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Michael Reedijk - One of the best experts on this subject based on the ideXlab platform.
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cyclin d1 is a direct target of JAG1 mediated notch signaling in breast cancer
Breast Cancer Research and Treatment, 2010Co-Authors: Brenda Cohen, Mamiko Shimizu, Julia Izrailit, Nancy F Ng, Yuri Buchman, Judy Dering, Michael ReedijkAbstract:The Notch ligand, JAG1 is associated with breast cancer recurrence. Herein, we report on a genomics approach to elucidate mechanisms downstream of JAG1 that promote breast cancer growth. In a survey of 46 breast cancer cell lines, we found that triple negative (TN; basal and mesenchymal ER-, PR-, and Her2-negative) lines express JAG1 at significantly higher levels than do HER2+ or luminal (ER+) Her2− cell lines. In contrast to the luminal lines tested (T47D and MCF7), TN breast cancer cell lines (HCC1143 and MDA MB231) display high-level JAG1 expression and growth inhibition with RNA interference-induced JAG1 down-regulation. We used microarray profiling of TN tumor cells transfected with JAG1 siRNA to identify JAG1-regulated genes (P ≤ 0.005; fold change ≥1.5). Among JAG1-regulated genes identified, cyclin D1 was found to be a direct target of NOTCH1 and NOTCH3. We show that JAG1 down-regulation reduces direct binding of Notch to the cyclin D1 promoter, reduced cyclin D1 expression and inhibition of cell cycle progression through the cyclin D1-dependant G1/S checkpoint. Furthermore, we show that cyclin D1 and JAG1 expression correlate in TN breast cancer expression datasets. These data suggest a model whereby JAG1 promotes cyclin D1-mediated proliferation of TN breast cancers.
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JAG1 expression is associated with a basal phenotype and recurrence in lymph node negative breast cancer
Breast Cancer Research and Treatment, 2008Co-Authors: Michael Reedijk, Brendan C Dickson, Anna Marie Mulligan, Hui Zhang, Dushanthi Pinnaduwage, Shelley B Bull, Frances P OmalleyAbstract:Expression of the JAG1 Notch ligand has previously been shown to correlate with poor overall survival in women with advanced breast cancer. We undertook to test whether expression of JAG1 is associated with reduced disease free survival (DFS) in 887 samples from a prospectively accrued LNN cohort with a median follow-up greater than 8 years. Moderate to high JAG1 mRNA expression was associated with reduced DFS in univariate analysis (hazard ratio of 1.58; 95% confidence interval, 1.03–2.40; P = 0.034) and correlated with large tumor size, ER and PgR negativity, high tumor grade, and p53 antibody reactivity. Although elevated risk of reduced DFS in patients with high JAG1 mRNA did not persist with adjustment for other prognostic factors, it did in combination with HER2. JAG1 mRNA was positively associated with expression of basal breast cancer markers, however, in contrast to the finding that basal gene expression is most strongly associated with reduced DFS in the first 36 months of follow-up, JAG1 mRNA expression was associated with reduced DFS through the full follow-up period. Also, tumors expressing high levels of both mRNA and protein showed reduced DFS as compared to all other groups in univariate analysis (hazard ratio of 1.73; 95% confidence interval, 1.09–2.74; P = 0.020). Thus, JAG1 expression is associated with poor DFS in LNN breast cancer. As JAG1 is a target of several oncogenic signaling pathways, and is a ligand for Notch, these data provide novel insights into signaling that may contribute to progression of early stage breast cancer.
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high level JAG1 mrna and protein predict poor outcome in breast cancer
Modern Pathology, 2007Co-Authors: Brendan C Dickson, Michael Reedijk, Anna Marie Mulligan, Hui Zhang, G Lockwood, Frances P Omalley, Sean E EganAbstract:Notch receptors regulate cell fate determination, stem cell self-renewal, proliferation and apoptosis. We previously reported that elevated mRNA expression of the Notch ligand JAG1 identifies breast cancer patients with a poor prognosis. Here we show through immunohistochemical analysis of the same breast cancer cases (N=127) that patients with tumors expressing high levels of JAG1 protein had a worse outcome than those with tumors expressing low levels (10-year survival 26 vs 48%, and median survival 63 vs 108 months, respectively; P=0.03). We also describe the novel application of the Allred score to quantify JAG1 mRNA and protein expression levels. Using the Allred score, patients with tumors expressing high levels of JAG1 mRNA had a worse outcome than those with tumors expressing low levels (10-year survival 16 vs 47%, and median survival 43 months vs 100 months, respectively; P<0.001). Interestingly, when tumors were classified as either high or low for JAG1 mRNA or protein expression, there was only 65% agreement (κ=0.08) between the two methods of expression analysis. When JAG1 mRNA and protein data were combined, patients with tumors expressing low levels of both had a 10-year survival of 53% and median survival of 131 months. In comparison, patients with tumors expressing either high levels of JAG1 protein, mRNA or both had reduced 10-year survival and median survival (31%, 19%, 11% and 77, 43, 23 months respectively; P<0.0001). There was marginal evidence of an interaction effect (P=0.055), which indicated that the prognostic value of JAG1 protein was limited to the JAG1 mRNA-low subgroup. These data show that the Allred score can be used to rapidly quantify JAG1 mRNA and protein levels in breast cancer to identify patients who have a significant survival disadvantage and who may benefit from therapies (such as γ-secretase inhibitors) that target signaling through the Notch pathway.
