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Bart O Williams - One of the best experts on this subject based on the ideXlab platform.
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Co-deletion of Lrp5 and LRP6 in the skeleton severely diminishes bone gain from sclerostin antibody administration.
Bone, 2020Co-Authors: Kyung-eun Lim, Bart O Williams, Whitney A. Bullock, Daniel J. Horan, Matthew L. Warman, Alexander G. RoblingAbstract:Abstract The cysteine knot protein sclerostin is an osteocyte-derived secreted inhibitor of the Wnt co-receptors LRP5 and LRP6. LRP5 plays a dominant role in bone homeostasis, but we previously reported that Sost/sclerostin suppression significantly increased osteogenesis regardless of Lrp5 presence or absence. Those observations suggested that the bone forming effects of sclerostin inhibition can occur through LRP6 (when Lrp5 is suppressed), or through other yet undiscovered mechanisms independent of Lrp5/6. To distinguish between these two possibilities, we generated mice with compound deletion of Lrp5 and LRP6 selectively in bone, and treated them with sclerostin monoclonal antibody (Scl-mAb). All mice were homozygous flox for both Lrp5 and LRP6 (Lrp5f/f; LRP6f/f), and varied only in whether or not they carried the Dmp1-Cre transgene. Positive (Cre+) and negative (Cre−) mice were injected with Scl-mAb or vehicle from 4.5 to 14 weeks of age. Vehicle-treated Cre+ mice exhibited significantly reduced skeletal properties compared to vehicle-treated Cre− mice, as assessed by DXA, μCT, pQCT, and histology, indicating that Lrp5/6 deletions were effective and efficient. Scl-mAb treatment improved nearly every bone-related parameter among Cre− mice, but the same treatment in Cre+ mice resulted in little to no improvement in skeletal properties. For the few endpoints where Cre+ mice responded to Scl-mAb, it is likely that antibody-induced promotion of Wnt signaling occurred in cell types earlier in the mesenchymal/osteoblast differentiation pathway than the Dmp1-expressing stage. This latter conclusion was supported by changes in some histomorphometric parameters. In conclusion, unlike with the deletion of Lrp5 alone, the bone-selective late-stage co-deletion of Lrp5 and LRP6 significantly impairs or completely nullifies the osteogenic action of Scl-mAb, and highlights a major role for both Lrp5 and LRP6 in the mechanism of action for the bone-building effects of sclerostin antibody.
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LRP5: From bedside to bench to bone
Bone, 2017Co-Authors: Bart O WilliamsAbstract:A role for low-density lipoprotein-related receptor 5 (LRP5) in human bone was first established by the identification of genetic alterations that led to dramatic changes in bone mass. Shortly thereafter, mutations that altered the function of the sclerostin (SOST) gene were also associated with altered human bone mass. Subsequent studies of LRP5 and sclerostin have provided important insights into the mechanisms by which these proteins regulate skeletal homeostasis. Sclerostin normally binds to LRP5 and the related LRP6 protein and prevents their activation by Wnts, the LRP5/LRP6 ligands. The interaction of sclerostin with LRP5 or LRP6 is facilitated by the LRP4 protein. Loss of LRP5 leads to defective osteoblast function and low bone mass, while loss of SOST or mutations in LRP5, which produce a protein that can no longer be bound by SOST, result in high bone mass. Insights gained from the use of genetically engineered mouse models are presented, as well as a brief summary of the status of antibodies in clinical trials that block the function of SOST as a mechanism to increase bone mass.
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Mice with a heterozygous LRP6 deletion have impaired fracture healing.
