The Experts below are selected from a list of 158697303 Experts worldwide ranked by ideXlab platform
Krzysztof Kobielak - One of the best experts on this subject based on the ideXlab platform.
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wnt7b is an important intrinsic regulator of hair follicle stem cell homeostasis and hair follicle cycling
Stem Cells, 2014Co-Authors: Eve Kandyba, Krzysztof KobielakAbstract:The hair follicle (HF) is an exceptional mini-organ to study the mechanisms which regulate HF morphogenesis, cycling, hair follicle stem cell (hfSCs) homeostasis, and progeny differentiation. During morphogenesis, Wnt signaling is well-characterized in the initiation of HF patterning but less is known about which particular Wnt ligands are required and whether individual Wnt ligands act in an indispensable or redundant manner during postnatal hfSCs anagen onset and HF cycle progression. Previously, we described the function of the bone morphogenetic protein (BMP) signaling target gene WNT7A in intrinsic regulation of hfSCs homeostasis in vivo. Here, we investigated the role of Wnt7b, which was also intrinsically upregulated in hfSCs during physiological and precocious anagen after BMP inhibition in vivo. We demonstrated Wnt7b to be a direct target of canonical BMP signaling in hfSCs and using Wnt7b conditional gene targeting during HF morphogenesis revealed disrupted HF cycling including a shorter anagen, premature catagen onset with overall shorter hair production, and diminished HF differentiation marker expression. Additionally, we observed that postnatal ablation of Wnt7b resulted in delayed HF activation, affecting both the hair germ and bulge hfSCs but still maintaining a two-step sequence of HF stimulation. Interestingly, Wnt7b cKO hfSCs participated in reformation of the new HF bulge, but with slower self-renewal. These findings demonstrate the importance of intrinsic Wnt7b expression in hfSCs regulation and normal HF cycling and surprisingly reveal a nonredundant role for Wnt7b in the control of HF anagen length and catagen entry which was not compensated by other Wnt ligands. Stem Cells 2014;32:886–901
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competitive balance of intrabulge bmp wnt signaling reveals a robust gene network ruling stem cell homeostasis and cyclic activation
Proceedings of the National Academy of Sciences of the United States of America, 2013Co-Authors: Eve Kandyba, Yi-bu Chen, Randall B. Widelitz, Yvonne Leung, Cheng-ming Chuong, Krzysztof KobielakAbstract:Hair follicles facilitate the study of stem cell behavior because stem cells in progressive activation stages, ordered within the follicle architecture, are capable of cyclic regeneration. To study the gene network governing the homeostasis of hair bulge stem cells, we developed a Keratin 15-driven genetic model to directly perturb molecular signaling in the stem cells. We visualize the behavior of these modified stem cells, evaluating their hair-regenerating ability and profile their molecular expression. Bone morphogenetic protein (BMP)-inactivated stem cells exhibit molecular profiles resembling those of hair germs, yet still possess multipotentiality in vivo. These cells also exhibit up-regulation of WNT7A, Wnt7b, and Wnt16 ligands and Frizzled (Fzd) 10 receptor. We demonstrate direct transcriptional modulation of the WNT7A promoter. These results highlight a previously unknown intra-stem cell antagonistic competition, between BMP and Wnt signaling, to balance stem cell activity. Reduced BMP signaling and increased Wnt signaling tilts each stem cell toward a hair germ fate and, vice versa, based on a continuous scale dependent on the ratio of BMP/Wnt activity. This work reveals one more hierarchical layer regulating stem cell homeostasis beneath the stem cell–dermal papilla-based epithelial–mesenchymal interaction layer and the hair follicle–intradermal adipocyte-based tissue interaction layer. Although hierarchical layers are all based on BMP/Wnt signaling, the multilayered control ensures that all information is taken into consideration and allows hair stem cells to sum up the total activators/inhibitors involved in making the decision of activation.
