Anne Prieto | Indiana University (original) (raw)
Papers by Anne Prieto
International journal of molecular sciences, Mar 16, 2024
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
Journal of comparative neurology, 2000
Proceedings of the National Academy of Sciences of the United States of America, Aug 1, 1991
International Journal of Developmental Neuroscience, Nov 16, 2006
Introduction: Recently, it has been shown that H2O2, one of the most stable and long-lived ROS, a... more Introduction: Recently, it has been shown that H2O2, one of the most stable and long-lived ROS, are produced and released during neuromuscular activity and play important roles in modulating synaptic activity. Less is known, however, about the underlying mechanism of H2O2 on synaptic facilitation. Methods: Whole-cell patch clamp recording in one-day-old Xenopus nerve–muscle coculture were used. Results: Exclusion of calcium from culture medium or bath application of calcium channel inhibitor either cadmium, nifedipine, verapamil or -conotoxin significantly reduced H2O2 induced synaptic facilitation. Pretreatment of catalase and N-acetyl-l-cysteine effectively hampered H2O2 induced SSC frequency elevation. Preincubation of the cultures with either inhibitor of IP3 receptor XeC or ryanodine receptor TMB8 or depletion of ER Ca stores with thapsigargin failed to abolish the synaptic facilitation elicited by H2O2. Bath application of mitochondrial Na–Ca exchanger inhibitor FCCP or CGP37157 partialy obstructed the increase of SSC frequency in response to H2O2. Treating cells with protein kinase C inhibitors either H-9 or GF 109203X effectively occluded the increase of SSC frequency elicited by H2O2. Furthermore, application of either phosphatidyl inositol 3 kinase (PI3K) inhibitor wortmannin or phospholipase C (PLC ) inhibitor U73122, also abolished H2O2-induced facilitation of synaptic transmission. Discussion: Overall, results from our current study indicate that both calcium influx through Land N-type Ca channel and efflux from mitochondrial are responsible for H2O2-induced SSC frequency facilitation. This is done via PLC /PI3K, PKC activation and Na–Ca exchanger blockade.
Carolina Digital Repository (University of North Carolina at Chapel Hill), 2013
MerTK, a receptor tyrosine kinase (RTK) of the TYRO3/AXL/MerTK family, is expressed in myeloid li... more MerTK, a receptor tyrosine kinase (RTK) of the TYRO3/AXL/MerTK family, is expressed in myeloid lineage cells in which it acts to suppress proinflammatory cytokines following ingestion of apoptotic material. Using syngeneic mouse models of breast cancer, melanoma, and colon cancer, we found that tumors grew slowly and were poorly metastatic in MerTK-/mice. Transplantation of MerTK-/bone marrow, but not wild-type bone marrow, into lethally irradiated MMTV-PyVmT mice (a model of metastatic breast cancer) decreased tumor growth and altered cytokine production by tumor CD11b + cells. Although MerTK expression was not required for tumor infiltration by leukocytes, MerTK-/leukocytes exhibited lower tumor cell-induced expression of wound healing cytokines, e.g., IL-10 and growth arrest-specific 6 (GAS6), and enhanced expression of acute inflammatory cytokines, e.g., IL-12 and IL-6. Intratumoral CD8 + T lymphocyte numbers were higher and lymphocyte proliferation was increased in tumor-bearing MerTK-/mice compared with tumor-bearing wild-type mice. Antibody-mediated CD8 + T lymphocyte depletion restored tumor growth in MerTK-/mice. These data demonstrate that MerTK signaling in tumor-associated CD11b + leukocytes promotes tumor growth by dampening acute inflammatory cytokines while inducing wound healing cytokines. These results suggest that inhibition of MerTK in the tumor microenvironment may have clinical benefit, stimulating antitumor immune responses or enhancing immunotherapeutic strategies.
Molecular and Cellular Neuroscience, Jun 1, 2004
Acetylcholine receptor (AChR) genes are transcribed selectively in synaptic nuclei of skeletal mu... more Acetylcholine receptor (AChR) genes are transcribed selectively in synaptic nuclei of skeletal muscle fibers, leading to accumulation of the mRNAs encoding AChR subunits at synaptic sites. The signals that regulate synapse-specific transcription remain elusive, though Neuregulin-1 is considered a favored candidate. Here, we show that motor neurons and terminal Schwann cells express neuregulin-2, a neuregulin-1-related gene. In skeletal muscle, Neuregulin-2 protein is concentrated at synaptic sites, where it accumulates adjacent to terminal Schwann cells. Neuregulin-2 stimulates AChR transcription in cultured myotubes expressing ErbB4, as well as ErbB3 and ErbB2, but not in myotubes expressing only ErbB3 and ErbB2. Thus, Neuregulin-2 is a candidate for a signal that regulates synaptic differentiation.