Bone research, 2016Co-Authors: Travis A. Burgers, Juan Vivanco, Juraj Zahatnansky, Andrew J Vander Moren, James J. Mason, Bart O WilliamsAbstract:Bone fracture non-unions, the failure of a fracture to heal, occur in 10%–20% of fractures and are a costly and debilitating clinical problem. The Wnt/β-catenin pathway is critical in bone development and fracture healing. Polymorphisms of linking low-density lipoprotein receptor-related protein 6 (LRP6), a Wnt-binding receptor, have been associated with decreased bone mineral density and fragility fractures, although this remains controversial. Mice with a homozygous deletion of LRP6 have severe skeletal abnormalities and are not viable, whereas mice with a heterozygous deletion have a combinatory effect with Lrp5 to decrease bone mineral density. As fracture healing closely models embryonic skeletal development, we investigated the process of fracture healing in mice heterozygous for LRP6 (LRP6+/−) and hypothesized that the heterozygous deletion of LRP6 would impair fracture healing. Mid-diaphyseal femur fractures were induced in LRP6+/− mice and wild-type controls (LRP6+/+). Fractures were analyzed using micro-computed tomography (μCT) scans, biomechanical testing, and histological analysis. LRP6+/− mice had significantly decreased stiffness and strength at 28 days post fracture (PF) and significantly decreased BV/TV, total density, immature bone density, and mature area within the callus on day-14 and -21 PF; they had significantly increased empty callus area at days 14 and 21 PF. Our results demonstrate that the heterozygous deletion of LRP6 impairs fracture healing, which suggests that LRP6 has a role in fracture healing. A gene defect linked to changes in bone mass in humans also impairs fracture healing in mouse models. The Wnt signaling pathway has emerged as a central regulator of skeletal remodeling, and some studies have found that people with mutations in one Wnt-associated gene called LRP6 have decreased bone mineral density and more fractures resulting from normal activities. To investigate the role of LRP6 in fracture healing, a team led by Bart Williams from the Van Andel Research Institute in Grand Rapids, Michigan, USA, bred mice with only one working copy of the gene. (Mice with two defective copies die before birth.) They induced fractures in the leg bone and four weeks later observed that the mutant mice had decreased density and volume in the healing bone, with fewer signs of tissue repair than normal mice.
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structure and function of norrin in assembly and activation of a frizzled 4 lrp5 6 complex
Genes & Development, 2013Co-Authors: Kaleeckal G Harikumar, Clara Erice, Chen Chen, Liren Wang, Naomi R Parker, Zhihong Cheng, Bart O Williams, Karsten Melcher, Laurence J MillerAbstract:Norrin is a cysteine-rich growth factor that is required for angiogenesis in the eye, ear, brain, and female reproductive organs. It functions as an atypical Wnt ligand by specifically binding to the Frizzled 4 (Fz4) receptor. Here we report the crystal structure of Norrin, which reveals a unique dimeric structure with each monomer adopting a conserved cystine knot fold. Functional studies demonstrate that the novel Norrin dimer interface is required for Fz4 activation. Furthermore, we demonstrate that Norrin contains separate binding sites for Fz4 and for the Wnt ligand coreceptor Lrp5 (low-density lipoprotein-related protein 5) or LRP6. Instead of inducing Fz4 dimerization, Norrin induces the formation of a ternary complex with Fz4 and Lrp5/6 by binding to their respective extracellular domains. These results provide crucial insights into the assembly and activation of the Norrin-Fz4-Lrp5/6 signaling complex.
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Lrp5 and LRP6 exert overlapping functions in osteoblasts during postnatal bone acquisition.
PloS one, 2013Co-Authors: Ryan C. Riddle, Cassandra R. Diegel, Julie M. Leslie, Kyle K. Van Koevering, Marie Claude Faugere, Thomas L. Clemens, Bart O WilliamsAbstract:The canonical Wnt signaling pathway is critical for skeletal development and maintenance, but the precise roles of the individual Wnt co-receptors, Lrp5 and LRP6, that enable Wnt signals to be transmitted in osteoblasts remain controversial. In these studies, we used Cre-loxP recombination, in which Cre-expression is driven by the human osteocalcin promoter, to determine the individual contributions of Lrp5 and LRP6 in postnatal bone acquisition and osteoblast function. Mice selectively lacking either Lrp5 or LRP6 in mature osteoblasts were born at the expected Mendelian frequency but demonstrated significant reductions in whole-body bone mineral density. Bone architecture measured by microCT revealed that LRP6 mutant mice failed to accumulate normal amounts of trabecular bone. By contrast, Lrp5 mutants had normal trabecular bone volume at 8 weeks of age, but with age, these mice also exhibited trabecular bone loss. Both mutants also exhibited significant alterations in cortical bone structure. In vitro differentiation was impaired in both Lrp5 and LRP6 null osteoblasts as indexed by alkaline phosphatase and Alizarin red staining, but the defect was more pronounced in LRP6 mutant cells. Mice lacking both Wnt co-receptors developed severe osteopenia similar to that observed previously in mice lacking β-catenin in osteoblasts. Likewise, calvarial cells doubly deficient for Lrp5 and LRP6 failed to form osteoblasts when cultured in osteogenic media, but instead attained a chondrocyte-like phenotype. These results indicate that expression of both Lrp5 and LRP6 are required within mature osteoblasts for normal postnatal bone development.