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Competitive balance of intrabulge BMP/Wnt signaling reveals a robust gene network ruling stem cell homeostasis and cyclic activation
Proceedings of the National Academy of Sciences of the United States of America, 2013Co-Authors: Eve Kandyba, Yi-bu Chen, Randall B. Widelitz, Yvonne Leung, Cheng-ming Chuong, Krzysztof KobielakAbstract:Hair follicles facilitate the study of stem cell behavior because stem cells in progressive activation stages, ordered within the follicle architecture, are capable of cyclic regeneration. To study the gene network governing the homeostasis of hair bulge stem cells, we developed a Keratin 15-driven genetic model to directly perturb molecular signaling in the stem cells. We visualize the behavior of these modified stem cells, evaluating their hair-regenerating ability and profile their molecular expression. Bone morphogenetic protein (BMP)-inactivated stem cells exhibit molecular profiles resembling those of hair germs, yet still possess multipotentiality in vivo. These cells also exhibit up-regulation of WNT7A, Wnt7b, and Wnt16 ligands and Frizzled (Fzd) 10 receptor. We demonstrate direct transcriptional modulation of the WNT7A promoter. These results highlight a previously unknown intra-stem cell antagonistic competition, between BMP and Wnt signaling, to balance stem cell activity. Reduced BMP signaling and increased Wnt signaling tilts each stem cell toward a hair germ fate and, vice versa, based on a continuous scale dependent on the ratio of BMP/Wnt activity. This work reveals one more hierarchical layer regulating stem cell homeostasis beneath the stem cell–dermal papilla-based epithelial–mesenchymal interaction layer and the hair follicle–intradermal adipocyte-based tissue interaction layer. Although hierarchical layers are all based on BMP/Wnt signaling, the multilayered control ensures that all information is taken into consideration and allows hair stem cells to sum up the total activators/inhibitors involved in making the decision of activation.
Jeremy Nathans - One of the best experts on this subject based on the ideXlab platform.
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Molecular determinants in Frizzled, Reck, and WNT7A for ligand-specific signaling in neurovascular development
eLife, 2019Co-Authors: Chris Cho, Yanshu Wang, John C. Williams, Philip M. Smallwood, Jeremy NathansAbstract:The molecular basis of Wnt-Frizzled specificity is a central question in developmental biology. Reck, a multi-domain and multi-functional glycosylphosphatidylinositol-anchored protein, specifically enhances beta-catenin signaling by WNT7A and Wnt7b in cooperation with the 7-transmembrane protein Gpr124. Among amino acids that distinguish WNT7A and Wnt7b from other Wnts, two clusters are essential for signaling in a Reck- and Gpr124-dependent manner. Both clusters are far from the site of Frizzled binding: one resides at the amino terminus and the second resides in a protruding loop. Within Reck, the fourth of five tandem repeats of an unusual domain with six-cysteines (the CC domain) is essential for WNT7A stimulation: substitutions P256A and W261A in CC4 eliminate this activity without changing protein abundance or surface localization. Mouse embryos carrying ReckP256A,W261A have severe defects in forebrain angiogenesis, providing the strongest evidence to date that Reck promotes CNS angiogenesis by specifically stimulating WNT7A and Wnt7b signaling.