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High-level JAG1 mRNA and protein predict poor outcome in breast cancer
Modern Pathology, 2007Co-Authors: Brendan C Dickson, Anna Marie Mulligan, Hui Zhang, G Lockwood, Sean E Egan, Frances P. O'malley, Michael ReedijkAbstract:Notch receptors regulate cell fate determination, stem cell self-renewal, proliferation and apoptosis. We previously reported that elevated mRNA expression of the Notch ligand JAG1 identifies breast cancer patients with a poor prognosis. Here we show through immunohistochemical analysis of the same breast cancer cases (N=127) that patients with tumors expressing high levels of JAG1 protein had a worse outcome than those with tumors expressing low levels (10-year survival 26 vs 48%, and median survival 63 vs 108 months, respectively; P=0.03). We also describe the novel application of the Allred score to quantify JAG1 mRNA and protein expression levels. Using the Allred score, patients with tumors expressing high levels of JAG1 mRNA had a worse outcome than those with tumors expressing low levels (10-year survival 16 vs 47%, and median survival 43 months vs 100 months, respectively; P
Kathleen M Loomes - One of the best experts on this subject based on the ideXlab platform.
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jagged1 JAG1 structure expression and disease associations
Gene, 2016Co-Authors: Christopher M Grochowski, Kathleen M Loomes, Nancy B SpinnerAbstract:Jagged1 (JAG1) is one of the 5 cell surface ligands that functions primarily in the highly conserved Notch signaling pathway. Notch signaling plays a critical role in cellular fate determination and is active throughout development and across many organ systems. The classic JAG1-NOTCH interaction leads to a cascade of proteolytic cleavages resulting in the NOTCH intracellular domain being transported into the nucleus where it functions to activate downstream transcription of target genes. JAG1 mutations have been associated with several disorders including the multi-system dominant disorder Alagille syndrome, and some cases of tetralogy of Fallot (although these may represent variable expressivity of Alagille syndrome). In addition, variations in JAG1 have been found to be associated with multiple types of cancer including breast cancer and adrenocortical carcinoma. Alagille syndrome, which primarily affects the liver, heart, skeleton, eye, face, kidney and vasculature is caused by loss of function mutations in JAG1, demonstrating that haploinsufficiency for JAG1 is disease causing, at least in these tissues. Expression and conditional gene knockout studies of JAG1 (JAG1) have correlated with tissue-specific disease phenotypes and have provided insight into both disease pathogenesis and human development.
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microarray data reveal relationship between JAG1 and ddr1 in mouse liver
PLOS ONE, 2013Co-Authors: Lara A Underkoffler, Anthony Nelson, Erikka Carr, Matthew J Ryan, Reiner Schultz, Kathleen M LoomesAbstract:Alagille syndrome is an autosomal dominant disorder involving bile duct paucity and cholestasis in addition to cardiac, skeletal, ophthalmologic, renal and vascular manifestations. Mutations in JAG1, encoding a ligand in the Notch signaling pathway, are found in 95% of patients meeting clinical criteria for Alagille syndrome. In order to define the role of JAG1 in the bile duct developmental abnormalities seen in ALGS, we previously created a JAG1 conditional knockout mouse model. Mice heterozygous for the JAG1 conditional and null alleles demonstrate abnormalities in postnatal bile duct growth and remodeling, with portal expansion and increased numbers of malformed bile ducts. In this study we report the results of microarray analysis and identify genes and pathways differentially expressed in the JAG1 conditional/null livers as compared with littermate controls. In the initial microarray analysis, we found that many of the genes up-regulated in the JAG1 conditional/null mutant livers were related to extracellular matrix (ECM) interactions, cell adhesion and cell migration. One of the most highly up-regulated genes was Ddr1, encoding a receptor tyrosine kinase (RTK) belonging to a large RTK family. We have found extensive co-localization of JAG1 and Ddr1 in bile ducts and blood vessels in postnatal liver. In addition, co-immunoprecipitation data provide evidence for a novel protein interaction between JAG1 and Ddr1. Further studies will be required to define the nature of this interaction and its functional consequences, which may have significant implications for bile duct remodeling and repair of liver injury.
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bile duct proliferation in liver specific JAG1 conditional knockout mice effects of gene dosage
Hepatology, 2007Co-Authors: Kathleen M Loomes, Pierre Russo, Matt E Ryan, Anthony Nelson, Lara A Underkoffler, Curtis L Glover, Hong FuAbstract:The Notch signaling pathway is involved in determination of cell fate and control of cell proliferation in multiple organ systems. JAG1 encodes a ligand in the Notch pathway and has been identified as the disease-causing gene for the developmental disorder Alagille syndrome. Evidence from the study of human disease and mouse models has implicated JAG1 as having an important role in the development of bile ducts. We have derived a conditional knockout allele (JAG1loxP) to study the role of JAG1 and Notch signaling in liver and bile duct development. We crossed JAG1loxP mice with a transgenic line carrying Cre recombinase under the control of the albumin promoter and α-fetoprotein enhancer to ablate JAG1 in hepatoblasts. The liver-specific JAG1 conditional knockout mice showed normal bile duct development. To further decrease Notch pathway function, we crossed the JAG1 conditional knockout mice with mice carrying the hypomorphic Notch2 allele, and bile duct anatomy remained normal. When JAG1 conditional mice were crossed with mice carrying the JAG1 null allele, the adult progeny exhibited striking bile duct proliferation. Conclusion: These results indicate that Notch signaling in the liver is sensitive to JAG1 gene dosage and suggest a role for the Notch pathway in postnatal growth and morphogenesis of bile ducts. (HEPATOLOGY 2007.)