Alexander G. Robling - One of the best experts on this subject based on the ideXlab platform.
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Co-deletion of Lrp5 and LRP6 in the skeleton severely diminishes bone gain from sclerostin antibody administration.
Bone, 2020Co-Authors: Kyung-eun Lim, Bart O Williams, Whitney A. Bullock, Daniel J. Horan, Matthew L. Warman, Alexander G. RoblingAbstract:Abstract The cysteine knot protein sclerostin is an osteocyte-derived secreted inhibitor of the Wnt co-receptors LRP5 and LRP6. LRP5 plays a dominant role in bone homeostasis, but we previously reported that Sost/sclerostin suppression significantly increased osteogenesis regardless of Lrp5 presence or absence. Those observations suggested that the bone forming effects of sclerostin inhibition can occur through LRP6 (when Lrp5 is suppressed), or through other yet undiscovered mechanisms independent of Lrp5/6. To distinguish between these two possibilities, we generated mice with compound deletion of Lrp5 and LRP6 selectively in bone, and treated them with sclerostin monoclonal antibody (Scl-mAb). All mice were homozygous flox for both Lrp5 and LRP6 (Lrp5f/f; LRP6f/f), and varied only in whether or not they carried the Dmp1-Cre transgene. Positive (Cre+) and negative (Cre−) mice were injected with Scl-mAb or vehicle from 4.5 to 14 weeks of age. Vehicle-treated Cre+ mice exhibited significantly reduced skeletal properties compared to vehicle-treated Cre− mice, as assessed by DXA, μCT, pQCT, and histology, indicating that Lrp5/6 deletions were effective and efficient. Scl-mAb treatment improved nearly every bone-related parameter among Cre− mice, but the same treatment in Cre+ mice resulted in little to no improvement in skeletal properties. For the few endpoints where Cre+ mice responded to Scl-mAb, it is likely that antibody-induced promotion of Wnt signaling occurred in cell types earlier in the mesenchymal/osteoblast differentiation pathway than the Dmp1-expressing stage. This latter conclusion was supported by changes in some histomorphometric parameters. In conclusion, unlike with the deletion of Lrp5 alone, the bone-selective late-stage co-deletion of Lrp5 and LRP6 significantly impairs or completely nullifies the osteogenic action of Scl-mAb, and highlights a major role for both Lrp5 and LRP6 in the mechanism of action for the bone-building effects of sclerostin antibody.
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New Insights into Wnt–Lrp5/6–β-Catenin Signaling in Mechanotransduction
Frontiers in endocrinology, 2015Co-Authors: Kyung Shin Kang, Alexander G. RoblingAbstract:Mechanical loading is essential to maintain normal bone metabolism and the balance between bone formation and resorption. The cellular mechanisms that control mechanotransduction are not fully defined, but several key pathways have been identified. We discuss the roles of several components of the Wnt signaling cascade, namely Lrp5, LRP6, and β-catenin in mechanical loading-induced bone formation. Lrp5 is an important Wnt co-receptor for regulating bone mass and mechanotransduction, and appears to function principally by augmenting bone formation. LRP6 also regulates bone mass but its action might involve resorption as well as formation. The role of LRP6 in mechanotransduction is unclear. Studies addressing the role of β-catenin in bone metabolism and mechanotransduction highlight the uncertainties in downstream modulators of Lrp5 and LRP6. Taken together, these data indicate that mechanical loading might affect bone regulation triggering the canonical Wnt signaling (and perhaps other pathways) not only via Lrp5 but also via LRP6. Further work is needed to clarify the role of the Wnt signaling pathway in Lrp5 and/or LRP6-mediated mechanotransduction, which could eventually lead to powerful therapeutic agents that might mimic the anabolic effects of mechanical stimulation.