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interplay of the norrin and WNT7A wnt7b signaling systems in blood brain barrier and blood retina barrier development and maintenance
Proceedings of the National Academy of Sciences of the United States of America, 2018Co-Authors: Yanshu Wang, Chris Cho, John C. Williams, Philip M. Smallwood, Chi Zhang, Harald J. Junge, Jeremy NathansAbstract:β-Catenin signaling controls the development and maintenance of the blood–brain barrier (BBB) and the blood–retina barrier (BRB), but the division of labor and degree of redundancy between the two principal ligand–receptor systems—the Norrin and WNT7A/Wnt7b systems—are incompletely defined. Here, we present a loss-of-function genetic analysis of postnatal BBB and BRB maintenance in mice that shows striking threshold and partial redundancy effects. In particular, the combined loss of WNT7A and Norrin or WNT7A and Frizzled4 (Fz4) leads to anatomically localized BBB defects that are far more severe than observed with loss of WNT7A, Norrin, or Fz4 alone. In the cerebellum, selective loss of WNT7A in glia combined with ubiquitous loss of Norrin recapitulates the phenotype observed with ubiquitous loss of both WNT7A and Norrin, implying that glia are the source of WNT7A in the cerebellum. Tspan12, a coactivator of Norrin signaling in the retina, is also active in BBB maintenance but is less potent than Norrin, consistent with a model in which Tspan12 enhances the amplitude of the Norrin signal in vascular endothelial cells. Finally, in the context of a partially impaired Norrin system, the retina reveals a small contribution to BRB development from the WNT7A/Wnt7b system. Taken together, these experiments define the extent of CNS region-specific cooperation for several components of the Norrin and WNT7A/Wnt7b systems, and they reveal substantial regional heterogeneity in the extent to which partially redundant ligands, receptors, and coactivators maintain the BBB and BRB.
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Interplay of the Norrin and WNT7A/Wnt7b Signaling Systems in Blood-Brain Barrier and Blood-Retina Barrier Development and Maintenance
Proceedings of the National Academy of Sciences of the United States of America, 2018Co-Authors: Yanshu Wang, Chris Cho, John C. Williams, Philip M. Smallwood, Chi Zhang, Harald J. Junge, Jeremy NathansAbstract:β-Catenin signaling controls the development and maintenance of the blood–brain barrier (BBB) and the blood–retina barrier (BRB), but the division of labor and degree of redundancy between the two principal ligand–receptor systems—the Norrin and WNT7A/Wnt7b systems—are incompletely defined. Here, we present a loss-of-function genetic analysis of postnatal BBB and BRB maintenance in mice that shows striking threshold and partial redundancy effects. In particular, the combined loss of WNT7A and Norrin or WNT7A and Frizzled4 (Fz4) leads to anatomically localized BBB defects that are far more severe than observed with loss of WNT7A, Norrin, or Fz4 alone. In the cerebellum, selective loss of WNT7A in glia combined with ubiquitous loss of Norrin recapitulates the phenotype observed with ubiquitous loss of both WNT7A and Norrin, implying that glia are the source of WNT7A in the cerebellum. Tspan12, a coactivator of Norrin signaling in the retina, is also active in BBB maintenance but is less potent than Norrin, consistent with a model in which Tspan12 enhances the amplitude of the Norrin signal in vascular endothelial cells. Finally, in the context of a partially impaired Norrin system, the retina reveals a small contribution to BRB development from the WNT7A/Wnt7b system. Taken together, these experiments define the extent of CNS region-specific cooperation for several components of the Norrin and WNT7A/Wnt7b systems, and they reveal substantial regional heterogeneity in the extent to which partially redundant ligands, receptors, and coactivators maintain the BBB and BRB.
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reck and gpr124 are essential receptor cofactors for WNT7A wnt7b specific signaling in mammalian cns angiogenesis and blood brain barrier regulation
Neuron, 2017Co-Authors: Philip M. Smallwood, Jeremy NathansAbstract:Summary Reck, a GPI-anchored membrane protein, and Gpr124, an orphan GPCR, have been implicated in WNT7A/Wnt7b signaling in the CNS vasculature. We show here that vascular endothelial cell (EC)-specific reduction in Reck impairs CNS angiogenesis and that EC-specific postnatal loss of Reck, combined with loss of Norrin, impairs blood-brain barrier (BBB) maintenance. The most N-terminal domain of Reck binds to the leucine-rich repeat (LRR) and immunoglobulin (Ig) domains of Gpr124, and weakening this interaction by targeted mutagenesis reduces Reck/Gpr124 stimulation of WNT7A signaling in cell culture and impairs CNS angiogenesis. Finally, a soluble Gpr124(LRR-Ig) probe binds to cells expressing Frizzled, WNT7A or Wnt7b, and Reck, and a soluble Reck(CC1-5) probe binds to cells expressing Frizzled, WNT7A or Wnt7b, and Gpr124. These experiments indicate that Reck and Gpr124 are part of the cell surface protein complex that transduces WNT7A- and Wnt7b-specific signals in mammalian CNS ECs to promote angiogenesis and regulate the BBB.