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Bile duct proliferation in liver‐specific JAG1 conditional knockout mice: Effects of gene dosage
Hepatology, 2007Co-Authors: Kathleen M Loomes, Thomas Gridley, Pierre Russo, Matt E Ryan, Anthony Nelson, Lara A Underkoffler, Curtis L Glover, Hong Fu, Klaus H. Kaestner, Rebecca J. OakeyAbstract:The Notch signaling pathway is involved in determination of cell fate and control of cell proliferation in multiple organ systems. JAG1 encodes a ligand in the Notch pathway and has been identified as the disease-causing gene for the developmental disorder Alagille syndrome. Evidence from the study of human disease and mouse models has implicated JAG1 as having an important role in the development of bile ducts. We have derived a conditional knockout allele (JAG1loxP) to study the role of JAG1 and Notch signaling in liver and bile duct development. We crossed JAG1loxP mice with a transgenic line carrying Cre recombinase under the control of the albumin promoter and α-fetoprotein enhancer to ablate JAG1 in hepatoblasts. The liver-specific JAG1 conditional knockout mice showed normal bile duct development. To further decrease Notch pathway function, we crossed the JAG1 conditional knockout mice with mice carrying the hypomorphic Notch2 allele, and bile duct anatomy remained normal. When JAG1 conditional mice were crossed with mice carrying the JAG1 null allele, the adult progeny exhibited striking bile duct proliferation. Conclusion: These results indicate that Notch signaling in the liver is sensitive to JAG1 gene dosage and suggest a role for the Notch pathway in postnatal growth and morphogenesis of bile ducts. (HEPATOLOGY 2007.)
Lara A Underkoffler - One of the best experts on this subject based on the ideXlab platform.
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microarray data reveal relationship between JAG1 and ddr1 in mouse liver
PLOS ONE, 2013Co-Authors: Lara A Underkoffler, Anthony Nelson, Erikka Carr, Matthew J Ryan, Reiner Schultz, Kathleen M LoomesAbstract:Alagille syndrome is an autosomal dominant disorder involving bile duct paucity and cholestasis in addition to cardiac, skeletal, ophthalmologic, renal and vascular manifestations. Mutations in JAG1, encoding a ligand in the Notch signaling pathway, are found in 95% of patients meeting clinical criteria for Alagille syndrome. In order to define the role of JAG1 in the bile duct developmental abnormalities seen in ALGS, we previously created a JAG1 conditional knockout mouse model. Mice heterozygous for the JAG1 conditional and null alleles demonstrate abnormalities in postnatal bile duct growth and remodeling, with portal expansion and increased numbers of malformed bile ducts. In this study we report the results of microarray analysis and identify genes and pathways differentially expressed in the JAG1 conditional/null livers as compared with littermate controls. In the initial microarray analysis, we found that many of the genes up-regulated in the JAG1 conditional/null mutant livers were related to extracellular matrix (ECM) interactions, cell adhesion and cell migration. One of the most highly up-regulated genes was Ddr1, encoding a receptor tyrosine kinase (RTK) belonging to a large RTK family. We have found extensive co-localization of JAG1 and Ddr1 in bile ducts and blood vessels in postnatal liver. In addition, co-immunoprecipitation data provide evidence for a novel protein interaction between JAG1 and Ddr1. Further studies will be required to define the nature of this interaction and its functional consequences, which may have significant implications for bile duct remodeling and repair of liver injury.
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bile duct proliferation in JAG1 fringe heterozygous mice identifies candidate modifiers of the alagille syndrome hepatic phenotype
Hepatology, 2008Co-Authors: Anthony Nelson, Lara A Underkoffler, Matthew J Ryan, Christina Bales, Dorian M Gonzalez, Michelle Segalov, Jeanne Wilsonrawls, Susan E ColeAbstract:Alagille syndrome (AGS) is a heterogeneous developmental disorder associated with bile duct paucity and various organ anomalies. The syndrome is caused by mutations in JAG1, which encodes a ligand in the Notch signaling pathway, in the majority of cases and mutations in the NOTCH2 receptor gene in less than 1% of patients. Although a wide array of JAG1 mutations have been identified in the AGS population, these mutational variants have not accounted for the wide phenotypic variability observed in patients with this syndrome. The Fringe genes encode glycosyltransferases, which modify Notch and alter ligand-receptor affinity. In this study, we analyzed double heterozygous mouse models to examine the Fringe genes as potential modifiers of the Notch-mediated hepatic phenotype observed in AGS. We generated mice that were haploinsufficient for both JAG1 and one of three paralogous Fringe genes: Lunatic (Lfng), Radical (Rfng), and Manic (Mfng). Adult JAG1+/−Lfng+/− and JAG1+/−Rfng+/− mouse livers exhibited widespread bile duct proliferation beginning at 5 weeks of age and persisting up to 1 year. The JAG1+/−Mfng+/− livers showed a subtle, yet significant increase in bile duct numbers and bile duct to portal tract ratios. These abnormalities were not observed in the newborn period. Despite the portal tract expansion by bile ducts, fibrosis was not increased and epithelial to mesenchymal transition was not shown in the affected portal tracts. Conclusion: Mice heterozygous for mutations in JAG1 and the Fringe genes display striking bile duct proliferation, which is not apparent at birth. These findings suggest that the Fringe genes may regulate postnatal bile duct growth and remodeling, and serve as candidate modifiers of the hepatic phenotype in AGS. (HEPATOLOGY 2008;48:1989–1997.)
