Wnt signaling through the Frizzled (FZD) family of serpentine receptors is essential for embryoge... more Wnt signaling through the Frizzled (FZD) family of serpentine receptors is essential for embryogenesis and homeostasis, and stringent control of the FZD protein level is critical for stem cell regulation. Through CRISPR/Cas9 genome-wide screening in human cells, we identified TMEM79/MATTRIN, an orphan multi-span transmembrane protein, as a specific inhibitor of Wnt/FZD signaling. TMEM79 interacts with FZD during biogenesis and promotes FZD degradation independent of ZNRF3/RNF43 ubiquitin ligases (R-spondin receptors). TMEM79 interacts with ubiquitin-specific protease 8 (USP8), whose activating mutations underlie human tumorigenesis. TMEM79 specifically inhibits USP8 deubiquitination of FZD, thereby governing USP8 substrate specificity and promoting FZD degradation. Tmem79 and Usp8 genes have a pre-bilaterian origin, and Tmem79 inhibition of Usp8 and Wnt signaling is required for anterior neural development and gastrulation in Xenopus embryos. TMEM79 is a predisposition gene for Atop...
Glioblastoma is an extremely aggressive and deadly brain tumor known for its striking cellular he... more Glioblastoma is an extremely aggressive and deadly brain tumor known for its striking cellular heterogeneity and capability to communicate with microenvironment components, such as microglia. Microglia-glioblastoma interaction contributes to an increase in tumor invasiveness, and Wnt signaling pathway is one of the main cascades related to tumor progression through changes in cell migration and invasion. However, very little is known about the role of canonical Wnt signaling during microglia-glioblastoma crosstalk. Here, we show for the first time that Wnt3a is one of the factors that regulate interactions between microglia and glioblastoma cells. Wnt3a activates the Wnt/β-catenin signaling of both glioblastoma and microglial cells. Glioblastoma-conditioned medium not only induces nuclear translocation of microglial β-catenin but also increases microglia viability and proliferation as well as Wnt3a, cyclin-D1, and c-myc expression. Moreover, glioblastoma-derived Wnt3a increases micr...
Understanding embryogenesis currently relies largely on the control of gene expression via severa... more Understanding embryogenesis currently relies largely on the control of gene expression via several signaling pathways. Many of the embryonic signaling pathways guiding embryological events are implicated in diseases that lack effective cure or treatment. Because of the large number and size of the eggs, the rapid development of the embryos and the fact they are amenable to pharmacological, surgical and genetic techniques, Xenopus laevis has been successfully used in searching for compounds that target embryonic signaling pathways. Areas covered: Here, the authors address the use of amphibian eggs/embryos in successful chemical screenings; egg extracts as well as embryo phenotypes have been assayed to reveal drug toxicology effects and novel compounds acting in the Wnt/β-catenin signaling pathway. They do not discuss the use of Xenopus oocyte two-electrode voltage clamps or genome editing tools as approaches for drug discovery because they have been discussed elsewhere. Expert opinion: While high-throughput screening is commonly performed in egg extracts, the embryo axes perturbation system is more suited to the refinement and/or the validation of drug discovery targeting embryonic signaling (particularly the Wnt/β-catenin pathway). In addition, Xenopus has also been used in FETAX (frog embryo teratogenesis assay: Xenopus) to address chemical toxic/teratogenic effects. However, further studies are necessary.
Myc proto-oncogenes regulate diverse cellular processes during development, but their roles durin... more Myc proto-oncogenes regulate diverse cellular processes during development, but their roles during morphogenesis of specific tissues are not fully understood. We found that c-myc regulates cell proliferation in mouse lens development and previous genome-wide studies suggested functional roles for N-myc in developing lens. Here, we examined the role of N-myc in mouse lens development. Genetic inactivation of N-myc in the surface ectoderm or lens vesicle impaired eye and lens growth, while "late" inactivation in lens fibers had no effect. Unexpectedly, defective growth of N-myc-deficient lenses was not associated with alterations in lens progenitor cell proliferation or survival. Notably, N-myc-deficient lens exhibited a delay in degradation of DNA in terminally differentiating lens fiber cells. RNA-sequencing analysis of N-myc-deficient lenses identified a cohort of down-regulated genes associated with fiber cell differentiation that included DNaseIIβ. Further, an integrate...