The Journal of Neuroscience, May 14, 2008
The TAM family of receptor protein tyrosine kinases comprises three known members, namely Tyro3, ... more The TAM family of receptor protein tyrosine kinases comprises three known members, namely Tyro3, Axl, and Mer. These receptors are widely expressed in the nervous system, including by oligodendrocytes, the cell type responsible for myelinating the CNS. We examined the potential role of the TAM family and of their principle cognate ligand, Gas6 (growth arrest gene 6), in modulating the phenotype of the cuprizone model of demyelination. We found that the expression profiles of Axl, Mer, and Gas6 mRNA were increased in the corpus callosum in a temporal profile correlating with the increased migration and proliferation of microglia/macrophages in this model. In contrast, expression of Tyro3 decreased, correlating with the loss of oligodendrocytes. Gas6 both promoted in vitro survival of oligodendrocytes (39.3 Ϯ 3.1 vs 11.8 Ϯ 2.4%) and modulated markers of activation in purified cultures of microglia (tumor necrosis factor ␣ mRNA expression was reduced ϳ48%). In Gas6 Ϫ/Ϫ mice subjected to cuprizone-challenge, demyelination was greater than in control mice, within the rostral region of the corpus callosum, as assessed by luxol fast blue staining (myelination reduced by 36%) and by ultrastructural analysis. An increased loss of Gst-(glutathione S-transferase-)-positive oligodendrocytes was also identified throughout the corpus callosum of Gas6 Ϫ/Ϫ mice. Microglial marker expression (ionized calcium-binding adapter molecule 1) was increased in Gas6 Ϫ/Ϫ mice but was restricted to the rostral corpus callosum. Therefore, TAM receptor activation and regulation can independently influence both oligodendrocyte survival and the microglial response after CNS damage.
Experimental Eye Research, Oct 1, 2001
The function and viability of vertebrate photoreceptors requires the daily phagocytosis of photor... more The function and viability of vertebrate photoreceptors requires the daily phagocytosis of photoreceptor outer segments (OS) by the adjacent retinal pigment epithelium (RPE). We demonstrate here a critical role in this process for Gas6 and by implication one of its receptor protein tyrosine kinases (RTKs), Mertk (Mer). Gas6 speci®cally and selectively stimulates the phagocytosis of OS by normal cultured rat RPE cells. The magnitude of the response is dose-dependent and shows an absolute requirement for calcium. By contrast the Royal College of Surgeons (RCS) rat RPE cells, in which a mutation in the gene Mertk results in the expression of a truncated, non-functional receptor, does not respond to Gas6. These data strongly suggest that activation of Mertk by its ligand, Gas6, is the speci®c signaling pathway responsible for initiating the ingestion of shed OS. Moreover, photoreceptor degeneration in the RCS rat retina, which lacks Mertk, and in humans with a mutation in Mertk, strongly suggests that the Gas6/Mertk signaling pathway is essential for photoreceptor viability. We believe that this is the ®rst demonstration of a speci®c function for Gas6 in the eye.
Experimental Neurology, Jul 1, 1990
Evidence is accumulating that molecules involved in cell-cell and cell-substratum interactions ar... more Evidence is accumulating that molecules involved in cell-cell and cell-substratum interactions are important in the establishment and maintenance of borders between cell groups during development. In this report, we review evidence supporting this conclusion, particularly in regard to the role of adhesion molecules in the formation of cell collectives and in the modulation of cell and neurite movements.
Brain Research, 1999
Gas6 (growth arrest specific gene-6) is a ligand for members of the Axl subfamily of receptor pro... more Gas6 (growth arrest specific gene-6) is a ligand for members of the Axl subfamily of receptor protein-tyrosine kinases. One of these receptors, Tyro-3, is widely expressed in the central nervous system. We have used biochemical and histological techniques, including in situ hybridization, to determine the expression patterns of Gas6 mRNA and protein during development. Gas6 is widely expressed in the rat central nervous system (CNS) beginning at late embryonic stages and its levels remain high in the adult. Gas6 is detected as a single 85 kDa protein, which is encoded by a single 2.5 kb mRNA species. At embryonic day 14 it is detected in the heart, blood vessels, testes, choroid plexus, and in the ventral spinal cord. In the adult, Gas6 is expressed in the cerebral cortex, (predominantly in layer V), the piriform cortex, and the hippocampus (areas CA1, CA3 and the dentate gyrus). It is also expressed in thalamic and hypothalamic structures, the midbrain, and in a subset of motor and trigeminal nuclei. In the cerebellum, it is expressed in Purkinje neurons and deep cerebellar nuclei. Protein S, a protein related to Gas6, is only detected at low levels in the CNS. The spatial and temporal profiles of Gas6 expression suggest that it could potentially serve as the physiologically relevant ligand for Tyro-3 in the postnatal rat nervous system.