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Bile duct proliferation in JAG1/fringe heterozygous mice identifies candidate modifiers of the Alagille syndrome hepatic phenotype.
Hepatology, 2008Co-Authors: Matthew J Ryan, Anthony Nelson, Lara A Underkoffler, Christina Bales, Dorian M Gonzalez, Michelle Segalov, Susan E Cole, Jeanne Wilson-rawls, Jennifer L. Moran, Pierre RussoAbstract:Alagille syndrome (AGS) is a heterogeneous developmental disorder associated with bile duct paucity and various organ anomalies. The syndrome is caused by mutations in JAG1, which encodes a ligand in the Notch signaling pathway, in the majority of cases and mutations in the NOTCH2 receptor gene in less than 1% of patients. Although a wide array of JAG1 mutations have been identified in the AGS population, these mutational variants have not accounted for the wide phenotypic variability observed in patients with this syndrome. The Fringe genes encode glycosyltransferases, which modify Notch and alter ligand-receptor affinity. In this study, we analyzed double heterozygous mouse models to examine the Fringe genes as potential modifiers of the Notch-mediated hepatic phenotype observed in AGS. We generated mice that were haploinsufficient for both JAG1 and one of three paralogous Fringe genes: Lunatic (Lfng), Radical (Rfng), and Manic (Mfng). Adult JAG1+/−Lfng+/− and JAG1+/−Rfng+/− mouse livers exhibited widespread bile duct proliferation beginning at 5 weeks of age and persisting up to 1 year. The JAG1+/−Mfng+/− livers showed a subtle, yet significant increase in bile duct numbers and bile duct to portal tract ratios. These abnormalities were not observed in the newborn period. Despite the portal tract expansion by bile ducts, fibrosis was not increased and epithelial to mesenchymal transition was not shown in the affected portal tracts. Conclusion: Mice heterozygous for mutations in JAG1 and the Fringe genes display striking bile duct proliferation, which is not apparent at birth. These findings suggest that the Fringe genes may regulate postnatal bile duct growth and remodeling, and serve as candidate modifiers of the hepatic phenotype in AGS. (HEPATOLOGY 2008;48:1989–1997.)
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bile duct proliferation in liver specific JAG1 conditional knockout mice effects of gene dosage
Hepatology, 2007Co-Authors: Kathleen M Loomes, Pierre Russo, Matt E Ryan, Anthony Nelson, Lara A Underkoffler, Curtis L Glover, Hong FuAbstract:The Notch signaling pathway is involved in determination of cell fate and control of cell proliferation in multiple organ systems. JAG1 encodes a ligand in the Notch pathway and has been identified as the disease-causing gene for the developmental disorder Alagille syndrome. Evidence from the study of human disease and mouse models has implicated JAG1 as having an important role in the development of bile ducts. We have derived a conditional knockout allele (JAG1loxP) to study the role of JAG1 and Notch signaling in liver and bile duct development. We crossed JAG1loxP mice with a transgenic line carrying Cre recombinase under the control of the albumin promoter and α-fetoprotein enhancer to ablate JAG1 in hepatoblasts. The liver-specific JAG1 conditional knockout mice showed normal bile duct development. To further decrease Notch pathway function, we crossed the JAG1 conditional knockout mice with mice carrying the hypomorphic Notch2 allele, and bile duct anatomy remained normal. When JAG1 conditional mice were crossed with mice carrying the JAG1 null allele, the adult progeny exhibited striking bile duct proliferation. Conclusion: These results indicate that Notch signaling in the liver is sensitive to JAG1 gene dosage and suggest a role for the Notch pathway in postnatal growth and morphogenesis of bile ducts. (HEPATOLOGY 2007.)
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Bile duct proliferation in liver‐specific JAG1 conditional knockout mice: Effects of gene dosage
Hepatology, 2007Co-Authors: Kathleen M Loomes, Thomas Gridley, Pierre Russo, Matt E Ryan, Anthony Nelson, Lara A Underkoffler, Curtis L Glover, Hong Fu, Klaus H. Kaestner, Rebecca J. OakeyAbstract:The Notch signaling pathway is involved in determination of cell fate and control of cell proliferation in multiple organ systems. JAG1 encodes a ligand in the Notch pathway and has been identified as the disease-causing gene for the developmental disorder Alagille syndrome. Evidence from the study of human disease and mouse models has implicated JAG1 as having an important role in the development of bile ducts. We have derived a conditional knockout allele (JAG1loxP) to study the role of JAG1 and Notch signaling in liver and bile duct development. We crossed JAG1loxP mice with a transgenic line carrying Cre recombinase under the control of the albumin promoter and α-fetoprotein enhancer to ablate JAG1 in hepatoblasts. The liver-specific JAG1 conditional knockout mice showed normal bile duct development. To further decrease Notch pathway function, we crossed the JAG1 conditional knockout mice with mice carrying the hypomorphic Notch2 allele, and bile duct anatomy remained normal. When JAG1 conditional mice were crossed with mice carrying the JAG1 null allele, the adult progeny exhibited striking bile duct proliferation. Conclusion: These results indicate that Notch signaling in the liver is sensitive to JAG1 gene dosage and suggest a role for the Notch pathway in postnatal growth and morphogenesis of bile ducts. (HEPATOLOGY 2007.)
Thomas Gridley - One of the best experts on this subject based on the ideXlab platform.