Direct development in amphibians is characterized by the loss of aquatic breeding. The anuran Ade... more Direct development in amphibians is characterized by the loss of aquatic breeding. The anuran Adelophryne maranguapensis is one example of a species with direct development, and it is endemic to the state of Ceará, Brazil. Detailed morphological features of A. maranguapensis embryos and the stages of sequential development have not been described before. Here, we analyzed all available genetic sequence tags in A. maranguapensis (tyr exon 1, pomc and rag1) and compared them with sequences from other species of Adelophryne frogs. We describe the A. maranguapensis reproductive tract and embryonic body development, with a focus on the limbs, tail, ciliated cells of the skin, and the egg tooth, which were analyzed using scanning electron microscopy. Histological analyses revealed ovaries containing oocytes surrounded by follicular cells, displaying large nuclei with nucleoli inside. Early in development, the body is unpigmented, and the neural tube forms dorsally to the yolk vesicle, typ...
The International Journal of Developmental Biology, Feb 1, 2001
We review how studies on the first Spemann-Mangold organizer marker, the homeobox gene goosecoid,... more We review how studies on the first Spemann-Mangold organizer marker, the homeobox gene goosecoid, led to the discovery of secreted factors that pattern the vertebrate embryo. Microinjection of goosecoid mRNA formed secondary axes and recruited neighboring cells. These non-cell autonomous effects are mediated in part by the expression of secreted factors such as chordin, cerberus and Frzb-1. Unexpectedly, many of the molecules secreted by the Spemann-Mangold organizer turned out to be antagonists that bind growth factors in the extracellular space and prevent them from binding to their receptors. The case of chordin is reviewed in detail, for this molecule has provided biochemical insights into how patterning by Spemann's organizer can be regulated by diffusion and proteolytic control. The study of the BMP-binding repeats of Chordin, which are present in many extracellular proteins, may provide a new paradigm for how cell-cell signaling is regulated in the extracellular space not only in embryos, but also in adult tissues.
Brazilian journal of medical and biological research = Revista brasileira de pesquisas médicas e biológicas / Sociedade Brasileira de Biofísica ... [et al.], 2011
Connective tissue growth factor (CCN2/CTGF) is a matricellular-secreted protein involved in extra... more Connective tissue growth factor (CCN2/CTGF) is a matricellular-secreted protein involved in extracellular matrix remodeling. The P19 cell line is an embryonic carcinoma line widely used as a cellular model for differentiation and migration studies. In the present study, we employed an exogenous source of CCN2 and small interference RNA to address the role of CCN2 in the P19 cell aggregation phenomenon. Our data showed that increasing CCN2 protein concentrations from 0.1 to 20 nM decreased the number of cell clusters and dramatically increased cluster size without changing proliferation or cell survival, suggesting that CCN2 induced aggregation. In addition, CCN2 specific silencing inhibited typical P19 cell aggregation, which could be partially rescued by 20 nM CCN2. The present study demonstrates that CCN2 is a key molecule for cell aggregation of embryonic P19 cells.
