but aggravates apoptosis
    Scientific Reports, 2017
    Co-Authors: Pauliina Saurus, Tuomas Aleksi Tolvanen, Sonja Lindfors, Sara Kuusela, Harry Holthofer, Eero Lehtonen, Sanna Lehtonen
    Abstract:

    Lack of CD2-Associated Protein (CD2AP) in mice increases podocyte apoptosis and leads to glomerulosclerosis and renal failure. We showed previously that SHIP2, a negative regulator of the PI3K/AKT signalling pathway, interacts with CD2AP. Here, we found that the expression level and activity of SHIP2 and production of reactive oxygen species (ROS) are increased in cultured CD2AP knockout (CD2AP−/−) mouse podocytes. Oxidative stress was also increased in CD2AP−/− mouse glomeruli in vivo. We found that puromycin aminonucleoside (PA), known to increase ROS production and apoptosis, increases SHIP2 activity and reduces CD2AP expression in cultured human podocytes. PDK1 and CDK2, central regulators of AKT, were downregulated in CD2AP−/− or PA-treated podocytes. Downregulation of PDK1 and CDK2, ROS generation and apoptosis were prevented by CD2AP overexpression in both models. Notably, inhibition of SHIP2 activity with a small molecule inhibitor AS1949490 ameliorated ROS production in CD2AP−/− podocytes, but, surprisingly, further reduced PDK1 expression and aggravated apoptosis. AKT- and ERK-mediated signalling was diminished and remained reduced after AS1949490 treatment in the absence of CD2AP. The data suggest that inhibition of the catalytic activity of SHIP2 is beneficial in reducing oxidative stress, but leads to deleterious increase in apoptosis in podocytes with reduced expression of CD2AP.

  • lack of CD2ap disrupts glut4 trafficking and attenuates glucose uptake in podocytes
    Journal of Cell Science, 2015
    Co-Authors: Tuomas Aleksi Tolvanen, Surjya Narayan Dash, Zydrune Polianskyteprause, Vincent Dumont, Sanna Lehtonen
    Abstract:

    ABSTRACT The adapter Protein CD2-Associated Protein (CD2AP) functions in various signaling and vesicle trafficking pathways, including endosomal sorting and/or trafficking and degradation pathways. Here, we investigated the role of CD2AP in insulin-dependent glucose transporter 4 (Glut4, also known as SLC2A4) trafficking and glucose uptake. Glucose uptake was attenuated in CD2AP−/− podocytes compared with wild-type podocytes in the basal state, and CD2AP−/− podocytes failed to increase glucose uptake in response to insulin. Live-cell imaging revealed dynamic trafficking of HA–Glut4–GFP in wild-type podocytes, whereas in CD2AP−/− podocytes, HA–Glut4–GFP clustered perinuclearly. In subcellular membrane fractionations, CD2AP co-fractionated with Glut4, IRAP (also known as LNPEP) and sortilin, constituents of Glut4 storage vesicles (GSVs). We further found that CD2AP forms a complex with GGA2, a clathrin adaptor, which sorts Glut4 to GSVs, suggesting a role for CD2AP in this process. We also found that CD2AP forms a complex with clathrin and connects clathrin to actin in the perinuclear region. Furthermore, clathrin recycling back to trans-Golgi membranes from the vesicular fraction containing GSVs was defective in the absence of CD2AP. This leads to reduced insulin-stimulated trafficking of GSVs and attenuated glucose uptake into CD2AP−/− podocytes.

  • septin 7 forms a complex with CD2ap and nephrin and regulates glucose transporter trafficking
    Molecular Biology of the Cell, 2012
    Co-Authors: Anita A Wasik, Andrey S Shaw, Zydrune Polianskyteprause, Mengqiu Dong, John R Yates, Marilyn G Farquhar, Sanna Lehtonen
    Abstract:

    Podocytes are insulin-sensitive and take up glucose in response to insulin. This requires nephrin, which interacts with vesicle-Associated membrane Protein 2 (VAMP2) on GLUT4 storage vesicles (GSVs) and facilitates their fusion with the plasma membrane. In this paper, we show that the filament-forming GTPase septin 7 is expressed in podocytes and associates with CD2-Associated Protein (CD2AP) and nephrin, both essential for glomerular ultrafiltration. In addition, septin 7 coimmunoprecipitates with VAMP2. Subcellular fractionation of cultured podocytes revealed that septin 7 is found in both cytoplasmic and membrane fractions, and immunofluorescence microscopy showed that septin 7 is expressed in a filamentous pattern and is also found on vesicles and the plasma membrane. The filamentous localization of septin 7 depends on CD2AP and intact actin organization. A 2-deoxy-d-glucose uptake assay indicates that depletion of septin 7 by small interfering RNA or alteration of septin assembly by forchlorfenuron facilitates glucose uptake into cells and further, knockdown of septin 7 increased the interaction of VAMP2 with nephrin and syntaxin 4. The data indicate that septin 7 hinders GSV trafficking and further, the interaction of septin 7 with nephrin in glomeruli suggests that septin 7 may participate in the regulation of glucose transport in podocytes.

  • Lipid phosphatase SHIP2 downregulates insulin signalling in podocytes.
    Molecular and cellular endocrinology, 2010
    Co-Authors: Mervi E. Hyvönen, Pauliina Saurus, Harry Holthofer, Anita Wasik, Eija Heikkilä, Marika Havana, Ras Trokovic, Moin Saleem, Sanna Lehtonen
    Abstract:

    Podocyte injury plays an important role in the development of diabetic nephropathy. Podocytes are insulin-responsive and can develop insulin resistance, but the mechanisms are unknown. To study the role of CD2-Associated Protein (CD2AP) in podocyte injury, we performed a yeast two-hybrid screening on a glomerular library, and found that CD2AP bound to SH2-domain-containing inositol polyphosphate 5-phosphatase 2 (SHIP2), a negative regulator of insulin signalling. SHIP2 interacts with CD2AP in glomeruli and is expressed in podocytes, where it translocates to plasma membrane after insulin stimulation. Overexpression of SHIP2 in cultured podocytes reduces Akt activation in response to insulin, and promotes apoptosis. SHIP2 is upregulated in glomeruli of insulin resistant obese Zucker rats. These results indicate that SHIP2 downregulates insulin signalling in podocytes. The upregulation of SHIP2 in Zucker rat glomeruli prior to the age of onset of Proteinuria suggests a possible role for SHIP2 in the development of podocyte injury.

  • CD2 Associated Protein is widely expressed and differentially regulated during embryonic development
    Differentiation, 2008
    Co-Authors: Eero Lehtonen, Sanna Lehtonen, Jukka Tienari, Anou Londesborough, Ulla Pirvola, Ilkka Reima
    Abstract:

    CD2-Associated Protein (CD2AP) is an adapter Protein that is involved in various signaling and vesicular trafficking processes and also functions as a linker between plasma membrane Proteins and the actin cytoskeleton. The Protein is known to have important functions in T cells and glomerular podocytes, but it is also expressed by many other adult-type tissues and cells. Here we analyzed the expression of the Protein during early embryonic development and organogenesis of the mouse. The results showed differential tissue-specific regulation of CD2AP in developing and maturing organs. In oocytes and pre-implantation embryos, CD2AP was located diffusely in the cytoplasm, whereas in late blastocysts it was concentrated to the intercellular contacts. During organogenesis, CD2AP was distinctly upregulated upon, e.g., the pretubular aggregation of metanephric mesenchyme cells and the appearance of the osteoblastic rim around cartilages during endochondral ossification. High CD2AP expression was also observed during epithelial-like conversion of some highly specialized secretory cell types such as the odontoblasts, the cells of the choroid plexus and the decidualized cells of the endometrial stroma. In other instances, such as the development of the proximal tubuli of the kidney and the flat alveolar epithelium of the lung, the Protein was downregulated upon differentiation and maturation of the cells. Finally, certain cells, e.g., glomerular podocytes, those forming the collecting ducts of the kidney, and the urothelium of the kidney pelvis, expressed CD2AP throughout their differentiation and maturation. Multiple molecules and complex pathways regulate embryogenesis, and scaffolding Proteins apparently have pivotal roles in targeting and finetuning, e.g., growth factor- or hormone-induced processes. The cell-type specific spatio-temporal regulation of CD2AP during development suggests that this adapter Protein is a key regulatory partner in many signaling pathways and cellular processes governing differentiation and morphogenesis.

Erwin P. Bottinger - One of the best experts on this subject based on the ideXlab platform.