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Generation of mice with a conditional null allele of the Jagged2 gene
Genesis, 2010Co-Authors: Jingxia Xu, Luke T. Krebs, Thomas GridleyAbstract:The Notch signaling pathway is an evolutionarily conserved intercellular signaling mechanism. Mutations in Notch pathway components disrupt embryonic development in diverse multicellular organisms and cause cancers and inherited disease syndromes in humans (Bray, 2006; Fiuza and Arias, 2007). In mammals, genes of the Notch family (Notch1 through Notch4) encode Type 1 transmembrane protein receptors that interact with Type 1 transmembrane ligands encoded by genes of the Delta-like (Dll1, Dll3 and Dll4) and Jagged (JAG1 and Jag2) families. We have previously described the construction and analysis of a targeted null allele of the Jagged2 (Jag2) gene, termed Jag2del1 (originally named Jag2ΔDSL) (Jiang et al., 1998). Jag2del1/Jag2del1 homozygous mutant mice die at birth from completely penetrant cleft palate. The Jag2del1/Jag2del1 mice also exhibit soft tissue syndactyly, the fusion of the digits on the fore- and hindlimbs (Jiang et al., 1998). Cleft palate and soft tissue syndactyly are also exhibited at reduced penetrance by mice homozygous for the syndactylism mutation, a spontaneous Jag2 missense mutant allele (Jag2sm) that behaves genetically as a Jag2 hypomorphic allele (Casey et al., 2006; Sidow et al., 1997). To circumvent the neonatal lethality exhibited by Jag2del1/Jag2del1 null mutant mice and permit the study of Jag2 function in postnatal and adult mice, we describe here construction of an allele for conditional inactivation of Jag2 gene function using the Cre-loxP system. The Jag2 gene spans approximately 21 kb on mouse Chromosome 12, and consists of 26 exons. To generate the Jag2floxneo targeting vector (Fig. 1a), a PGKneo selection cassette was introduced into intron 2. The PGKneo cassette was flanked by FRT sites for removal by Flpe recombinase (Farley et al., 2000), and by a loxP site distal to the PGKneo cassette in intron 2. A second loxP site was introduced approximately 1.1 kb upstream of the Jag2 translational start site. A diphtheria toxin cassette, at the terminus of the targeting vector, was introduced for negative selection against random integration of the targeting vector into the ES cell genome. The design of the Jag2floxneo allele permits removal of the PGKneo cassette by mating to a Flpe deleter mouse line (generating the Jag2flox allele), or removal of Jag2 genomic sequence (including promoter sequences and exons 1 and 2) and the PGKneo cassette by mating to a Cre deleter line. To distinguish the Jag2 mutant allele generated by Cre-mediated deletion of the Jag2flox or Jag2floxneo alleles from the Jag2del1 allele, our previously published Jag2 targeted null mutant allele (Jiang et al., 1998), we designate the Jag2 null allele generated by Cre recombinase-mediated deletion of the Jag2flox or Jag2floxneo alleles the Jag2del2 allele. FIG. 1 Generation of a Jag2 conditional null allele. (a) Schematic representation of a portion of the wildtype Jag2 allele, the Jag2floxneo targeting vector, and the targeted Jag2floxneo allele. Exons are indicated by boxes with coding sequences designated by ... The Jag2floxneo targeting construct was electroporated into R1 ES cells (Nagy et al., 1993), and three correctly targeted clones (Fig. 1b) were injected into C57BL/6J blastocysts. Chimeras were mated to C57BL/6J female mice, and germline transmission was obtained from one clone. Jag2floxneo/+ heterozygous mice were mated to a deleter line expressing the Flpe recombinase (Farley et al., 2000) to excise the PGKneo cassette and generate the Jag2flox allele. Mice heterozygous for the Jag2del2 null allele were generated by mating Jag2flox/+ heterozygous mice to Meox2-Cre mice which, in addition to other tissues, express Cre recombinase in the germline (Tallquist and Soriano, 2000). As with the Jag2del1 allele, heterozygous Jag2del2/+ mice were viable, fertile and displayed no obvious phenotypic abnormalities. Cre-mediated excision of the Jag2flox allele to generate the Jag2del2 allele deletes predicted Jag2 promoter sequences, the ATG translation start site and exons 1 and 2, which we predict would create a Jag2 null allele. In order to assess the functionality of the Jag2del2 allele, embryos and neonatal mice homozygous for the Jag2del2 allele were obtained by intercrossing Jag2del2/+ heterozygous mice. We assessed Jag2 RNA expression in Jag2del2/Jag2del2 and Jag2del2/+ littermate control embryos at E10.5 by whole mount in situ hybridization. Jag2 RNA was not expressed at levels detectable by in situ hybridization in Jag2del2/Jag2del2 homozygous mutant embryos (Fig. 2). We also assessed Jag2 RNA expression in homozygous Jag2del2/Jag2del2 mice by quantitative RT-PCR. Using a primer set that spanned exons 3 and 4, Jag2del2/Jag2del2 homozygous mutants expressed 11.5 ± 1.3% of wildtype Jag2 transcript levels. Utilizing a primer set spanning exons 6–8 of the Jag2 gene, Jag2del2/Jag2del2 mutant embryos expressed 21.8± 3.6% of wildtype Jag2 transcript levels. However, these levels of the Jag2del2 mutant transcript do not appear to have any functional consequences, since Jag2del2/Jag2del2 homozygous mutant mice exhibited a phenotype identical to that exhibited by Jag2del1/Jag2del1 null mutant mice (Fig. 3). Jag2del2/Jag2del2 neonates died the first day of birth, exhibiting syndactyly of the fore- and hindlimbs and palate-tongue fusions that caused palatal clefting (Fig. 3). These data demonstrate that the Jag2del2 allele is a Jag2 null allele, functionally equivalent to the Jag2del1 allele, and that the extent of Cre-mediated deletion of the Jag2flox allele can be assessed by in situ hybridization. The neonatal lethality of Jag2 null mice has precluded analysis of Jag2 function in postnatal and adult mice. Utilization of the Jag2flox conditional null allele will enable the investigation of Jag2 function in a tissue-specific manner throughout the mouse life span. FIG. 2 Absence of Jag2 RNA expression in Jag2del2/Jag2del2 embryos. (a,b) Whole mount in situ hybridization for Jag2 transcripts in Jag2del2/+ control littermate (a,c,e,g) and Jag2del2/Jag2del2 (b,d,f,h) embryos at E10.5. Jag2 RNA was not detectable by in situ ... FIG. 3 Phenotype of Jag2del2 homozygous mutant embryos. (a–c) Cleft palate in Jag2del2/Jag2del2 homozygotes. Coronal sections of E16.5 embryos show that the Jag2del2/Jag2del2 homozygote has cleft palate and fused tongue. (c) Fusion of the tongue with ...