The International journal of developmental biology, 2001
We review how studies on the first Spemann-Mangold organizer marker, the homeobox gene goosecoid,... more We review how studies on the first Spemann-Mangold organizer marker, the homeobox gene goosecoid, led to the discovery of secreted factors that pattern the vertebrate embryo. Microinjection of goosecoid mRNA formed secondary axes and recruited neighboring cells. These non-cell autonomous effects are mediated in part by the expression of secreted factors such as chordin, cerberus and Frzb-1. Unexpectedly, many of the molecules secreted by the Spemann-Mangold organizer turned out to be antagonists that bind growth factors in the extracellular space and prevent them from binding to their receptors. The case of chordin is reviewed in detail, for this molecule has provided biochemical insights into how patterning by Spemann's organizer can be regulated by diffusion and proteolytic control. The study of the BMP-binding repeats of Chordin, which are present in many extracellular proteins, may provide a new paradigm for how cell-cell signaling is regulated in the extracellular space not...
The kidney is a homeostatic organ required for waste excretion and reabsorption of water, salts a... more The kidney is a homeostatic organ required for waste excretion and reabsorption of water, salts and other macromolecules. To this end, a complex series of developmental steps ensures the formation of a correctly patterned and properly proportioned organ. While previous studies have mainly focused on the individual signaling pathways, the formation of higher order receptor complexes in lipid rafts is an equally important aspect. These membrane platforms are characterized by differences in local lipid and protein compositions. Indeed, the cells in the Xenopus pronephric kidney were positive for the lipid raft markers ganglioside GM1 and Caveolin-1. To specifically interfere with lipid raft function in vivo, we focused on the Sterol Carrier Protein 2 (scp2), a multifunctional protein that is an important player in remodeling lipid raft composition. In Xenopus, scp2 mRNA was strongly expressed in differentiated epithelial structures of the pronephric kidney. Knockdown of scp2 did not interfere with the patterning of the kidney along its proximo-distal axis, but dramatically decreased the size of the kidney, in particular the proximal tubules. This phenotype was accompanied by a reduction of lipid rafts, but was independent of the peroxisomal or transcriptional activities of scp2. Finally, disrupting lipid micro-domains by inhibiting cholesterol synthesis using Mevinolin phenocopied the defects seen in scp2 morphants. Together these data underscore the importance for localized signaling platforms in the proper formation of the Xenopus kidney.
Wnt signaling through the Frizzled (FZD) family of serpentine receptors is essential for embryoge... more Wnt signaling through the Frizzled (FZD) family of serpentine receptors is essential for embryogenesis and homeostasis, and stringent control of the FZD protein level is critical for stem cell regulation. Through CRISPR/Cas9 genome-wide screening in human cells, we identified TMEM79/MATTRIN, an orphan multi-span transmembrane protein, as a specific inhibitor of Wnt/FZD signaling. TMEM79 interacts with FZD during biogenesis and promotes FZD degradation independent of ZNRF3/RNF43 ubiquitin ligases (R-spondin receptors). TMEM79 interacts with ubiquitin-specific protease 8 (USP8), whose activating mutations underlie human tumorigenesis. TMEM79 specifically inhibits USP8 deubiquitination of FZD, thereby governing USP8 substrate specificity and promoting FZD degradation. Tmem79 and Usp8 genes have a pre-bilaterian origin, and Tmem79 inhibition of Usp8 and Wnt signaling is required for anterior neural development and gastrulation in Xenopus embryos. TMEM79 is a predisposition gene for Atop...
Glioblastoma is an extremely aggressive and deadly brain tumor known for its striking cellular he... more Glioblastoma is an extremely aggressive and deadly brain tumor known for its striking cellular heterogeneity and capability to communicate with microenvironment components, such as microglia. Microglia-glioblastoma interaction contributes to an increase in tumor invasiveness, and Wnt signaling pathway is one of the main cascades related to tumor progression through changes in cell migration and invasion. However, very little is known about the role of canonical Wnt signaling during microglia-glioblastoma crosstalk. Here, we show for the first time that Wnt3a is one of the factors that regulate interactions between microglia and glioblastoma cells. Wnt3a activates the Wnt/β-catenin signaling of both glioblastoma and microglial cells. Glioblastoma-conditioned medium not only induces nuclear translocation of microglial β-catenin but also increases microglia viability and proliferation as well as Wnt3a, cyclin-D1, and c-myc expression. Moreover, glioblastoma-derived Wnt3a increases micr...