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Erratum: Reck and Gpr124 Are Essential Receptor Cofactors for WNT7A/Wnt7b-Specific Signaling in Mammalian CNS Angiogenesis and Blood-Brain Barrier Regulation (Neuron (2017) 95 (5)(1056–1073.e5) (S0896627317306797) (10.1016/j.neuron.2017.07.031))
Neuron, 2017Co-Authors: Chris Cho, Philip M. Smallwood, Jeremy NathansAbstract:Summary Reck, a GPI-anchored membrane protein, and Gpr124, an orphan GPCR, have been implicated in WNT7A/Wnt7b signaling in the CNS vasculature. We show here that vascular endothelial cell (EC)-specific reduction in Reck impairs CNS angiogenesis and that EC-specific postnatal loss of Reck, combined with loss of Norrin, impairs blood-brain barrier (BBB) maintenance. The most N-terminal domain of Reck binds to the leucine-rich repeat (LRR) and immunoglobulin (Ig) domains of Gpr124, and weakening this interaction by targeted mutagenesis reduces Reck/Gpr124 stimulation of WNT7A signaling in cell culture and impairs CNS angiogenesis. Finally, a soluble Gpr124(LRR-Ig) probe binds to cells expressing Frizzled, WNT7A or Wnt7b, and Reck, and a soluble Reck(CC1-5) probe binds to cells expressing Frizzled, WNT7A or Wnt7b, and Gpr124. These experiments indicate that Reck and Gpr124 are part of the cell surface protein complex that transduces WNT7A- and Wnt7b-specific signals in mammalian CNS ECs to promote angiogenesis and regulate the BBB.
Eve Kandyba - One of the best experts on this subject based on the ideXlab platform.
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wnt7b is an important intrinsic regulator of hair follicle stem cell homeostasis and hair follicle cycling
Stem Cells, 2014Co-Authors: Eve Kandyba, Krzysztof KobielakAbstract:The hair follicle (HF) is an exceptional mini-organ to study the mechanisms which regulate HF morphogenesis, cycling, hair follicle stem cell (hfSCs) homeostasis, and progeny differentiation. During morphogenesis, Wnt signaling is well-characterized in the initiation of HF patterning but less is known about which particular Wnt ligands are required and whether individual Wnt ligands act in an indispensable or redundant manner during postnatal hfSCs anagen onset and HF cycle progression. Previously, we described the function of the bone morphogenetic protein (BMP) signaling target gene WNT7A in intrinsic regulation of hfSCs homeostasis in vivo. Here, we investigated the role of Wnt7b, which was also intrinsically upregulated in hfSCs during physiological and precocious anagen after BMP inhibition in vivo. We demonstrated Wnt7b to be a direct target of canonical BMP signaling in hfSCs and using Wnt7b conditional gene targeting during HF morphogenesis revealed disrupted HF cycling including a shorter anagen, premature catagen onset with overall shorter hair production, and diminished HF differentiation marker expression. Additionally, we observed that postnatal ablation of Wnt7b resulted in delayed HF activation, affecting both the hair germ and bulge hfSCs but still maintaining a two-step sequence of HF stimulation. Interestingly, Wnt7b cKO hfSCs participated in reformation of the new HF bulge, but with slower self-renewal. These findings demonstrate the importance of intrinsic Wnt7b expression in hfSCs regulation and normal HF cycling and surprisingly reveal a nonredundant role for Wnt7b in the control of HF anagen length and catagen entry which was not compensated by other Wnt ligands. Stem Cells 2014;32:886–901
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competitive balance of intrabulge bmp wnt signaling reveals a robust gene network ruling stem cell homeostasis and cyclic activation