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genetic background modifies inner ear and eye phenotypes of JAG1 heterozygous mice
Genetics, 2007Co-Authors: Amy E Kiernan, Renhua Li, Norman L Hawes, Gary A Churchill, Thomas GridleyAbstract:Mice heterozygous for missense mutations of the Notch ligand Jagged1 (JAG1) exhibit head-shaking behavior indicative of an inner ear vestibular defect. In contrast, mice heterozygous for a targeted deletion of the JAG1 gene (JAG1del1) do not demonstrate obvious head-shaking behavior. To determine whether the differences in inner ear phenotypes were due to the types of JAG1 mutations or to differences in genetic background, we crossed JAG1del1 heterozygous mice onto the same genetic background as the missense mutants. This analysis revealed that variation of the JAG1 mutant inner ear phenotype is caused by genetic background differences and is not due to the type of JAG1 mutation. Genome scans of N2 backcross mice identified a significant modifier locus on chromosome 7, as well as a suggestive locus on chromosome 14. We also analyzed modifiers of an eye defect in JAG1del1 heterozygous mice from this same cross.
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Bile duct proliferation in liver‐specific JAG1 conditional knockout mice: Effects of gene dosage
Hepatology, 2007Co-Authors: Kathleen M Loomes, Thomas Gridley, Pierre Russo, Matt E Ryan, Anthony Nelson, Lara A Underkoffler, Curtis L Glover, Hong Fu, Klaus H. Kaestner, Rebecca J. OakeyAbstract:The Notch signaling pathway is involved in determination of cell fate and control of cell proliferation in multiple organ systems. JAG1 encodes a ligand in the Notch pathway and has been identified as the disease-causing gene for the developmental disorder Alagille syndrome. Evidence from the study of human disease and mouse models has implicated JAG1 as having an important role in the development of bile ducts. We have derived a conditional knockout allele (JAG1loxP) to study the role of JAG1 and Notch signaling in liver and bile duct development. We crossed JAG1loxP mice with a transgenic line carrying Cre recombinase under the control of the albumin promoter and α-fetoprotein enhancer to ablate JAG1 in hepatoblasts. The liver-specific JAG1 conditional knockout mice showed normal bile duct development. To further decrease Notch pathway function, we crossed the JAG1 conditional knockout mice with mice carrying the hypomorphic Notch2 allele, and bile duct anatomy remained normal. When JAG1 conditional mice were crossed with mice carrying the JAG1 null allele, the adult progeny exhibited striking bile duct proliferation. Conclusion: These results indicate that Notch signaling in the liver is sensitive to JAG1 gene dosage and suggest a role for the Notch pathway in postnatal growth and morphogenesis of bile ducts. (HEPATOLOGY 2007.)
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the notch ligand JAG1 is required for sensory progenitor development in the mammalian inner ear
PLOS Genetics, 2005Co-Authors: Amy E Kiernan, Jingxia Xu, Thomas GridleyAbstract:In mammals, six separate sensory regions in the inner ear are essential for hearing and balance function. Each sensory region is made up of hair cells, which are the sensory cells, and their associated supporting cells, both arising from a common progenitor. Little is known about the molecular mechanisms that govern the development of these sensory organs. Notch signaling plays a pivotal role in the differentiation of hair cells and supporting cells by mediating lateral inhibition via the ligands Delta-like 1 and Jagged (JAG) 2. However, another Notch ligand, JAG1, is expressed early in the sensory patches prior to cell differentiation, indicating that there may be an earlier role for Notch signaling in sensory development in the ear. Here, using conditional gene targeting, we show that the JAG1 gene is required for the normal development of all six sensory organs within the inner ear. Cristae are completely lacking in JAG1-conditional knockout (cko) mutant inner ears, whereas the cochlea and utricle show partial sensory development. The saccular macula is present but malformed. Using SOX2 and p27kip1 as molecular markers of the prosensory domain, we show that JAG1 is initially expressed in all the prosensory regions of the ear, but becomes down-regulated in the nascent organ of Corti by embryonic day 14.5, when the cells exit the cell cycle and differentiate. We also show that both SOX2 and p27kip1 are down-regulated in JAG1-cko inner ears. Taken together, these data demonstrate that JAG1 is expressed early in the prosensory domains of both the cochlear and vestibular regions, and is required to maintain the normal expression levels of both SOX2 and p27kip1. These data demonstrate that JAG1-mediated Notch signaling is essential during early development for establishing the prosensory regions of the inner ear.