Experimental Eye Research, Oct 1, 2002
The phagocytosis of photoreceptor outer segments by retinal pigment epithelial (RPE) cells plays ... more The phagocytosis of photoreceptor outer segments by retinal pigment epithelial (RPE) cells plays a critical role in preserving normal retinal function. Recently the receptor protein tyrosine kinase (RTK) Mer, has been shown to be necessary for this cellular process to take place. Gas6, the ligand for the Mer RTK, can speci®cally and selectively stimulate the phagocytosis of photoreceptor outer segments (OS) by normal cultured rat RPE cells, as we have previously reported. The Gas6 protein has been shown to associate with plasma membrane phosphatidylserine by its amino-terminal portion, while its carboxylterminal portion can bind and activate Mer and its related RTKs, Axl and Tyro-3. Given the capability of Gas6 to interact with more than one molecule, we have performed a series of experiments to further dissect the interactions of Gas6 with the OS and RPE and to determine the speci®c calcium requirements necessary for Gas6 to exert its stimulatory effect on phagocytosis. These experiments show that Gas6 must bind to OS before the stimulation of OS ingestion can occur and that this binding requires a Ca 2 concentration of 500±600 mM. The same Ca 2 concentration is required for the Gas6 mediated stimulation of OS ingestion. We further demonstrate that in order to bind to OS and to stimulate OS phagocytosis, Gas6 requires g-carboxylation in a vitamin K-dependent reaction. By analogy with other systems, we propose that Ca 2 mediates the linkage between the g-carboxyglutamic acid (Gla)-rich N-terminal region of Gas6 with phospholipids, presumably phosphatidylserine, in the plasma membrane of the OS. Only after this binding has occurred can Gas6 interact with receptor molecule(s) on the surface of the RPE, and activate RPE cell signaling pathways leading to OS ingestion. These studies further underscore the importance of Gas6 in the phagocytic function of the RPE and open new avenues of investigation to understand the molecular events mediated and triggered by Gas6, and its interaction with the OS and RPE.
Neuroscience, Apr 1, 1993
Neuron-glia-related cell adhesion molecule (Nr-CAM) is a recently characterized cell adhesion mol... more Neuron-glia-related cell adhesion molecule (Nr-CAM) is a recently characterized cell adhesion molecule in the family of immunoglobulin-related molecules of which the neural cell adhesion molecule, N-CAM, is the prototype. Nr-CAM shares structural properties with another member of this family (neuron-glia CAM, Ng-CAM) and both molecules exhibit homophilic and heterophilic binding propcrties. To understand better the role of such molecules in development, we have examined the sites of synthesis and expression of Nr-CAM by means of in situ hybridization and immunohistochemistry. Both methods indicated that N&AM is expressed only in the nervous system. The molecule was observed on neurons in both the peripheral and central nervous systems and on epitheliai floor plate cells in the spinal cord, but it was absent in the germinal zones. The protein was present on perikarya, but was found preferentially on axonal tracts. As observed for messenger RNAs specifying other cell adhesion molecules, messenger RNA for NC-CAM was localized in the perikarya.
Journal of Cell Biology, Nov 1, 1992
The extracellular matrix molecule cytotactin
Journal of comparative neurology, Sep 18, 2000
Tyro-3, Axl, and Mer are three related receptor protein-tyrosine kinases (RPTKs) characterized by... more Tyro-3, Axl, and Mer are three related receptor protein-tyrosine kinases (RPTKs) characterized by an extracellular domain exhibiting significant amino acid sequence similarity to neural cell adhesion molecules. The molecule Gas6 (for growth arrest-specific gene-6) has been shown to activate each of these receptors. Gas6 is expressed extensively in the central nervous system (CNS), suggesting that interactions between Gas6 and its receptors are likely to have physiologically relevant functions. To identify and localize the relevant Gas6/RPTK pairs, we have characterized the developmental expression of Tyro-3, Axl, and Mer in rat CNS using blotting and mRNA in situ hybridization analyses. Throughout development, Tyro-3 was the most widely expressed of the three receptors in the CNS, with Axl and Mer detected in only a limited number of sites in the adult. Tyro-3 expression was low in the embryo and increased markedly during early postnatal stages, with a time course paralleling that of synaptogenesis. Axl and Mer were expressed at low but relatively constant levels throughout development. In the cerebellum, all three receptors were found in Purkinje cells, and Tyro-3 was also detected in both granule neurons and Bergmann glia. Insofar as Gas6 has been previously shown to also be expressed by Purkinje cells, it may be engaged in both autocrine and paracrine signaling. The three receptors were also detected in cerebellar white matter, primarily during myelination. In the cortex, Tyro-3 was expressed at high levels during postnatal development and in the adult. Beginning at P6 in the hippocampus, Tyro-3 was expressed at high levels in CA1 pyramidal neurons and at lower levels in CA3 and was not detected in dentate granule neurons. Axl and Mer were found in the molecular layer of the dentate gyrus and were absent from the pyramidal and dentate granule neurons. In that Gas6 is expressed throughout the pyramidal cell layer, it may activate these cells in both an autocrine and a paracrine manner. These studies provide initial clues for elucidating the cellular functions of the Axl subfamily members and suggest potential complex Gas6/RPTK as well as RPTK/RPTK signaling interactions in the mature and developing CNS.
Cells Tissues Organs, 1995
The functional units in most inductive and morphogenetic processes in the embryo are not single c... more The functional units in most inductive and morphogenetic processes in the embryo are not single cells, but rather collectives of interacting cells that give rise to the tissues and organs. Cell adhesion molecules (CAMs) are involved in defining cell collectives and their borders as they interact during inductive events in morphogenesis. The expression patterns of CAMs are highly dynamic and changes are known to occur during epithelial-mesenchymal transformations. Alterations in CAM expression are correlated with changes in morphology. Conversely, experimentally induced changes in morphology result in changes in CAM expression. The structure, function, distribution, and control of CAM gene expression are presented in this review, and discussed with regard to their importance to normal developmental processes, particularly epithelial-mesenchymal transformations.