Proceedings of the National Academy of Sciences of the United States of America, 2013Co-Authors: Eve Kandyba, Yi-bu Chen, Randall B. Widelitz, Yvonne Leung, Cheng-ming Chuong, Krzysztof KobielakAbstract:Hair follicles facilitate the study of stem cell behavior because stem cells in progressive activation stages, ordered within the follicle architecture, are capable of cyclic regeneration. To study the gene network governing the homeostasis of hair bulge stem cells, we developed a Keratin 15-driven genetic model to directly perturb molecular signaling in the stem cells. We visualize the behavior of these modified stem cells, evaluating their hair-regenerating ability and profile their molecular expression. Bone morphogenetic protein (BMP)-inactivated stem cells exhibit molecular profiles resembling those of hair germs, yet still possess multipotentiality in vivo. These cells also exhibit up-regulation of WNT7A, Wnt7b, and Wnt16 ligands and Frizzled (Fzd) 10 receptor. We demonstrate direct transcriptional modulation of the WNT7A promoter. These results highlight a previously unknown intra-stem cell antagonistic competition, between BMP and Wnt signaling, to balance stem cell activity. Reduced BMP signaling and increased Wnt signaling tilts each stem cell toward a hair germ fate and, vice versa, based on a continuous scale dependent on the ratio of BMP/Wnt activity. This work reveals one more hierarchical layer regulating stem cell homeostasis beneath the stem cell–dermal papilla-based epithelial–mesenchymal interaction layer and the hair follicle–intradermal adipocyte-based tissue interaction layer. Although hierarchical layers are all based on BMP/Wnt signaling, the multilayered control ensures that all information is taken into consideration and allows hair stem cells to sum up the total activators/inhibitors involved in making the decision of activation.
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Competitive balance of intrabulge BMP/Wnt signaling reveals a robust gene network ruling stem cell homeostasis and cyclic activation
Proceedings of the National Academy of Sciences of the United States of America, 2013Co-Authors: Eve Kandyba, Yi-bu Chen, Randall B. Widelitz, Yvonne Leung, Cheng-ming Chuong, Krzysztof KobielakAbstract:Hair follicles facilitate the study of stem cell behavior because stem cells in progressive activation stages, ordered within the follicle architecture, are capable of cyclic regeneration. To study the gene network governing the homeostasis of hair bulge stem cells, we developed a Keratin 15-driven genetic model to directly perturb molecular signaling in the stem cells. We visualize the behavior of these modified stem cells, evaluating their hair-regenerating ability and profile their molecular expression. Bone morphogenetic protein (BMP)-inactivated stem cells exhibit molecular profiles resembling those of hair germs, yet still possess multipotentiality in vivo. These cells also exhibit up-regulation of WNT7A, Wnt7b, and Wnt16 ligands and Frizzled (Fzd) 10 receptor. We demonstrate direct transcriptional modulation of the WNT7A promoter. These results highlight a previously unknown intra-stem cell antagonistic competition, between BMP and Wnt signaling, to balance stem cell activity. Reduced BMP signaling and increased Wnt signaling tilts each stem cell toward a hair germ fate and, vice versa, based on a continuous scale dependent on the ratio of BMP/Wnt activity. This work reveals one more hierarchical layer regulating stem cell homeostasis beneath the stem cell–dermal papilla-based epithelial–mesenchymal interaction layer and the hair follicle–intradermal adipocyte-based tissue interaction layer. Although hierarchical layers are all based on BMP/Wnt signaling, the multilayered control ensures that all information is taken into consideration and allows hair stem cells to sum up the total activators/inhibitors involved in making the decision of activation.
Marilyn B. Renfree - One of the best experts on this subject based on the ideXlab platform.
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Hormone-responsive genes in the SHH and WNT/β-catenin signaling pathways influence urethral closure and phallus growth.