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new insights into wnt lrp5 6 β catenin signaling in mechanotransduction
Frontiers in Endocrinology, 2015Co-Authors: Kyung Shin Kang, Alexander G. RoblingAbstract:Mechanical loading is essential to maintain normal bone metabolism and the balance between bone formation and resorption. The cellular mechanisms that control mechanotransduction are not fully defined, but several key pathways have been identified. We discuss the roles of several components of the Wnt signaling cascade, namely Lrp5, LRP6, and β-catenin in mechanical loading-induced bone formation. Lrp5 is an important Wnt co-receptor for regulating bone mass and mechanotransduction, and appears to function principally by augmenting bone formation. LRP6 also regulates bone mass but its action might involve resorption as well as formation. The role of LRP6 in mechanotransduction is unclear. Studies addressing the role of β-catenin in bone metabolism and mechanotransduction highlight the uncertainties in downstream modulators of Lrp5 and LRP6. Taken together, these data indicate that mechanical loading might affect bone regulation triggering the canonical Wnt signaling (and perhaps other pathways) not only via Lrp5 but also via LRP6. Further work is needed to clarify the role of the Wnt signaling pathway in Lrp5 and/or LRP6-mediated mechanotransduction, which could eventually lead to powerful therapeutic agents that might mimic the anabolic effects of mechanical stimulation.
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Sclerostin inhibition reverses skeletal fragility in an Lrp5-deficient mouse model of OPPG syndrome.
Science translational medicine, 2013Co-Authors: Rajendra Kedlaya, Daniel J. Horan, Matthew L. Warman, Shreya Veera, Rachel E. Moss, Ugur M. Ayturk, Christina M. Jacobsen, Margot E. Bowen, Chris Paszty, Alexander G. RoblingAbstract:Osteoporosis pseudoglioma syndrome (OPPG) is a rare genetic disease that produces debilitating effects in the skeleton. OPPG is caused by mutations in LRP5, a WNT co-receptor that mediates osteoblast activity. WNT signaling through LRP5, and also through the closely related receptor LRP6, is inhibited by the protein sclerostin (SOST). It is unclear whether OPPG patients might benefit from the anabolic action of sclerostin neutralization therapy (an approach currently being pursued in clinical trials for postmenopausal osteoporosis) in light of their LRP5 deficiency and consequent osteoblast impairment. To assess whether loss of sclerostin is anabolic in OPPG, we measured bone properties in a mouse model of OPPG (Lrp5−/−), a mouse model of sclerosteosis (Sost−/−), and in mice with both genes knocked out (Lrp5−/−;Sost−/−). Lrp5−/−;Sost−/− mice have larger, denser, and stronger bones than do Lrp5−/− mice, indicating that SOST deficiency can improve bone properties via pathways that do not require LRP5. Next, we determined whether the anabolic effects of sclerostin depletion in Lrp5−/− mice are retained in adult mice by treating 17-week-old Lrp5−/− mice with a sclerostin antibody for 3 weeks. Lrp5+/+ and Lrp5−/− mice each exhibited osteoanabolic responses to antibody therapy, as indicated by increased bone mineral density, content, and formation rates. Collectively, our data show that inhibiting sclerostin can improve bone mass whether LRP5 is present or not. In the absence of LRP5, the anabolic effects of SOST depletion can occur via other receptors (such as LRP4/6). Regardless of the mechanism, our results suggest that humans with OPPG might benefit from sclerostin neutralization therapies.
Christof Niehrs - One of the best experts on this subject based on the ideXlab platform.
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RAB8B is required for activity and caveolar endocytosis of LRP6.
Cell reports, 2013Co-Authors: Kubilay Demir, Christof Niehrs, Dierk Ingelfinger, Nadine Kirsch, Carlo A. Beretta, Gerrit Erdmann, Enrico Moro, Francesco Argenton, Matthias Carl, Michael BoutrosAbstract:Summary Wnt/β-catenin signaling plays an important role in embryonic development and adult tissue homeostasis. When Wnt ligands bind to the receptor complex, LRP5/6 coreceptors are activated by phosphorylation and concomitantly endocytosed. In vertebrates, Wnt ligands induce caveolin-dependent endocytosis of LRP6 to relay signal downstream, whereas antagonists such as Dickkopf promote clathrin-dependent endocytosis, leading to inhibition. However, little is known about how LRP6 is directed to different internalization mechanisms, and how caveolin-dependent endocytosis is mediated. In an RNAi screen, we identified the Rab GTPase RAB8B as being required for Wnt/β-catenin signaling. RAB8B depletion reduces LRP6 activity, β-catenin accumulation, and induction of Wnt target genes, whereas RAB8B overexpression promotes LRP6 activity and internalization and rescues inhibition of caveolar endocytosis. In Xenopus laevis and Danio rerio , RAB8B morphants show lower Wnt activity during embryonic development. Our results implicate RAB8B as an essential evolutionary conserved component of Wnt/β-catenin signaling through regulation of LRP6 activity and endocytosis.