International Journal of Developmental Neuroscience, Oct 31, 2014
PubMed, Oct 1, 2002
The phagocytosis of photoreceptor outer segments by retinal pigment epithelial (RPE) cells plays ... more The phagocytosis of photoreceptor outer segments by retinal pigment epithelial (RPE) cells plays a critical role in preserving normal retinal function. Recently the receptor protein tyrosine kinase (RTK) Mer, has been shown to be necessary for this cellular process to take place. Gas6, the ligand for the Mer RTK, can specifically and selectively stimulate the phagocytosis of photoreceptor outer segments (OS) by normal cultured rat RPE cells, as we have previously reported. The Gas6 protein has been shown to associate with plasma membrane phosphatidylserine by its amino-terminal portion, while its carboxyl-terminal portion can bind and activate Mer and its related RTKs, Axl and Tyro-3. Given the capability of Gas6 to interact with more than one molecule, we have performed a series of experiments to further dissect the interactions of Gas6 with the OS and RPE and to determine the specific calcium requirements necessary for Gas6 to exert its stimulatory effect on phagocytosis. These experiments show that Gas6 must bind to OS before the stimulation of OS ingestion can occur and that this binding requires a Ca(2+) concentration of 500-600 microM. The same Ca(2+) concentration is required for the Gas6 mediated stimulation of OS ingestion. We further demonstrate that in order to bind to OS and to stimulate OS phagocytosis, Gas6 requires gamma-carboxylation in a vitamin K-dependent reaction. By analogy with other systems, we propose that Ca(2+) mediates the linkage between the gamma-carboxyglutamic acid (Gla)-rich N-terminal region of Gas6 with phospholipids, presumably phosphatidylserine, in the plasma membrane of the OS. Only after this binding has occurred can Gas6 interact with receptor molecule(s) on the surface of the RPE, and activate RPE cell signaling pathways leading to OS ingestion. These studies further underscore the importance of Gas6 in the phagocytic function of the RPE and open new avenues of investigation to understand the molecular events mediated and triggered by Gas6, and its interaction with the OS and RPE.
Neuroscience, Dec 1, 2007
Protein phosphorylation serves as a critical biochemical regulator of short-term and long-term sy... more Protein phosphorylation serves as a critical biochemical regulator of short-term and long-term synaptic plasticity. Receptor protein tyrosine kinases (RPTKs) including members of the trk, eph and erbB subfamilies have been shown to modulate signaling cascades that influence synaptic function in the central nervous system (CNS). Tyro3 is one of 3 RPTKs belonging to the "TAM" receptor family, which also includes Axl and Mer. Tyro3 is the most widely expressed of these receptors in the CNS. Despite recent advances suggesting roles for members of this receptor family in the reproductive and immune systems, their functions in the CNS remain largely unexplored. In an effort to elucidate the roles of Tyro3 and its ligand Gas6 in the hippocampus and cortex, we performed a detailed study of the localization and signaling of Tyro3 polypeptides in hippocampal and cortical neurons. Tyro3 was readily detected in dendrites and in the soma where it was distributed in a punctate pattern. Tyro3 exhibited only a limited level of co-localization with PSD-95, suggesting that while located within dendrites, it was not confined to the postsynaptic compartment. In addition, Tyro3 was also identified in the axons and growth cones of immature neurons. The prominent expression of Tyro3 in dendrites suggested that it may be capable of modulating signaling pathways triggered by synaptic transmission. We have provided evidence in support of this role by demonstrating that Gas6 induced the phosphorylation of Tyro3 in cortical neurons in vitro, resulting in the recruitment of the mitogen-activated protein kinase (MAPK) and the phosphoinositide-3 kinase (PI(3)K) signaling pathways. As these pathways play critical roles in the induction of hippocampal long-term potentiation (LTP), these findings suggest that Tyro3 signaling may influence synaptic plasticity in the dendritic compartment of hippocampal and cortical neurons.
Journal of Neuroinflammation, May 15, 2011
Background: Axl, together with Tyro3 and Mer, constitute the TAM family of receptor tyrosine kina... more Background: Axl, together with Tyro3 and Mer, constitute the TAM family of receptor tyrosine kinases. In the nervous system, Axl and its ligand Growth-arrest-specific protein 6 (Gas6) are expressed on multiple cell types. Axl functions in dampening the immune response, regulating cytokine secretion, clearing apoptotic cells and debris, and maintaining cell survival. Axl is upregulated in various disease states, such as in the cuprizone toxicity-induced model of demyelination and in multiple sclerosis (MS) lesions, suggesting that it plays a role in disease pathogenesis. To test for this, we studied the susceptibility of Axl-/-mice to experimental autoimmune encephalomyelitis (EAE), an animal model for multiple sclerosis. Methods: WT and Axl-/-mice were immunized with myelin oligodendrocyte glycoprotein (MOG) 35-55 peptide emulsified in complete Freund's adjuvant and injected with pertussis toxin on day 0 and day 2. Mice were monitored daily for clinical signs of disease and analyzed for pathology during the acute phase of disease. Immunological responses were monitored by flow cytometry, cytokine analysis and proliferation assays. Results: Axl-/-mice had a significantly more severe acute phase of EAE than WT mice. Axl-/-mice had more spinal cord lesions with larger inflammatory cuffs, more demyelination, and more axonal damage than WT mice during EAE. Strikingly, lesions in Axl-/-mice had more intense Oil-RedO staining indicative of inefficient clearance of myelin debris. Fewer activated microglia/macrophages (Iba1+) were found in and/or surrounding lesions in Axl-/-mice relative to WT mice. In contrast, no significant differences were noted in immune cell responses between naïve and sensitized animals. Conclusions: These data show that Axl alleviates EAE disease progression and suggests that in EAE Axl functions in the recruitment of microglia/macrophages and in the clearance of debris following demyelination. In addition, these data provide further support that administration of the Axl ligand Gas6 could be therapeutic for immunemediated demyelinating diseases.