Understanding embryogenesis currently relies largely on the control of gene expression via severa... more Understanding embryogenesis currently relies largely on the control of gene expression via several signaling pathways. Many of the embryonic signaling pathways guiding embryological events are implicated in diseases that lack effective cure or treatment. Because of the large number and size of the eggs, the rapid development of the embryos and the fact they are amenable to pharmacological, surgical and genetic techniques, Xenopus laevis has been successfully used in searching for compounds that target embryonic signaling pathways. Areas covered: Here, the authors address the use of amphibian eggs/embryos in successful chemical screenings; egg extracts as well as embryo phenotypes have been assayed to reveal drug toxicology effects and novel compounds acting in the Wnt/β-catenin signaling pathway. They do not discuss the use of Xenopus oocyte two-electrode voltage clamps or genome editing tools as approaches for drug discovery because they have been discussed elsewhere. Expert opinion: While high-throughput screening is commonly performed in egg extracts, the embryo axes perturbation system is more suited to the refinement and/or the validation of drug discovery targeting embryonic signaling (particularly the Wnt/β-catenin pathway). In addition, Xenopus has also been used in FETAX (frog embryo teratogenesis assay: Xenopus) to address chemical toxic/teratogenic effects. However, further studies are necessary.
Myc proto-oncogenes regulate diverse cellular processes during development, but their roles durin... more Myc proto-oncogenes regulate diverse cellular processes during development, but their roles during morphogenesis of specific tissues are not fully understood. We found that c-myc regulates cell proliferation in mouse lens development and previous genome-wide studies suggested functional roles for N-myc in developing lens. Here, we examined the role of N-myc in mouse lens development. Genetic inactivation of N-myc in the surface ectoderm or lens vesicle impaired eye and lens growth, while "late" inactivation in lens fibers had no effect. Unexpectedly, defective growth of N-myc-deficient lenses was not associated with alterations in lens progenitor cell proliferation or survival. Notably, N-myc-deficient lens exhibited a delay in degradation of DNA in terminally differentiating lens fiber cells. RNA-sequencing analysis of N-myc-deficient lenses identified a cohort of down-regulated genes associated with fiber cell differentiation that included DNaseIIβ. Further, an integrate...
Direct development in amphibians is characterized by the loss of aquatic breeding. The anuran Ade... more Direct development in amphibians is characterized by the loss of aquatic breeding. The anuran Adelophryne maranguapensis is one example of a species with direct development, and it is endemic to the state of Ceará, Brazil. Detailed morphological features of A. maranguapensis embryos and the stages of sequential development have not been described before. Here, we analyzed all available genetic sequence tags in A. maranguapensis (tyr exon 1, pomc and rag1) and compared them with sequences from other species of Adelophryne frogs. We describe the A. maranguapensis reproductive tract and embryonic body development, with a focus on the limbs, tail, ciliated cells of the skin, and the egg tooth, which were analyzed using scanning electron microscopy. Histological analyses revealed ovaries containing oocytes surrounded by follicular cells, displaying large nuclei with nucleoli inside. Early in development, the body is unpigmented, and the neural tube forms dorsally to the yolk vesicle, typ...
The International Journal of Developmental Biology, Feb 1, 2001
We review how studies on the first Spemann-Mangold organizer marker, the homeobox gene goosecoid,... more We review how studies on the first Spemann-Mangold organizer marker, the homeobox gene goosecoid, led to the discovery of secreted factors that pattern the vertebrate embryo. Microinjection of goosecoid mRNA formed secondary axes and recruited neighboring cells. These non-cell autonomous effects are mediated in part by the expression of secreted factors such as chordin, cerberus and Frzb-1. Unexpectedly, many of the molecules secreted by the Spemann-Mangold organizer turned out to be antagonists that bind growth factors in the extracellular space and prevent them from binding to their receptors. The case of chordin is reviewed in detail, for this molecule has provided biochemical insights into how patterning by Spemann's organizer can be regulated by diffusion and proteolytic control. The study of the BMP-binding repeats of Chordin, which are present in many extracellular proteins, may provide a new paradigm for how cell-cell signaling is regulated in the extracellular space not only in embryos, but also in adult tissues.