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Regulation of LRP6 phosphorylation
Cellular and molecular life sciences : CMLS, 2010Co-Authors: Christof Niehrs, Jinlong ShenAbstract:The Wnt/β-catenin signaling pathway plays important roles in embryonic development and tissue homeostasis, and is implicated in human disease. Wnts transduce signals via transmembrane receptors of the Frizzled (Fzd/Fz) family and the low density lipoprotein receptor-related protein 5/6 (Lrp5/6). A key mechanism in their signal transduction is that Wnts induce LRP6 signalosomes, which become phosphorylated at multiple conserved sites, notably at PPSPXS motifs. LRP6 phosphorylation is crucial to β-catenin stabilization and pathway activation by promoting Axin and Gsk3 recruitment to phosphorylated sites. Here, we summarize how proline-directed kinases (Gsk3, PKA, Pftk1, Grk5/6) and non-proline-directed kinases (CK1 family) act upon LRP6, how the phosphorylation is regulated by ligand binding and mitosis, and how LRP6 phosphorylation leads to β-catenin stabilization.
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O7. Regulation of Wnt/LRP6 signalling
Differentiation, 2010Co-Authors: Christof NiehrsAbstract:The Wnt/β-catenin signaling pathway plays important roles in embryonic development and disease. Wnts transduce signals via transmembrane receptors of the Frizzled (Fzd) family and the low density lipoprotein receptor-related protein 5/6 (Lrp5/6). A key mechanism in their signal transduction is that Wnts induce LRP6 signalosomes, which become phosphorylated at multiple conserved sites. LRP6 phosphorylation is crucial to beta-catenin stabilization and pathway activation by promoting Axin and Gsk3 recruitment. I will discuss the regulation of this co-receptor and the significance for Xenopus early development.
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Kremen is required for neural crest induction in Xenopus and promotes LRP6-mediated Wnt signaling
Development (Cambridge England), 2007Co-Authors: Christine Hassler, Cristina-maria Cruciat, Ya Lin Huang, Sei Kuriyama, Roberto Mayor, Christof NiehrsAbstract:Kremen 1 and 2 (Krm1/2) are transmembrane receptors for Wnt antagonists of the Dickkopf (Dkk) family and function by inhibiting the Wnt co-receptors LRP5/6. Here we show that Krm2 functions independently from Dkks during neural crest (NC) induction in Xenopus. Krm2 is co-expressed with, and regulated by, canonical Wnts. Krm2 is differentially expressed in the NC, and morpholino-mediated Krm2 knockdown inhibits NC induction, which is mimicked by LRP6 depletion. Conversely, krm2 overexpression induces ectopic NC. Kremens bind to LRP6, promote its cell-surface localization and stimulate LRP6 signaling. Furthermore, Krm2 knockdown specifically reduces LRP6 protein levels in NC explants. The results indicate that in the absence of Dkks, Kremens activate Wnt/β-catenin signaling through LRP6.
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casein kinase 1γ couples wnt receptor activation to cytoplasmic signal transduction
Nature, 2005Co-Authors: Gary Davidso, Jinlong She, Josipa Ilic, Ursula Fenge, Pete Stannek, Andrei Glinka, Christof NiehrsAbstract:Signalling by Wnt proteins (Wingless in Drosophila) has diverse roles during embryonic development and in adults, and is implicated in human diseases, including cancer. LDL-receptor-related proteins 5 and 6 (LRP5 and LRP6; Arrow in Drosophila) are key receptors required for transmission of Wnt/beta-catenin signalling in metazoa. Although the role of these receptors in Wnt signalling is well established, their coupling with the cytoplasmic signalling apparatus remains poorly defined. Using a protein modification screen for regulators of LRP6, we describe the identification of Xenopus Casein kinase 1 gamma (CK1gamma), a membrane-bound member of the CK1 family. Gain-of-function and loss-of-function experiments show that CK1gamma is both necessary and sufficient to transduce LRP6 signalling in vertebrates and Drosophila cells. In Xenopus embryos, CK1gamma is required during anterio-posterior patterning to promote posteriorizing Wnt/beta-catenin signalling. CK1gamma is associated with LRP6, which has multiple, modular CK1 phosphorylation sites. Wnt treatment induces the rapid CK1gamma-mediated phosphorylation of these sites within LRP6, which, in turn, promotes the recruitment of the scaffold protein Axin. Our results reveal an evolutionarily conserved mechanism that couples Wnt receptor activation to the cytoplasmic signal transduction apparatus.