International journal of molecular sciences, Mar 16, 2024
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
Journal of comparative neurology, 2000
Proceedings of the National Academy of Sciences of the United States of America, Aug 1, 1991
International Journal of Developmental Neuroscience, Nov 16, 2006
Introduction: Recently, it has been shown that H2O2, one of the most stable and long-lived ROS, a... more Introduction: Recently, it has been shown that H2O2, one of the most stable and long-lived ROS, are produced and released during neuromuscular activity and play important roles in modulating synaptic activity. Less is known, however, about the underlying mechanism of H2O2 on synaptic facilitation. Methods: Whole-cell patch clamp recording in one-day-old Xenopus nerve–muscle coculture were used. Results: Exclusion of calcium from culture medium or bath application of calcium channel inhibitor either cadmium, nifedipine, verapamil or -conotoxin significantly reduced H2O2 induced synaptic facilitation. Pretreatment of catalase and N-acetyl-l-cysteine effectively hampered H2O2 induced SSC frequency elevation. Preincubation of the cultures with either inhibitor of IP3 receptor XeC or ryanodine receptor TMB8 or depletion of ER Ca stores with thapsigargin failed to abolish the synaptic facilitation elicited by H2O2. Bath application of mitochondrial Na–Ca exchanger inhibitor FCCP or CGP37157 partialy obstructed the increase of SSC frequency in response to H2O2. Treating cells with protein kinase C inhibitors either H-9 or GF 109203X effectively occluded the increase of SSC frequency elicited by H2O2. Furthermore, application of either phosphatidyl inositol 3 kinase (PI3K) inhibitor wortmannin or phospholipase C (PLC ) inhibitor U73122, also abolished H2O2-induced facilitation of synaptic transmission. Discussion: Overall, results from our current study indicate that both calcium influx through Land N-type Ca channel and efflux from mitochondrial are responsible for H2O2-induced SSC frequency facilitation. This is done via PLC /PI3K, PKC activation and Na–Ca exchanger blockade.
Carolina Digital Repository (University of North Carolina at Chapel Hill), 2013
MerTK, a receptor tyrosine kinase (RTK) of the TYRO3/AXL/MerTK family, is expressed in myeloid li... more MerTK, a receptor tyrosine kinase (RTK) of the TYRO3/AXL/MerTK family, is expressed in myeloid lineage cells in which it acts to suppress proinflammatory cytokines following ingestion of apoptotic material. Using syngeneic mouse models of breast cancer, melanoma, and colon cancer, we found that tumors grew slowly and were poorly metastatic in MerTK-/mice. Transplantation of MerTK-/bone marrow, but not wild-type bone marrow, into lethally irradiated MMTV-PyVmT mice (a model of metastatic breast cancer) decreased tumor growth and altered cytokine production by tumor CD11b + cells. Although MerTK expression was not required for tumor infiltration by leukocytes, MerTK-/leukocytes exhibited lower tumor cell-induced expression of wound healing cytokines, e.g., IL-10 and growth arrest-specific 6 (GAS6), and enhanced expression of acute inflammatory cytokines, e.g., IL-12 and IL-6. Intratumoral CD8 + T lymphocyte numbers were higher and lymphocyte proliferation was increased in tumor-bearing MerTK-/mice compared with tumor-bearing wild-type mice. Antibody-mediated CD8 + T lymphocyte depletion restored tumor growth in MerTK-/mice. These data demonstrate that MerTK signaling in tumor-associated CD11b + leukocytes promotes tumor growth by dampening acute inflammatory cytokines while inducing wound healing cytokines. These results suggest that inhibition of MerTK in the tumor microenvironment may have clinical benefit, stimulating antitumor immune responses or enhancing immunotherapeutic strategies.
Molecular and Cellular Neuroscience, Jun 1, 2004
Acetylcholine receptor (AChR) genes are transcribed selectively in synaptic nuclei of skeletal mu... more Acetylcholine receptor (AChR) genes are transcribed selectively in synaptic nuclei of skeletal muscle fibers, leading to accumulation of the mRNAs encoding AChR subunits at synaptic sites. The signals that regulate synapse-specific transcription remain elusive, though Neuregulin-1 is considered a favored candidate. Here, we show that motor neurons and terminal Schwann cells express neuregulin-2, a neuregulin-1-related gene. In skeletal muscle, Neuregulin-2 protein is concentrated at synaptic sites, where it accumulates adjacent to terminal Schwann cells. Neuregulin-2 stimulates AChR transcription in cultured myotubes expressing ErbB4, as well as ErbB3 and ErbB2, but not in myotubes expressing only ErbB3 and ErbB2. Thus, Neuregulin-2 is a candidate for a signal that regulates synaptic differentiation.