  • Glomerular expression of transforming growth factor–β (TGF-β) isoforms in mice lacking CD2-Associated Protein
    Pediatric Nephrology, 2006
    Co-Authors: Robert P. Woroniecki, Andrey S Shaw, Mario Schiffer, Frederick J. Kaskel, Erwin P. Bottinger
    Abstract:

    Mice lacking CD2-Associated Protein (CD2AP−/−) develop glomerular lesions resembling human focal segmental glomerulosclerosis (FSGS) between 3–4 weeks of age and die approximately 2 weeks later from massive Proteinuria and renal failure. The mechanisms involved in the glomerular injury in this model are unclear. In this study, we used laser capture microdissection (LCM) and real-time PCR, and examined expression of TGF-ß isoforms in CD2AP−/− mice at the level of isolated glomeruli. Total RNA yield from cryosections of 30 glomeruli was 10.71 ng (SD, 5.45) in CD2AP+/+ group ( n =7), and 4.20 ng (SD, 2.04) in CD2AP−/− group ( n =8), p =0.008. Expression of TGF-ß1 mRNA was increased 1.5-fold in the whole kidney ( p =0.030), and twofold in isolated CD2AP−/− glomeruli ( p =0.026). Whole kidney mRNA of TGF-ß receptor I (RI) and II (RII) was not different in CD2AP−/− and CD2AP+/+ animals, but it was increased in CD2AP−/− glomerular samples by 4.38-fold ( p =0.001) and 11.37-fold ( p =0.0163), respectively. By using LCM we confirmed increased glomerular expression levels of TGF-ß isoforms previously described by our group in glomeruli isolated by sieving in CD2AP KO mice and underscored the importance of local factors in the development of glomerulosclerosis.

  • glomerular expression of transforming growth factor β tgf β isoforms in mice lacking CD2 Associated Protein
    Pediatric Nephrology, 2006
    Co-Authors: Robert P. Woroniecki, Andrey S Shaw, Mario Schiffer, Frederick J. Kaskel, Erwin P. Bottinger
    Abstract:

    Mice lacking CD2-Associated Protein (CD2AP−/−) develop glomerular lesions resembling human focal segmental glomerulosclerosis (FSGS) between 3–4 weeks of age and die approximately 2 weeks later from massive Proteinuria and renal failure. The mechanisms involved in the glomerular injury in this model are unclear. In this study, we used laser capture microdissection (LCM) and real-time PCR, and examined expression of TGF-s isoforms in CD2AP−/− mice at the level of isolated glomeruli. Total RNA yield from cryosections of 30 glomeruli was 10.71 ng (SD, 5.45) in CD2AP+/+ group (n =7), and 4.20 ng (SD, 2.04) in CD2AP−/− group (n =8), p =0.008. Expression of TGF-s1 mRNA was increased 1.5-fold in the whole kidney (p =0.030), and twofold in isolated CD2AP−/− glomeruli (p =0.026). Whole kidney mRNA of TGF-s receptor I (RI) and II (RII) was not different in CD2AP−/− and CD2AP+/+ animals, but it was increased in CD2AP−/− glomerular samples by 4.38-fold (p =0.001) and 11.37-fold (p =0.0163), respectively. By using LCM we confirmed increased glomerular expression levels of TGF-s isoforms previously described by our group in glomeruli isolated by sieving in CD2AP KO mice and underscored the importance of local factors in the development of glomerulosclerosis.

  • a novel role for the adaptor molecule CD2 Associated Protein in transforming growth factor β induced apoptosis
    Journal of Biological Chemistry, 2004
    Co-Authors: Mario Schiffer, Andrey S Shaw, Peter Mundel, Erwin P. Bottinger
    Abstract:

    Abstract CD2-Associated Protein (CD2AP) is an adaptor molecule involved in T cell receptor signaling and podocyte homeostasis. CD2AP-deficient mice develop nephrotic syndrome and renal failure caused by glomerulosclerosis. Here we report that increased transforming growth factor-β1 (TGF-β1) expression and apoptosis were present in podocytes at the onset of albuminuria and were followed by depletion of podocytes Associated with progressive focal-segmental glomerulosclerosis in CD2AP-/- mice. Conditionally immortalized podocytes derived from CD2AP-/- mice were more susceptible to TGF-β-induced apoptosis compared with CD2AP+/+ podocytes. Reconstitution of CD2AP rescued CD2AP-/- podocytes from TGF-β-induced apoptosis. CD2AP was required for early activation of anti-apoptotic phosphatidylinositol 3-kinase (PI3K)/AKT and extracellular signal-regulated kinase 1/2 by TGF-β. In contrast, activation of pro-apoptotic p38 MAPK by TGF-β was accelerated and enhanced in the absence of CD2AP. CD2AP was not required for PI3K/AKT activation by insulin and epidermal growth factor, indicating that CD2AP is a selective mediator of anti-apoptotic TGF-β signaling. In summary, we identified CD2AP as a novel mediator for selective activation of survival pathways and repression of apoptosis signaling by TGF-β in podocytes. Together, our in vitro and in vivo findings suggest that TGF-β-induced podocyte apoptosis is an early pathomechanism in mice developing focal-segmental glomerulosclerosis Associated with functional impairment of CD2AP.