Caroline M. Alexander - One of the best experts on this subject based on the ideXlab platform.
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Lrp5 Has a Wnt-Independent Role in Glucose Uptake and Growth for Mammary Epithelial Cells
Molecular and cellular biology, 2015Co-Authors: Emily N Chin, Soyoung Kim, Saja A Fakhraldeen, Joshua A. Martin, Caroline M. AlexanderAbstract:Lrp5 is typically described as a Wnt signaling receptor, albeit a less effective Wnt signaling receptor than the better-studied sister isoform, LRP6. Here we show that Lrp5 is only a minor player in the response to Wnt3a-type ligands in mammary epithelial cells; instead, Lrp5 is required for glucose uptake, and glucose uptake regulates the growth rate of mammary epithelial cells in culture. Thus, a loss of Lrp5 leads to profound growth suppression, whether growth is induced by serum or by specific growth factors, and this inhibition is not due to a loss of Wnt signaling. Depletion of Lrp5 decreases glucose uptake, lactate secretion, and oxygen consumption rates; inhibition of glucose consumption phenocopies the loss of Lrp5 function. Both Lrp5 knockdown and low external glucose induce mitochondrial stress, as revealed by the accumulation of reactive oxygen species (ROS) and the activation of the ROS-sensitive checkpoint, p38α. In contrast, loss of function of LRP6 reduces Wnt responsiveness but has little impact on growth. This highlights the distinct functions of these two Lrp receptors and an important Wnt ligand-independent role of Lrp5 in glucose uptake in mammary epithelial cells.
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both lrp5 and LRP6 receptors are required to respond to physiological wnt ligands in mammary epithelial cells and fibroblasts
Journal of Biological Chemistry, 2012Co-Authors: Shruti Goel, Emily N Chin, Saja A Fakhraldeen, Scott M Berry, David J Beebe, Caroline M. AlexanderAbstract:A canonical Wnt signal maintains adult mammary ductal stem cell activity, and this signal requires the Wnt signaling reception, LRP5. However, previous data from our laboratory have shown that LRP5 and LRP6 are co-expressed in mammary basal cells and that LRP6 is active, leading us to question why LRP6 is insufficient to mediate canonical signaling in the absence of LRP5. Here, we show that at endogenous levels of LRP5 and LRP6 both receptors are required to signal in response to some Wnt ligands both in vitro (in mouse embryonic fibroblasts and mammary epithelial cells) and in vivo (in mammary outgrowths). This subgroup of canonical ligands includes Wnt1, Wnt9b, and Wnt10b; the latter two are expressed in mammary gland. In contrast, the ligand commonly used experimentally, Wnt3a, prefers LRP6 and requires just one receptor regardless of cellular context. When either LRP5 or LRP6 is overexpressed, signaling remains ligand-dependent, but the requirement for both receptors is abrogated (regardless of ligand type). We have documented an LRP5-6 heteromer using immiscible filtration assisted by surface tension (IFAST) immunoprecipitation. Together, our data imply that under physiological conditions some Wnt ligands require both receptors to be present to generate a canonical signal. We have designed a model to explain our results based on the resistance of LRP5-6 heteromers to a selective inhibitor of E1/2-binding Wnt-LRP6 interaction. These data have implications for stem cell biology and for the analysis of the oncogenicity of LRP receptors that are often overexpressed in breast tumors.