The Journal of Neuroscience, May 14, 2008
The TAM family of receptor protein tyrosine kinases comprises three known members, namely Tyro3, ... more The TAM family of receptor protein tyrosine kinases comprises three known members, namely Tyro3, Axl, and Mer. These receptors are widely expressed in the nervous system, including by oligodendrocytes, the cell type responsible for myelinating the CNS. We examined the potential role of the TAM family and of their principle cognate ligand, Gas6 (growth arrest gene 6), in modulating the phenotype of the cuprizone model of demyelination. We found that the expression profiles of Axl, Mer, and Gas6 mRNA were increased in the corpus callosum in a temporal profile correlating with the increased migration and proliferation of microglia/macrophages in this model. In contrast, expression of Tyro3 decreased, correlating with the loss of oligodendrocytes. Gas6 both promoted in vitro survival of oligodendrocytes (39.3 Ϯ 3.1 vs 11.8 Ϯ 2.4%) and modulated markers of activation in purified cultures of microglia (tumor necrosis factor ␣ mRNA expression was reduced ϳ48%). In Gas6 Ϫ/Ϫ mice subjected to cuprizone-challenge, demyelination was greater than in control mice, within the rostral region of the corpus callosum, as assessed by luxol fast blue staining (myelination reduced by 36%) and by ultrastructural analysis. An increased loss of Gst-(glutathione S-transferase-)-positive oligodendrocytes was also identified throughout the corpus callosum of Gas6 Ϫ/Ϫ mice. Microglial marker expression (ionized calcium-binding adapter molecule 1) was increased in Gas6 Ϫ/Ϫ mice but was restricted to the rostral corpus callosum. Therefore, TAM receptor activation and regulation can independently influence both oligodendrocyte survival and the microglial response after CNS damage.
Experimental Eye Research, Oct 1, 2001
The function and viability of vertebrate photoreceptors requires the daily phagocytosis of photor... more The function and viability of vertebrate photoreceptors requires the daily phagocytosis of photoreceptor outer segments (OS) by the adjacent retinal pigment epithelium (RPE). We demonstrate here a critical role in this process for Gas6 and by implication one of its receptor protein tyrosine kinases (RTKs), Mertk (Mer). Gas6 speci®cally and selectively stimulates the phagocytosis of OS by normal cultured rat RPE cells. The magnitude of the response is dose-dependent and shows an absolute requirement for calcium. By contrast the Royal College of Surgeons (RCS) rat RPE cells, in which a mutation in the gene Mertk results in the expression of a truncated, non-functional receptor, does not respond to Gas6. These data strongly suggest that activation of Mertk by its ligand, Gas6, is the speci®c signaling pathway responsible for initiating the ingestion of shed OS. Moreover, photoreceptor degeneration in the RCS rat retina, which lacks Mertk, and in humans with a mutation in Mertk, strongly suggests that the Gas6/Mertk signaling pathway is essential for photoreceptor viability. We believe that this is the ®rst demonstration of a speci®c function for Gas6 in the eye.
Experimental Neurology, Jul 1, 1990
Evidence is accumulating that molecules involved in cell-cell and cell-substratum interactions ar... more Evidence is accumulating that molecules involved in cell-cell and cell-substratum interactions are important in the establishment and maintenance of borders between cell groups during development. In this report, we review evidence supporting this conclusion, particularly in regard to the role of adhesion molecules in the formation of cell collectives and in the modulation of cell and neurite movements.
Brain Research, 1999
Gas6 (growth arrest specific gene-6) is a ligand for members of the Axl subfamily of receptor pro... more Gas6 (growth arrest specific gene-6) is a ligand for members of the Axl subfamily of receptor protein-tyrosine kinases. One of these receptors, Tyro-3, is widely expressed in the central nervous system. We have used biochemical and histological techniques, including in situ hybridization, to determine the expression patterns of Gas6 mRNA and protein during development. Gas6 is widely expressed in the rat central nervous system (CNS) beginning at late embryonic stages and its levels remain high in the adult. Gas6 is detected as a single 85 kDa protein, which is encoded by a single 2.5 kb mRNA species. At embryonic day 14 it is detected in the heart, blood vessels, testes, choroid plexus, and in the ventral spinal cord. In the adult, Gas6 is expressed in the cerebral cortex, (predominantly in layer V), the piriform cortex, and the hippocampus (areas CA1, CA3 and the dentate gyrus). It is also expressed in thalamic and hypothalamic structures, the midbrain, and in a subset of motor and trigeminal nuclei. In the cerebellum, it is expressed in Purkinje neurons and deep cerebellar nuclei. Protein S, a protein related to Gas6, is only detected at low levels in the CNS. The spatial and temporal profiles of Gas6 expression suggest that it could potentially serve as the physiologically relevant ligand for Tyro-3 in the postnatal rat nervous system.