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a mouse model of alagille syndrome notch2 as a genetic modifier of JAG1 haploinsufficiency
Development, 2002Co-Authors: Brent Mccright, Julie Lozier, Thomas GridleyAbstract:Alagille syndrome is a human autosomal dominant developmental disorder characterized by liver, heart, eye, skeletal, craniofacial and kidney abnormalities. Alagille syndrome is caused by mutations in the Jagged 1 ( JAG1 ) gene, which encodes a ligand for Notch family receptors. The majority of JAG1 mutations seen in Alagille syndrome patients are null alleles, suggesting JAG1 haploinsufficiency as a primary cause of this disorder. Mice homozygous for a JAG1 null mutation die during embryogenesis and JAG1/+ heterozygous mice exhibit eye defects but do not exhibit other phenotypes characteristic of Alagille syndrome patients ( Xue, Y., Gao, X., Lindsell, C. E., Norton, C. R., Chang, B., Hicks, C., Gendron-Maguire, M., Rand, E. B., Weinmaster, G. and Gridley, T. (1999) Hum. Mol. Genet . , -730 ). Here we report that mice doubly heterozygous for the JAG1 null allele and a Notch2 hypomorphic allele exhibit developmental abnormalities characteristic of Alagille syndrome. Double heterozygous mice exhibit jaundice, growth retardation, impaired differentiation of intrahepatic bile ducts and defects in heart, eye and kidney development. The defects in bile duct epithelial cell differentiation and morphogenesis in the double heterozygous mice are similar to defects in epithelial morphogenesis of Notch pathway mutants in Drosophila , suggesting that a role for the Notch signaling pathway in regulating epithelial morphogenesis has been conserved between insects and mammals. This work also demonstrates that the Notch2 and JAG1 mutations interact to create a more representative mouse model of Alagille syndrome and provides a possible explanation of the variable phenotypic expression observed in Alagille syndrome patients.
Anthony Nelson - One of the best experts on this subject based on the ideXlab platform.
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microarray data reveal relationship between JAG1 and ddr1 in mouse liver
PLOS ONE, 2013Co-Authors: Lara A Underkoffler, Anthony Nelson, Erikka Carr, Matthew J Ryan, Reiner Schultz, Kathleen M LoomesAbstract:Alagille syndrome is an autosomal dominant disorder involving bile duct paucity and cholestasis in addition to cardiac, skeletal, ophthalmologic, renal and vascular manifestations. Mutations in JAG1, encoding a ligand in the Notch signaling pathway, are found in 95% of patients meeting clinical criteria for Alagille syndrome. In order to define the role of JAG1 in the bile duct developmental abnormalities seen in ALGS, we previously created a JAG1 conditional knockout mouse model. Mice heterozygous for the JAG1 conditional and null alleles demonstrate abnormalities in postnatal bile duct growth and remodeling, with portal expansion and increased numbers of malformed bile ducts. In this study we report the results of microarray analysis and identify genes and pathways differentially expressed in the JAG1 conditional/null livers as compared with littermate controls. In the initial microarray analysis, we found that many of the genes up-regulated in the JAG1 conditional/null mutant livers were related to extracellular matrix (ECM) interactions, cell adhesion and cell migration. One of the most highly up-regulated genes was Ddr1, encoding a receptor tyrosine kinase (RTK) belonging to a large RTK family. We have found extensive co-localization of JAG1 and Ddr1 in bile ducts and blood vessels in postnatal liver. In addition, co-immunoprecipitation data provide evidence for a novel protein interaction between JAG1 and Ddr1. Further studies will be required to define the nature of this interaction and its functional consequences, which may have significant implications for bile duct remodeling and repair of liver injury.
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bile duct proliferation in JAG1 fringe heterozygous mice identifies candidate modifiers of the alagille syndrome hepatic phenotype
Hepatology, 2008Co-Authors: Anthony Nelson, Lara A Underkoffler, Matthew J Ryan, Christina Bales, Dorian M Gonzalez, Michelle Segalov, Jeanne Wilsonrawls, Susan E ColeAbstract:Alagille syndrome (AGS) is a heterogeneous developmental disorder associated with bile duct paucity and various organ anomalies. The syndrome is caused by mutations in JAG1, which encodes a ligand in the Notch signaling pathway, in the majority of cases and mutations in the NOTCH2 receptor gene in less than 1% of patients. Although a wide array of JAG1 mutations have been identified in the AGS population, these mutational variants have not accounted for the wide phenotypic variability observed in patients with this syndrome. The Fringe genes encode glycosyltransferases, which modify Notch and alter ligand-receptor affinity. In this study, we analyzed double heterozygous mouse models to examine the Fringe genes as potential modifiers of the Notch-mediated hepatic phenotype observed in AGS. We generated mice that were haploinsufficient for both JAG1 and one of three paralogous Fringe genes: Lunatic (Lfng), Radical (Rfng), and Manic (Mfng). Adult JAG1+/−Lfng+/− and JAG1+/−Rfng+/− mouse livers exhibited widespread bile duct proliferation beginning at 5 weeks of age and persisting up to 1 year. The JAG1+/−Mfng+/− livers showed a subtle, yet significant increase in bile duct numbers and bile duct to portal tract ratios. These abnormalities were not observed in the newborn period. Despite the portal tract expansion by bile ducts, fibrosis was not increased and epithelial to mesenchymal transition was not shown in the affected portal tracts. Conclusion: Mice heterozygous for mutations in JAG1 and the Fringe genes display striking bile duct proliferation, which is not apparent at birth. These findings suggest that the Fringe genes may regulate postnatal bile duct growth and remodeling, and serve as candidate modifiers of the hepatic phenotype in AGS. (HEPATOLOGY 2008;48:1989–1997.)