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The Wnt Receptor, Lrp5, Is Expressed by Mouse Mammary Stem Cells and Is Required to Maintain the Basal Lineage
PloS one, 2009Co-Authors: Nisha M. Badders, Bart O Williams, Shruti Goel, Rod J. Clark, Kristine S. Klos, Soyoung Kim, Anna Bafico, Charlotta Lindvall, Caroline M. AlexanderAbstract:Background: Ectopic Wnt signaling induces increased stem/progenitor cell activity in the mouse mammary gland, followed by tumor development. The Wnt signaling receptors, Lrp5/6, are uniquely required for canonical Wnt activity. Previous data has shown that the absence of Lrp5 confers resistance to Wnt1-induced tumor development. Methodology/Principal Findings: Here, we show that all basal mammary cells express Lrp5, and co-express LRP6 in a similar fashion. Though Wnt dependent transcription of key target genes is relatively unchanged in mammary epithelial cell cultures, the absence of Lrp5 specifically depletes adult regenerative stem cell activity (to less than 1%). Stem cell activity can be enriched by .200 fold (over 80% of activity), based on high Lrp5 expression alone. Though Lrp5 null glands have apparent normal function, the basal lineage is relatively reduced (from 42% basal/total epithelial cells to 22%) and Lrp52/2 mammary epithelial cells show enhanced expression of senescence-associated markers in vitro, as measured by expression of p16 Ink4a and TA-p63. Conclusions/Significance: This is the first single biomarker that has been demonstrated to be functionally involved in stem cell maintenance. Together, these results demonstrate that Wnt signaling through Lrp5 is an important component of normal mammary stem cell function.
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The role of Lrp5 and LRP6 in mouse mammary development and Wnt-1-induced carcinogenesis
Cancer Research, 2005Co-Authors: Charlotta Lindvall, Troy A. Giambernardi, Cassandra R. Zylstra, Caroline M. Alexander, Nicole Evans, Michael T. Lewis, Bart O WilliamsAbstract:Proc Amer Assoc Cancer Res, Volume 46, 2005 1075 LRP5 and LRP6 form a distinct subfamily of low-density lipoprotein receptor (LDLR)-like proteins. They are co-receptors for Wnt ligands, which are a large family of secreted glycoproteins that initiate signaling by binding to members of the Frizzled (Fz) family of seven transmembrane receptors and to either Lrp5 or LRP6. We have used mice carrying mutations in Lrp5 and LRP6 to gain insight into the normal roles of the two genes in mammary development and in Wnt1-induced mammary tumorigenesis. Our results show that loss of either Lrp5 or LRP6 impairs normal mammary development. The most severe phenotype is obtained in Lrp5-/-;LRP6+/− virgin mice, who exhibit mammary glands lacking a ductal network. Besides a delay in development, the ductal network of adult virgin Lrp5 -mutant mice display a reduction in side branching. Because LRP6 -mutant mice die shortly after birth, we have examined mammary tissue from 19-day-old embryos. The mammary glands of LRP6 -mutant embryos contain an epithelial sprout, but in contrast to wild type the mutants lack side branching. Interestingly, LRP6 -mutant embryos seem to lack mammary fat pads. Female mice expressing the Wnt-1 gene under the control of the mouse mammary tumor virus (MMTV) long terminal repeats reproducibly develop mammary adenocarcinomas at approximately six months of age. To investigate the requirement of Lrp5 and LRP6 in Wnt-induced carcinogenesis we have generated MMTV-Wnt-1 transgenic female mice that are deficient for Lrp5. These homozygotes exhibit an almost complete resistance to Wnt-1-induced mammary tumors and the heterozygotes (Lrp5+/−) display a significant delay in tumorigenesis. After 300 days, less than 10% of the MMTV-Wnt-1;Lrp5 -mutant and more than 90% of the MMTV-Wnt-1;Lrp5 wild type mice had developed mammary tumors. The tumors that arise in a Lrp5 -deficient background are more heterogeneous than normal Wnt-1 tumors, and frequently display a papillary growth pattern. Current work is aimed at characterizing the differences in the tumors at the cellular and molecular level.
Vítězslav Bryja - One of the best experts on this subject based on the ideXlab platform.
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Genetic interaction between LRP6 and Wnt5a during mouse development.