Eero Lehtonen - One of the best experts on this subject based on the ideXlab platform.

  • inhibition of ship2 in CD2ap deficient podocytes ameliorates reactive oxygen species generation but aggravates apoptosis
    Scientific Reports, 2017
    Co-Authors: Pauliina Saurus, Tuomas Aleksi Tolvanen, Sonja Lindfors, Sara Kuusela, Harry Holthofer, Eero Lehtonen, Sanna Lehtonen
    Abstract:

    Lack of CD2-Associated Protein (CD2AP) in mice increases podocyte apoptosis and leads to glomerulosclerosis and renal failure. We showed previously that SHIP2, a negative regulator of the PI3K/AKT signalling pathway, interacts with CD2AP. Here, we found that the expression level and activity of SHIP2 and production of reactive oxygen species (ROS) are increased in cultured CD2AP knockout (CD2AP−/−) mouse podocytes. Oxidative stress was also increased in CD2AP−/− mouse glomeruli in vivo. We found that puromycin aminonucleoside (PA), known to increase ROS production and apoptosis, increases SHIP2 activity and reduces CD2AP expression in cultured human podocytes. PDK1 and CDK2, central regulators of AKT, were downregulated in CD2AP−/− or PA-treated podocytes. Downregulation of PDK1 and CDK2, ROS generation and apoptosis were prevented by CD2AP overexpression in both models. Notably, inhibition of SHIP2 activity with a small molecule inhibitor AS1949490 ameliorated ROS production in CD2AP−/− podocytes, but, surprisingly, further reduced PDK1 expression and aggravated apoptosis. AKT- and ERK-mediated signalling was diminished and remained reduced after AS1949490 treatment in the absence of CD2AP. The data suggest that inhibition of the catalytic activity of SHIP2 is beneficial in reducing oxidative stress, but leads to deleterious increase in apoptosis in podocytes with reduced expression of CD2AP.

  • CD2 Associated Protein is widely expressed and differentially regulated during embryonic development
    Differentiation, 2008
    Co-Authors: Eero Lehtonen, Sanna Lehtonen, Jukka Tienari, Anou Londesborough, Ulla Pirvola, Ilkka Reima
    Abstract:

    CD2-Associated Protein (CD2AP) is an adapter Protein that is involved in various signaling and vesicular trafficking processes and also functions as a linker between plasma membrane Proteins and the actin cytoskeleton. The Protein is known to have important functions in T cells and glomerular podocytes, but it is also expressed by many other adult-type tissues and cells. Here we analyzed the expression of the Protein during early embryonic development and organogenesis of the mouse. The results showed differential tissue-specific regulation of CD2AP in developing and maturing organs. In oocytes and pre-implantation embryos, CD2AP was located diffusely in the cytoplasm, whereas in late blastocysts it was concentrated to the intercellular contacts. During organogenesis, CD2AP was distinctly upregulated upon, e.g., the pretubular aggregation of metanephric mesenchyme cells and the appearance of the osteoblastic rim around cartilages during endochondral ossification. High CD2AP expression was also observed during epithelial-like conversion of some highly specialized secretory cell types such as the odontoblasts, the cells of the choroid plexus and the decidualized cells of the endometrial stroma. In other instances, such as the development of the proximal tubuli of the kidney and the flat alveolar epithelium of the lung, the Protein was downregulated upon differentiation and maturation of the cells. Finally, certain cells, e.g., glomerular podocytes, those forming the collecting ducts of the kidney, and the urothelium of the kidney pelvis, expressed CD2AP throughout their differentiation and maturation. Multiple molecules and complex pathways regulate embryogenesis, and scaffolding Proteins apparently have pivotal roles in targeting and finetuning, e.g., growth factor- or hormone-induced processes. The cell-type specific spatio-temporal regulation of CD2AP during development suggests that this adapter Protein is a key regulatory partner in many signaling pathways and cellular processes governing differentiation and morphogenesis.