Biology of Reproduction, 2018Co-Authors: Yu Chen, Andrew J. Pask, Geoff Shaw, Asao Fujiyama, Yutaka Suzuki, Sumio Sugano, Marilyn B. RenfreeAbstract:Environmental endocrine disruptors (EEDs) that affect androgen or estrogen activity may disrupt gene regulation during phallus development to cause hypospadias or a masculinized clitoris. We treated developing male tammar wallabies with estrogen and females with androgen from day 20-40 postpartum (pp) during the androgen imprinting window of sensitivity. Estrogen inhibited phallus elongation but had no effect on urethral closure and did not significantly depress testicular androgen synthesis. Androgen treatment in females did not promote phallus elongation but initiated urethral closure. Phalluses were collected for transcriptome sequencing at day 50 pp when they first become sexually dimorphic to examine changes in two signaling pathways, sonic hedgehog (SHH) and wingless-type MMTV integration site family (WNT)/β-catenin. SHH mRNA and β-catenin were predominantly expressed in the urethral epithelium in the tammar phallus, as in eutherian mammals. Estrogen treatment and castration of males induced an upregulation of SHH, while androgen treatment downregulated SHH. These effects appear to be direct since we detected putative estrogen receptor α (ERα) and androgen receptor (AR) binding sites near SHH. WNT5A, like SHH, was downregulated by androgen, while WNT4 was upregulated in female phalluses after androgen treatment. After estrogen treatment, WIF1 and WNT7A were both downregulated in male phalluses. After castration, WNT9A was upregulated. These results suggest that SHH and WNT pathways are regulated by both estrogen and androgen to direct the proliferation and elongation of the phallus during differentiation. Their response to exogenous hormones makes these genes potential targets of EEDs in the etiology of abnormal phallus development including hypospadias.
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hormone responsive genes in the shh and wnt β catenin signaling pathways influence urethral closure and phallus growth
Biology of Reproduction, 2018Co-Authors: Yu Chen, Andrew J. Pask, Geoff Shaw, Asao Fujiyama, Yutaka Suzuki, Sumio Sugano, Marilyn B. RenfreeAbstract:Environmental endocrine disruptors (EEDs) that affect androgen or estrogen activity may disrupt gene regulation during phallus development to cause hypospadias or a masculinized clitoris. We treated developing male tammar wallabies with estrogen and females with androgen from day 20-40 postpartum (pp) during the androgen imprinting window of sensitivity. Estrogen inhibited phallus elongation but had no effect on urethral closure and did not significantly depress testicular androgen synthesis. Androgen treatment in females did not promote phallus elongation but initiated urethral closure. Phalluses were collected for transcriptome sequencing at day 50 pp when they first become sexually dimorphic to examine changes in two signaling pathways, sonic hedgehog (SHH) and wingless-type MMTV integration site family (WNT)/β-catenin. SHH mRNA and β-catenin were predominantly expressed in the urethral epithelium in the tammar phallus, as in eutherian mammals. Estrogen treatment and castration of males induced an upregulation of SHH, while androgen treatment downregulated SHH. These effects appear to be direct since we detected putative estrogen receptor α (ERα) and androgen receptor (AR) binding sites near SHH. WNT5A, like SHH, was downregulated by androgen, while WNT4 was upregulated in female phalluses after androgen treatment. After estrogen treatment, WIF1 and WNT7A were both downregulated in male phalluses. After castration, WNT9A was upregulated. These results suggest that SHH and WNT pathways are regulated by both estrogen and androgen to direct the proliferation and elongation of the phallus during differentiation. Their response to exogenous hormones makes these genes potential targets of EEDs in the etiology of abnormal phallus development including hypospadias.
Akira Kikuchi - One of the best experts on this subject based on the ideXlab platform.
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Apical secretion of Wnt1 in polarized epithelial cells is regulated by exocyst-mediated trafficking.