Experimental Eye Research, Oct 1, 2002
The phagocytosis of photoreceptor outer segments by retinal pigment epithelial (RPE) cells plays ... more The phagocytosis of photoreceptor outer segments by retinal pigment epithelial (RPE) cells plays a critical role in preserving normal retinal function. Recently the receptor protein tyrosine kinase (RTK) Mer, has been shown to be necessary for this cellular process to take place. Gas6, the ligand for the Mer RTK, can speci®cally and selectively stimulate the phagocytosis of photoreceptor outer segments (OS) by normal cultured rat RPE cells, as we have previously reported. The Gas6 protein has been shown to associate with plasma membrane phosphatidylserine by its amino-terminal portion, while its carboxylterminal portion can bind and activate Mer and its related RTKs, Axl and Tyro-3. Given the capability of Gas6 to interact with more than one molecule, we have performed a series of experiments to further dissect the interactions of Gas6 with the OS and RPE and to determine the speci®c calcium requirements necessary for Gas6 to exert its stimulatory effect on phagocytosis. These experiments show that Gas6 must bind to OS before the stimulation of OS ingestion can occur and that this binding requires a Ca 2 concentration of 500±600 mM. The same Ca 2 concentration is required for the Gas6 mediated stimulation of OS ingestion. We further demonstrate that in order to bind to OS and to stimulate OS phagocytosis, Gas6 requires g-carboxylation in a vitamin K-dependent reaction. By analogy with other systems, we propose that Ca 2 mediates the linkage between the g-carboxyglutamic acid (Gla)-rich N-terminal region of Gas6 with phospholipids, presumably phosphatidylserine, in the plasma membrane of the OS. Only after this binding has occurred can Gas6 interact with receptor molecule(s) on the surface of the RPE, and activate RPE cell signaling pathways leading to OS ingestion. These studies further underscore the importance of Gas6 in the phagocytic function of the RPE and open new avenues of investigation to understand the molecular events mediated and triggered by Gas6, and its interaction with the OS and RPE.
Neuroscience, Apr 1, 1993
Neuron-glia-related cell adhesion molecule (Nr-CAM) is a recently characterized cell adhesion mol... more Neuron-glia-related cell adhesion molecule (Nr-CAM) is a recently characterized cell adhesion molecule in the family of immunoglobulin-related molecules of which the neural cell adhesion molecule, N-CAM, is the prototype. Nr-CAM shares structural properties with another member of this family (neuron-glia CAM, Ng-CAM) and both molecules exhibit homophilic and heterophilic binding propcrties. To understand better the role of such molecules in development, we have examined the sites of synthesis and expression of Nr-CAM by means of in situ hybridization and immunohistochemistry. Both methods indicated that N&AM is expressed only in the nervous system. The molecule was observed on neurons in both the peripheral and central nervous systems and on epitheliai floor plate cells in the spinal cord, but it was absent in the germinal zones. The protein was present on perikarya, but was found preferentially on axonal tracts. As observed for messenger RNAs specifying other cell adhesion molecules, messenger RNA for NC-CAM was localized in the perikarya.
Journal of Cell Biology, Nov 1, 1992
The extracellular matrix molecule cytotactin
Journal of comparative neurology, Sep 18, 2000
Tyro-3, Axl, and Mer are three related receptor protein-tyrosine kinases (RPTKs) characterized by... more Tyro-3, Axl, and Mer are three related receptor protein-tyrosine kinases (RPTKs) characterized by an extracellular domain exhibiting significant amino acid sequence similarity to neural cell adhesion molecules. The molecule Gas6 (for growth arrest-specific gene-6) has been shown to activate each of these receptors. Gas6 is expressed extensively in the central nervous system (CNS), suggesting that interactions between Gas6 and its receptors are likely to have physiologically relevant functions. To identify and localize the relevant Gas6/RPTK pairs, we have characterized the developmental expression of Tyro-3, Axl, and Mer in rat CNS using blotting and mRNA in situ hybridization analyses. Throughout development, Tyro-3 was the most widely expressed of the three receptors in the CNS, with Axl and Mer detected in only a limited number of sites in the adult. Tyro-3 expression was low in the embryo and increased markedly during early postnatal stages, with a time course paralleling that of synaptogenesis. Axl and Mer were expressed at low but relatively constant levels throughout development. In the cerebellum, all three receptors were found in Purkinje cells, and Tyro-3 was also detected in both granule neurons and Bergmann glia. Insofar as Gas6 has been previously shown to also be expressed by Purkinje cells, it may be engaged in both autocrine and paracrine signaling. The three receptors were also detected in cerebellar white matter, primarily during myelination. In the cortex, Tyro-3 was expressed at high levels during postnatal development and in the adult. Beginning at P6 in the hippocampus, Tyro-3 was expressed at high levels in CA1 pyramidal neurons and at lower levels in CA3 and was not detected in dentate granule neurons. Axl and Mer were found in the molecular layer of the dentate gyrus and were absent from the pyramidal and dentate granule neurons. In that Gas6 is expressed throughout the pyramidal cell layer, it may activate these cells in both an autocrine and a paracrine manner. These studies provide initial clues for elucidating the cellular functions of the Axl subfamily members and suggest potential complex Gas6/RPTK as well as RPTK/RPTK signaling interactions in the mature and developing CNS.