  • CD2-Associated Protein in human urogenital system and in adult kidney tumours
    Virchows Archiv, 2005
    Co-Authors: Jukka Tienari, Sanna Lehtonen, Eero Lehtonen
    Abstract:

    We studied expression of CD2-Associated Protein (CD2AP) in human urogenital system and in adult kidney tumours. In the cortex of normal kidney, CD2AP was expressed in all types of tubules and in the glomeruli. Labelling was more intense in cytokeratin 7- and in Tamm–Horsfall-positive tubules than in proximal tubules. In the medulla, expression was observed in the collecting ducts. Urothelium and the epithelium of prostatic acini, seminal vesicles, seminiferous tubules, epididymal ducts, Fallopian tube, endometrium and endocervix as well as granulosa cells showed moderate to strong CD2AP positivity. In syncytiotrophoblast, the expression was weaker than in cytotrophoblast. Endometrial stroma was negative, but decidualised stroma was weakly positive. Clear cell renal cell carcinoma (RCC) ( n =63) showed a weak expression. Type-I papillary RCCs ( n =4) and papillary adenomas ( n =3) were negative. The epithelium lining the cysts in multilocular cystic RCCs ( n =3) and in cystic nephroma ( n =1) was strongly positive. Chromophobe RCCs ( n =2), oncocytomas ( n =3) and urothelial carcinomas ( n =2) were moderately positive. The results show that CD2AP displays a specific expression pattern in human urogenital organs and that distinct expression is shown in several types of kidney tumours but not in type-I papillary RCCs or in papillary adenomas.

  • CD2 Associated Protein directly interacts with the actin cytoskeleton
    American Journal of Physiology-renal Physiology, 2002
    Co-Authors: Sanna Lehtonen, Fang Zhao, Eero Lehtonen
    Abstract:

    CD2-Associated Protein (CD2AP) is an adapter Protein associating with several membrane Proteins, including nephrin, mutated in congenital nephrotic syndrome of the Finnish type, and polycystin-2, mutated in type 2 autosomal dominant polycystic kidney disease. Both Proteins have critical roles in the maintenance of the integrity of the nephrons. Previous studies have suggested a role for CD2AP in the regulation of the organization of the actin cytoskeleton, but it has not been known whether the postulated association between CD2AP and actin is direct or mediated by other Proteins. In this study, we address this question by using various cellular and biochemical approaches. We show that CD2AP and F-actin partially colocalize in cultured cells and that disruption of the actin cytoskeleton results in disorganization of endogenous CD2AP. Using cytoskeletal fractionation by differential centrifugation, we demonstrate that a proportion of CD2AP associates with the actin cytoskeleton. Furthermore, using pure actin and purified CD2AP fusion Proteins in an F-actin coprecipitation assay, we show that CD2AP directly associates with filamentous actin and that this interaction is mediated by means of the COOH terminus of CD2AP. The present results suggest that CD2AP is involved in the regulation of the actin cytoskeleton and indicate that CD2AP may act as a direct adapter between the actin cytoskeleton and cell membrane Proteins, such as nephrin and polycystin-2. Alterations in these interactions could explain some of the pathophysiological changes in congenital nephrotic syndrome and polycystic kidney disease.

  • interaction of endogenous nephrin and CD2 Associated Protein in mouse epithelial m 1 cell line
    Journal of The American Society of Nephrology, 2002
    Co-Authors: Tuula Palmen, Eero Lehtonen, Sanna Lehtonen, Dontscho Kerjaschki, Corinne Antignac, Harry Holthofer
    Abstract:

    The interpodocyte slit diaphragm is an essential structure for maintaining the functional glomerular filtration barrier. The slit diaphragm is proposed to consist of an inter- acting meshwork of nephrin molecules. Earlier studies with tagged Proteins have suggested that the intracellular part of nephrin interacts with CD2-Associated Protein (CD2AP). This study was addressed to show by coimmunoprecipitation and pulldown assays an interaction of endogenously expressed nephrin and CD2AP in the kidney-derived mouse epithelial M-1 cell line, to provide evidence of the domain(s) of CD2AP involved in the interaction, and to show the localization of the respective Proteins by immunoelectron microscopy in kidney cortex. In addition, the localization of CD2AP, podocin, -ac- tinin 4, and nephrin was studied in human kidney glomeruli and in M-1 cells by immunofluorescence microscopy. The results indicate an endogenous interaction between nephrin and CD2AP in M-1 cells and suggest that this interaction is medi- ated by the third Src homology 3 (SH3) domain of CD2AP. We also show by immunoelectron microscopy that nephrin and CD2AP are detected at the slit diaphragm area, supporting their interaction in the glomeruli in vivo. In addition, nephrin was found to partially colocalize with CD2AP and podocin in double immunofluorescence microscopy, confirming the close proximity of these Proteins and proposing that these Proteins may belong to nephrin-Associated Protein complex in glomer- uli. The existence of nephrin, CD2AP, podocin, and -actinin 4 enables further characterization of their relationship in M-1 cells.

Guoping Zhou - One of the best experts on this subject based on the ideXlab platform.

  • mir 939 5p decreases the enrichment of rna polymerase ii in the promoter region of CD2ap involved in nephrotic syndrome
    Journal of Cellular Biochemistry, 2019
    Co-Authors: Jinya Wang, Daoqi Zhang, Xiaoqin Qiao, Guoping Zhou
    Abstract:

    : The expression changes of CD2-Associated Protein (CD2AP) can lead to kidney diseases with Proteinuria, including nephrotic syndrome (NS). A recent study reported that miRNAs may be important transcriptional regulators. In this study, we found increased expression of miR-939-5p and decreased expression of CD2AP in the peripheral blood of patients with NS. However, miR-939-5p did not show a regulatory effect on the 3'-untranslated region of CD2AP. The expression levels of specific Protein 1 and adenovirus E2 promoter-binding factor 1, important transcription regulators in the promoter region of CD2AP, were also not affected by microRNA (miR)-939-5p. We confirmed that miR-939-5p is in the nucleus by fluorescent in situ hybridization and cytoplasmic separation polymerase chain reaction. The promoter plasmid and miR-939-5p were cotransfected into HEK-293 cells, and the luciferase reporter gene assay was used to analyze the promoter activity. We found that miR-939-5p binds to a specific sequence in the CD2AP promoter. miR-939-5p was confirmed to reduce the recruitment of RNA polymerase II to the CD2AP promoter region by chromatin immunoprecipitation. These findings improve our understanding of the mechanism of miR-939-5p in NS and provide potential molecular therapeutic targets for NS.

  • microrna 939 down regulates CD2 Associated Protein by targeting promoter in hek 293t cells
    Renal Failure, 2016
    Co-Authors: Yuping Huang, Guoping Zhou
    Abstract:

    AbstractCD2-Associated Protein (CD2AP) serves as a slit diaphragm (SD) Protein and plays essential roles in maintaining podocyte integrity and reducing Proteinuria. MicroRNAs (miRNAs) are novel regulators of gene expression. Podocyte-specific loss of miRNAs would lead to significant Proteinuria. Here, we report new evidence in which miRNAs may function to suppress CD2AP expression through a transcriptional way. By scanning human CD2AP promoter in silico for sequences complementary to known miRNAs, we chose miR-939, miR-148b*, miR-191*, miR-638 as four candidates and transfected them into HEK-293T cells. Dual-luciferase reporter assay identified that only miR-939 significantly reduced the relative luciferase activity of CD2AP promoter region. Further analysis confirmed that the mRNA and Protein expressions of CD2AP were also down-regulated by miR-939. In conclusion, we have identified that miR-939 targets CD2AP promoter sequences and suppresses its gene expression. These findings suggest that miRNAs may me...

  • microrna 939 regulating CD2 Associated Protein expression by targeting promoter
    Chinese Journal of Applied Clinical Pediatrics, 2016
    Co-Authors: Yuping Huang, Jingwen Xu, Liping Zhao, Ling Li, Guoping Zhou
    Abstract:

    Objective To verify the targeting regulatory relationship between microRNA-939 (miR-939) and CD2-Associated Protein (CD2AP). Methods The online RegRNA software was used to predict the human CD2AP promoter for potential binding sites complementary to miR-939.HEK-293T cells were cotransfected with human CD2AP promoter plasmid pGL3-2K and microRNA negative control (miR-NC) or miR-939 mimics, and the relative luciferase activity(RLA) was detected at 24 h post-transfection.HEK-293T cells were transfected with miR-NC or miR-939 mimics for 48 h, and the CD2AP mRNA expression level was detected by adopting reverse transcript and real-time fluorescence quantification-PCR, while the CD2AP Protein expression level was detected by using Western blot. Results (1) There were 2 miR-939 binding sites at CD2AP promoter region, located at -468 to -491 and -654 to -677 upstream of initiation codon ATG (marked as + 1) relatively.(2)At 30 nmol/L, 50 nmol/L, the RLA in miR-NC group and miR-939 group were 6.81±0.88 vs 6.07±2.24, 5.88±1.44 vs 3.94±0.79 relatively, and there were no significant differences between the 2 groups (t=3.04, 2.06, all P>0.05), while the RLA between the 2 groups were 5.58±0.58 vs 3.29±0.64 at 100 nmol/L, and the difference was significant between the 2 groups(t=4.07, P<0.05). (3) At 30 nmol/L, 50 nmol/L and 100 nmol/L, the relative CD2AP mRNA expression in miR-NC group and miR-939 group were 1.00±0.01 vs 0.80±0.08, 1.00±0.00 vs 0.80±0.13 and 1.00±0.00 vs 0.72±0.07 relatively, while the CD2AP mRNA expression was decreased by 20%-30% at each concentration level, and there were significant differences between the 2 groups (t=4.44, 2.93, 6.84, all P<0.05). (4) At 50 nmol/L, the relative CD2AP Protein expression in miR-NC group and miR-939 group were 0.48±0.09 vs 0.19±0.12, and the CD2AP Protein expression was decreased, and the difference was significant (t=3.36, P<0.05). Conclusions CD2AP is the target gene of miR-939, and miR-939 can down-regulate the expression of CD2AP both in mRNA and Protein levels by targeting its promoter region, which indicates that miR-939 may mediate the podocyte injury. Key words: MicroRNA-939; CD2-Associated Protein; Podocyte; Promoter; Gene expression regulation

  • transcriptional activation of the human CD2ap promoter by e2f1
    PLOS ONE, 2012
    Co-Authors: Huaguo Xu, Yi Wang, Guoping Zhou
    Abstract:

    CD2-Associated Protein (CD2AP) is an adaptor molecule involved in T cell receptor signaling and podocyte homeostasis. CD2AP-deficient mice develop nephritic syndrome and renal failure caused by glomerulosclerosis. Transcription factor E2F1 is a key regulator of cell proliferation and apoptosis. Here we report that E2F1 up-regulates the human CD2AP promoter and further increases the mRNA and Protein levels of the human CD2AP in human embryonic kidney (HEK) 293 cells. By semi-quantitative RT-PCR and Western blot analysis we demonstrate that ectopic expression of E2F1 elevates the mRNA and Protein levels of CD2AP. Consistently, transient transfection assays prove that overexpression of E2F1 transactivates the CD2AP promoter while knocking-down of endogenous E2F1 by a shRNA strategy results in reduction of the CD2AP promoter activity. Toward understanding the underlying mechanism of this regulation, we performed chromatin immunoprecipitation and mutations of the putative Sp1 binding sites, demonstrating that E2F1 can bind to Sp1 binding site and overexpression of E2F1 is capable of increasing the binding of E2F1 and decreasing the binding of Sp1 to Sp1 binding sites.

  • transcriptional control of human CD2ap expression the role of sp1 and sp3
    Molecular Biology Reports, 2012
    Co-Authors: Huaguo Xu, Yi Wang, Guoping Zhou
    Abstract:

    The CD2 Associated Protein (CD2AP) is characterized as a T-lymphocyte CD2 adapter Protein and is found to be related to glomerulosclerosis, and CD2AP knockout mice develop a rapid onset nephrotic syndrome and die of renal failure. Here we report that the transcription factor Sp1 and Sp3 up-regulate the basal transcriptional activity of CD2AP and increase CD2AP expression at mRNA level. We show by Chromatin immunoprecipitation (ChIP) assay that Sp1 and Sp3 interact with the CD2AP promoter region in vivo. By transient transfection analysis we also demonstrate the mutations of Sp1/3 binding sites result in a profound reduction of CD2AP promoter activity. Overexpression of Sp1 and Sp3 transactivates the CD2AP promoter, whereas small interfering RNA-mediated (siRNA) blockage of Sp1 and Sp3 genes expressions inhibits markedly its activity. These results suggest that Sp1 and Sp3 play an important role in regulating CD2AP transcription through binding to the Sp1/3 binding sites.