The Journal of Biochemistry, 2017Co-Authors: Hideki Yamamoto, Akira Sato, Akira KikuchiAbstract:Wnts are glycosylated proteins secreted from various cell types including mesenchymal, hematopoietic and epithelial cells. Directional secretion of Wnts in polarized epithelial cells is unique; Wnt11 is secreted apically, whereas Wnt5a and Wnt3a are secreted basolaterally. Here, we found that Wnt1 is equivalently secreted both apically and basolaterally in MDCK cells. Wnt1 was modified with a complex- or hybrid-type glycan at Asn29 and Asn359 and the high-mannose- or hybrid-type glycan at Asn316. Although glycosylation of Wnt11 at the N-terminal site was shown to be essential for its apical secretion, glycosylation of Asn29 of Wnt1 was not required. Instead, the apical secretion of Wnt1 was inhibited by knockdown of Sec6 and Sec8, suggesting that Wnt1 is secreted apically via exocyst-mediated transport. Basolateral secretion of Wnt1 was mediated by clathrin and AP-1, in mechanism similar to that used by Wnt5a and Wnt3a. Although Wingless was reported to be transcytosed to the basolateral region in the Drosophila wing disc, transcytosis was not involved in the basolateral secretion of Wnt1. Thus, the polarized secretion of Wnt1 is regulated by different mechanisms than other Wnts.
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Localization of glypican-4 in different membrane microdomains is involved in the regulation of Wnt signaling.
Journal of cell science, 2012Co-Authors: Hiroshi Sakane, Shinji Matsumoto, Akira Sato, Hideki Yamamoto, Akira KikuchiAbstract:Glypicans are members of the heparan sulfate proteoglycans (HSPGs) and are involved in various growth factor signaling mechanisms. Although HSPGs affect the β-catenin-dependent and -independent pathways of Wnt signaling, how they regulate distinct Wnt pathways is not clear. It has been suggested that the β-catenin-dependent pathway is initiated through receptor endocytosis in lipid raft microdomains and the independent pathway is activated through receptor endocytosis in non-lipid raft microdomains. Here, evidence is presented that glypican-4 (GPC4) is localized to both membrane microdomains and that the localization affects its ability to regulate distinct Wnt pathways. GPC4 bound to Wnt3a and Wnt5a, which activate the β-catenin-dependent and -independent pathways, respectively, and colocalized with Wnts on the cell surface. LRP6, one of Wnt3a coreceptors, was present in lipid raft microdomains, whereas Ror2, one of Wnt5a coreceptors, was localized to non-lipid raft microdomains. Expression of GPC4 enhanced the Wnt3a-dependent β-catenin pathway and the Wnt5a-dependent β-catenin-independent pathway, and knockdown of GPC4 suppressed both pathways. A GPC4 mutant that was localized to only non-lipid raft microdomains inhibited the β-catenin-dependent pathway but enhanced the β-catenin-independent pathway. These results suggest that GPC4 concentrates Wnt3a and Wnt5a to the vicinity of their specific receptors in different membrane microdomains, thereby regulating distinct Wnt signaling.
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wnt5a regulates distinct signalling pathways by binding to frizzled2
The EMBO Journal, 2010Co-Authors: Akira Sato, Hirofumi Koyama, Hiroshi Sakane, Akira Kikuchi, Hideki YamamotoAbstract:Wnt5a regulates multiple intracellular signalling cascades, but how Wnt5a determines the specificity of these pathways is not well understood. This study examined whether the internalization of Wnt receptors affects the ability of Wnt5a to regulate its signalling pathways. Wnt5a activated Rac in the β-catenin-independent pathway, and Frizzled2 (Fz2) and Ror1 or Ror2 were required for this action. Fz2 was internalized through a clathrin-mediated route in response to Wnt5a, and inhibition of clathrin-dependent internalization suppressed the ability of Wnt5a to activate Rac. As another action of Wnt5a, it inhibited Wnt3a-dependent lipoprotein receptor-related protein 6 (LRP6) phosphorylation and β-catenin accumulation. Wnt3a-dependent phosphorylation of LRP6 was enhanced in Wnt5a knockout embryonic fibroblasts. Fz2 was also required for the Wnt3a-dependent accumulation of β-catenin, and Wnt5a competed with Wnt3a for binding to Fz2 in vitro and in intact cells, thereby inhibiting the β-catenin pathway. This inhibitory action of Wnt5a was not affected by the impairment of clathrin-dependent internalization. These results suggest that Wnt5a regulates distinct pathways through receptor internalization-dependent and -independent mechanisms.