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Bile duct proliferation in JAG1/fringe heterozygous mice identifies candidate modifiers of the Alagille syndrome hepatic phenotype.
Hepatology, 2008Co-Authors: Matthew J Ryan, Anthony Nelson, Lara A Underkoffler, Christina Bales, Dorian M Gonzalez, Michelle Segalov, Susan E Cole, Jeanne Wilson-rawls, Jennifer L. Moran, Pierre RussoAbstract:Alagille syndrome (AGS) is a heterogeneous developmental disorder associated with bile duct paucity and various organ anomalies. The syndrome is caused by mutations in JAG1, which encodes a ligand in the Notch signaling pathway, in the majority of cases and mutations in the NOTCH2 receptor gene in less than 1% of patients. Although a wide array of JAG1 mutations have been identified in the AGS population, these mutational variants have not accounted for the wide phenotypic variability observed in patients with this syndrome. The Fringe genes encode glycosyltransferases, which modify Notch and alter ligand-receptor affinity. In this study, we analyzed double heterozygous mouse models to examine the Fringe genes as potential modifiers of the Notch-mediated hepatic phenotype observed in AGS. We generated mice that were haploinsufficient for both JAG1 and one of three paralogous Fringe genes: Lunatic (Lfng), Radical (Rfng), and Manic (Mfng). Adult JAG1+/−Lfng+/− and JAG1+/−Rfng+/− mouse livers exhibited widespread bile duct proliferation beginning at 5 weeks of age and persisting up to 1 year. The JAG1+/−Mfng+/− livers showed a subtle, yet significant increase in bile duct numbers and bile duct to portal tract ratios. These abnormalities were not observed in the newborn period. Despite the portal tract expansion by bile ducts, fibrosis was not increased and epithelial to mesenchymal transition was not shown in the affected portal tracts. Conclusion: Mice heterozygous for mutations in JAG1 and the Fringe genes display striking bile duct proliferation, which is not apparent at birth. These findings suggest that the Fringe genes may regulate postnatal bile duct growth and remodeling, and serve as candidate modifiers of the hepatic phenotype in AGS. (HEPATOLOGY 2008;48:1989–1997.)
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bile duct proliferation in liver specific JAG1 conditional knockout mice effects of gene dosage
Hepatology, 2007Co-Authors: Kathleen M Loomes, Pierre Russo, Matt E Ryan, Anthony Nelson, Lara A Underkoffler, Curtis L Glover, Hong FuAbstract:The Notch signaling pathway is involved in determination of cell fate and control of cell proliferation in multiple organ systems. JAG1 encodes a ligand in the Notch pathway and has been identified as the disease-causing gene for the developmental disorder Alagille syndrome. Evidence from the study of human disease and mouse models has implicated JAG1 as having an important role in the development of bile ducts. We have derived a conditional knockout allele (JAG1loxP) to study the role of JAG1 and Notch signaling in liver and bile duct development. We crossed JAG1loxP mice with a transgenic line carrying Cre recombinase under the control of the albumin promoter and α-fetoprotein enhancer to ablate JAG1 in hepatoblasts. The liver-specific JAG1 conditional knockout mice showed normal bile duct development. To further decrease Notch pathway function, we crossed the JAG1 conditional knockout mice with mice carrying the hypomorphic Notch2 allele, and bile duct anatomy remained normal. When JAG1 conditional mice were crossed with mice carrying the JAG1 null allele, the adult progeny exhibited striking bile duct proliferation. Conclusion: These results indicate that Notch signaling in the liver is sensitive to JAG1 gene dosage and suggest a role for the Notch pathway in postnatal growth and morphogenesis of bile ducts. (HEPATOLOGY 2007.)
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Bile duct proliferation in liver‐specific JAG1 conditional knockout mice: Effects of gene dosage
Hepatology, 2007Co-Authors: Kathleen M Loomes, Thomas Gridley, Pierre Russo, Matt E Ryan, Anthony Nelson, Lara A Underkoffler, Curtis L Glover, Hong Fu, Klaus H. Kaestner, Rebecca J. OakeyAbstract:The Notch signaling pathway is involved in determination of cell fate and control of cell proliferation in multiple organ systems. JAG1 encodes a ligand in the Notch pathway and has been identified as the disease-causing gene for the developmental disorder Alagille syndrome. Evidence from the study of human disease and mouse models has implicated JAG1 as having an important role in the development of bile ducts. We have derived a conditional knockout allele (JAG1loxP) to study the role of JAG1 and Notch signaling in liver and bile duct development. We crossed JAG1loxP mice with a transgenic line carrying Cre recombinase under the control of the albumin promoter and α-fetoprotein enhancer to ablate JAG1 in hepatoblasts. The liver-specific JAG1 conditional knockout mice showed normal bile duct development. To further decrease Notch pathway function, we crossed the JAG1 conditional knockout mice with mice carrying the hypomorphic Notch2 allele, and bile duct anatomy remained normal. When JAG1 conditional mice were crossed with mice carrying the JAG1 null allele, the adult progeny exhibited striking bile duct proliferation. Conclusion: These results indicate that Notch signaling in the liver is sensitive to JAG1 gene dosage and suggest a role for the Notch pathway in postnatal growth and morphogenesis of bile ducts. (HEPATOLOGY 2007.)