Brazilian journal of medical and biological research = Revista brasileira de pesquisas médicas e biológicas / Sociedade Brasileira de Biofísica ... [et al.], 2011
Connective tissue growth factor (CCN2/CTGF) is a matricellular-secreted protein involved in extra... more Connective tissue growth factor (CCN2/CTGF) is a matricellular-secreted protein involved in extracellular matrix remodeling. The P19 cell line is an embryonic carcinoma line widely used as a cellular model for differentiation and migration studies. In the present study, we employed an exogenous source of CCN2 and small interference RNA to address the role of CCN2 in the P19 cell aggregation phenomenon. Our data showed that increasing CCN2 protein concentrations from 0.1 to 20 nM decreased the number of cell clusters and dramatically increased cluster size without changing proliferation or cell survival, suggesting that CCN2 induced aggregation. In addition, CCN2 specific silencing inhibited typical P19 cell aggregation, which could be partially rescued by 20 nM CCN2. The present study demonstrates that CCN2 is a key molecule for cell aggregation of embryonic P19 cells.
The International journal of developmental biology, 2001
We review how studies on the first Spemann-Mangold organizer marker, the homeobox gene goosecoid,... more We review how studies on the first Spemann-Mangold organizer marker, the homeobox gene goosecoid, led to the discovery of secreted factors that pattern the vertebrate embryo. Microinjection of goosecoid mRNA formed secondary axes and recruited neighboring cells. These non-cell autonomous effects are mediated in part by the expression of secreted factors such as chordin, cerberus and Frzb-1. Unexpectedly, many of the molecules secreted by the Spemann-Mangold organizer turned out to be antagonists that bind growth factors in the extracellular space and prevent them from binding to their receptors. The case of chordin is reviewed in detail, for this molecule has provided biochemical insights into how patterning by Spemann's organizer can be regulated by diffusion and proteolytic control. The study of the BMP-binding repeats of Chordin, which are present in many extracellular proteins, may provide a new paradigm for how cell-cell signaling is regulated in the extracellular space not...
The kidney is a homeostatic organ required for waste excretion and reabsorption of water, salts a... more The kidney is a homeostatic organ required for waste excretion and reabsorption of water, salts and other macromolecules. To this end, a complex series of developmental steps ensures the formation of a correctly patterned and properly proportioned organ. While previous studies have mainly focused on the individual signaling pathways, the formation of higher order receptor complexes in lipid rafts is an equally important aspect. These membrane platforms are characterized by differences in local lipid and protein compositions. Indeed, the cells in the Xenopus pronephric kidney were positive for the lipid raft markers ganglioside GM1 and Caveolin-1. To specifically interfere with lipid raft function in vivo, we focused on the Sterol Carrier Protein 2 (scp2), a multifunctional protein that is an important player in remodeling lipid raft composition. In Xenopus, scp2 mRNA was strongly expressed in differentiated epithelial structures of the pronephric kidney. Knockdown of scp2 did not interfere with the patterning of the kidney along its proximo-distal axis, but dramatically decreased the size of the kidney, in particular the proximal tubules. This phenotype was accompanied by a reduction of lipid rafts, but was independent of the peroxisomal or transcriptional activities of scp2. Finally, disrupting lipid micro-domains by inhibiting cholesterol synthesis using Mevinolin phenocopied the defects seen in scp2 morphants. Together these data underscore the importance for localized signaling platforms in the proper formation of the Xenopus kidney.
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Papers by Jose Abreu