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Wnt5a modulates glycogen synthase kinase 3 to induce phosphorylation of receptor tyrosine kinase Ror2
Genes to cells : devoted to molecular & cellular mechanisms, 2007Co-Authors: Hiroyuki Yamamoto, Michiru Nishita, Akira Kikuchi, Kan Yoo, Yasuhiro MinamiAbstract:The receptor tyrosine kinase Ror2 plays important roles in mediating non-canonical Wnt5a signaling by activating the Wnt–JNK pathway and inhibiting the β-catenin–TCF pathway. It has been shown that Ror2 is phosphorylated and activated by casein kinase Iɛ when both molecules are over-expressed in cultured cells. However, it remains unknown whether or not Ror2 is phosphorylated upon Wnt5a stimulation. Here we show that Ror2 is phosphorylated on serine/threonine residues upon stimulation of cultured cells, expressing Ror2 endogenously, with Wnt5a, but not Wnt3a. It was found that treatment of cells with glycogen synthase kinase-3 (GSK-3) inhibitors (LiCl and SB216763) or small interfering RNAs (siRNAs) for GSK-3 (mainly GSK-3α) can inhibit Wnt5a-induced phosphorylation of Ror2. Immunoprecipitated Ror2 can also be phosphorylated by purified GSK-3α or GSK-3βin vitro, and ectopic co-expression of Ror2 and GSK-3 (mainly GSK-3α) in cultured cells results in Ror2 phosphorylation, irrespective of Wnt5a, that is sensitive to SB216763. These results indicate that GSK-3 is involved in Wnt5a-induced phosphorylation of Ror2. Moreover, it was found that Wnt5a-induced cell migration can be inhibited by SB216763 or by siRNA-mediated suppression of GSK-3α (and GSK-3β) expression, further emphasizing the role(s) of GSK-3 in Wnt5a-induced signaling.
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filopodia formation mediated by receptor tyrosine kinase ror2 is required for wnt5a induced cell migration
Journal of Cell Biology, 2006Co-Authors: Michiru Nishita, Akira Nomachi, Yasutaka Ohta, Shinji Takada, Nagako Sougawa, Akira Kikuchi, Shuichi Kani, Yasuhiro MinamiAbstract:The receptor tyrosine kinase Ror2 plays important roles in developmental morphogenesis. It has recently been shown that Ror2 mediates Wnt5a-induced noncanonical Wnt signaling by activating the Wnt–JNK pathway and inhibiting the β-catenin–TCF pathway. However, the function of Ror2 in noncanonical Wnt signaling leading to cell migration is largely unknown. We show, using genetically different or manipulated cultured cells, that Ror2 is critical for Wnt5a-induced, but not Wnt3a-induced, cell migration. Ror2-mediated cell migration requires the extracellular cysteine-rich domain (CRD), which is the binding site for Wnt5a, and the cytoplasmic proline-rich domain (PRD) of Ror2. Furthermore, Ror2 can mediate filopodia formation via actin reorganization, irrespective of Wnt5a, and this Ror2-mediated filopodia formation requires the actin-binding protein filamin A, which associates with the PRD of Ror2. Intriguingly, disruption of filopodia formation by suppressing the expression of either Ror2 or filamin A inhibits Wnt5a-induced cell migration, indicating that Ror2-mediated filopodia formation is essential for Wnt5a-induced cell migration.