Developmental Dynamics, 2009Co-Authors: Emma R. Andersson, Lenka Bryjova, Kristin K. Biris, Terry P. Yamaguchi, Ernest Arenas, Vítězslav BryjaAbstract:LRP6 is generally described as a receptor required for signal transduction in the Wnt/β-catenin pathway. Wnt5a, however, is a Wnt ligand that usually does not activate Wnt/β-catenin but rather activates noncanonical Wnt signaling. We have previously shown that LRP6 can inhibit noncanonical Wnt5a/Wnt11 signaling and that Lrp5/6 loss-of-function produces noncanonical gain-of function defects, which can be rescued by loss of Wnt5a. Here, we describe other phenotypes found in Wnt5a/LRP6 compound mutant mice, including a worsening of individual Wnt5a or LRP6 loss of function phenotypes. LRP6 haploinsufficiency in a Wnt5a−/− background caused spina bifida and exacerbated posterior truncation. Wnt5a−/−LRP6−/− embryos displayed presomitic mesoderm morphogenesis, somitogenesis, and neurogenesis defects, which are much more severe than in either of the single mutants. Interestingly these results reveal a further level of complexity in processes in which Wnt5a and LRP6 cooperate, or oppose each other, during mouse development. Developmental Dynamics 239:237–245, 2010. Published 2009 Wiley-Liss, Inc.
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The Extracellular Domain of Lrp5/6 Inhibits Noncanonical Wnt Signaling In Vivo
Molecular biology of the cell, 2008Co-Authors: Vítězslav Bryja, Lenka Bryjova, Kristin K. Biris, Emma R. Andersson, Alexandra Schambony, Milan Esner, Anita C. Hall, Bianca Kraft, Lukas Cajanek, Terry P. YamaguchiAbstract:Lrp5/6 are crucial coreceptors for Wnt/β-catenin signaling, a pathway biochemically distinct from noncanonical Wnt signaling pathways. Here, we examined the possible participation of Lrp5/6 in noncanonical Wnt signaling. We found that LRP6 physically interacts with Wnt5a, but that this does not lead to phosphorylation of LRP6 or activation of the Wnt/β-catenin pathway. Overexpression of LRP6 blocks activation of the Wnt5a downstream target Rac1, and this effect is dependent on intact LRP6 extracellular domains. These results suggested that the extracellular domain of LRP6 inhibits noncanonical Wnt signaling in vitro. In vivo, LRP6−/− mice exhibited exencephaly and a heart phenotype. Surprisingly, these defects were rescued by deletion of Wnt5a, indicating that the phenotypes resulted from noncanonical Wnt gain-of-function. Similarly, Lrp5 and LRP6 antisense morpholino-treated Xenopus embryos exhibited convergent extension and heart phenotypes that were rescued by knockdown of noncanonical XWnt5a and XWnt11. Thus, we provide evidence that the extracellular domains of Lrp5/6 behave as physiologically relevant inhibitors of noncanonical Wnt signaling during Xenopus and mouse development in vivo.
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the extracellular domain of lrp5 6 inhibits noncanonical wnt signaling in vivo
Molecular Biology of the Cell, 2008Co-Authors: Lenka Bryjova, Kristin K. Biris, Vítězslav Bryja, Emma R. Andersson, Alexandra Schambony, Milan Esner, Anita C. Hall, Bianca KraftAbstract:Lrp5/6 are crucial coreceptors for Wnt/β-catenin signaling, a pathway biochemically distinct from noncanonical Wnt signaling pathways. Here, we examined the possible participation of Lrp5/6 in noncanonical Wnt signaling. We found that LRP6 physically interacts with Wnt5a, but that this does not lead to phosphorylation of LRP6 or activation of the Wnt/β-catenin pathway. Overexpression of LRP6 blocks activation of the Wnt5a downstream target Rac1, and this effect is dependent on intact LRP6 extracellular domains. These results suggested that the extracellular domain of LRP6 inhibits noncanonical Wnt signaling in vitro. In vivo, LRP6−/− mice exhibited exencephaly and a heart phenotype. Surprisingly, these defects were rescued by deletion of Wnt5a, indicating that the phenotypes resulted from noncanonical Wnt gain-of-function. Similarly, Lrp5 and LRP6 antisense morpholino-treated Xenopus embryos exhibited convergent extension and heart phenotypes that were rescued by knockdown of noncanonical XWnt5a and XWnt11. Thus, we provide evidence that the extracellular domains of Lrp5/6 behave as physiologically relevant inhibitors of noncanonical Wnt signaling during Xenopus and mouse development in vivo.