Cells Tissues Organs, 1995
The functional units in most inductive and morphogenetic processes in the embryo are not single c... more The functional units in most inductive and morphogenetic processes in the embryo are not single cells, but rather collectives of interacting cells that give rise to the tissues and organs. Cell adhesion molecules (CAMs) are involved in defining cell collectives and their borders as they interact during inductive events in morphogenesis. The expression patterns of CAMs are highly dynamic and changes are known to occur during epithelial-mesenchymal transformations. Alterations in CAM expression are correlated with changes in morphology. Conversely, experimentally induced changes in morphology result in changes in CAM expression. The structure, function, distribution, and control of CAM gene expression are presented in this review, and discussed with regard to their importance to normal developmental processes, particularly epithelial-mesenchymal transformations.
International Journal of Developmental Neuroscience, Oct 31, 2014
PubMed, Oct 1, 2002
The phagocytosis of photoreceptor outer segments by retinal pigment epithelial (RPE) cells plays ... more The phagocytosis of photoreceptor outer segments by retinal pigment epithelial (RPE) cells plays a critical role in preserving normal retinal function. Recently the receptor protein tyrosine kinase (RTK) Mer, has been shown to be necessary for this cellular process to take place. Gas6, the ligand for the Mer RTK, can specifically and selectively stimulate the phagocytosis of photoreceptor outer segments (OS) by normal cultured rat RPE cells, as we have previously reported. The Gas6 protein has been shown to associate with plasma membrane phosphatidylserine by its amino-terminal portion, while its carboxyl-terminal portion can bind and activate Mer and its related RTKs, Axl and Tyro-3. Given the capability of Gas6 to interact with more than one molecule, we have performed a series of experiments to further dissect the interactions of Gas6 with the OS and RPE and to determine the specific calcium requirements necessary for Gas6 to exert its stimulatory effect on phagocytosis. These experiments show that Gas6 must bind to OS before the stimulation of OS ingestion can occur and that this binding requires a Ca(2+) concentration of 500-600 microM. The same Ca(2+) concentration is required for the Gas6 mediated stimulation of OS ingestion. We further demonstrate that in order to bind to OS and to stimulate OS phagocytosis, Gas6 requires gamma-carboxylation in a vitamin K-dependent reaction. By analogy with other systems, we propose that Ca(2+) mediates the linkage between the gamma-carboxyglutamic acid (Gla)-rich N-terminal region of Gas6 with phospholipids, presumably phosphatidylserine, in the plasma membrane of the OS. Only after this binding has occurred can Gas6 interact with receptor molecule(s) on the surface of the RPE, and activate RPE cell signaling pathways leading to OS ingestion. These studies further underscore the importance of Gas6 in the phagocytic function of the RPE and open new avenues of investigation to understand the molecular events mediated and triggered by Gas6, and its interaction with the OS and RPE.
Neuroscience, Dec 1, 2007
Protein phosphorylation serves as a critical biochemical regulator of short-term and long-term sy... more Protein phosphorylation serves as a critical biochemical regulator of short-term and long-term synaptic plasticity. Receptor protein tyrosine kinases (RPTKs) including members of the trk, eph and erbB subfamilies have been shown to modulate signaling cascades that influence synaptic function in the central nervous system (CNS). Tyro3 is one of 3 RPTKs belonging to the "TAM" receptor family, which also includes Axl and Mer. Tyro3 is the most widely expressed of these receptors in the CNS. Despite recent advances suggesting roles for members of this receptor family in the reproductive and immune systems, their functions in the CNS remain largely unexplored. In an effort to elucidate the roles of Tyro3 and its ligand Gas6 in the hippocampus and cortex, we performed a detailed study of the localization and signaling of Tyro3 polypeptides in hippocampal and cortical neurons. Tyro3 was readily detected in dendrites and in the soma where it was distributed in a punctate pattern. Tyro3 exhibited only a limited level of co-localization with PSD-95, suggesting that while located within dendrites, it was not confined to the postsynaptic compartment. In addition, Tyro3 was also identified in the axons and growth cones of immature neurons. The prominent expression of Tyro3 in dendrites suggested that it may be capable of modulating signaling pathways triggered by synaptic transmission. We have provided evidence in support of this role by demonstrating that Gas6 induced the phosphorylation of Tyro3 in cortical neurons in vitro, resulting in the recruitment of the mitogen-activated protein kinase (MAPK) and the phosphoinositide-3 kinase (PI(3)K) signaling pathways. As these pathways play critical roles in the induction of hippocampal long-term potentiation (LTP), these findings suggest that Tyro3 signaling may influence synaptic plasticity in the dendritic compartment of hippocampal and cortical neurons.
Journal of Neuroinflammation, May 15, 2011
Background: Axl, together with Tyro3 and Mer, constitute the TAM family of receptor tyrosine kina... more Background: Axl, together with Tyro3 and Mer, constitute the TAM family of receptor tyrosine kinases. In the nervous system, Axl and its ligand Growth-arrest-specific protein 6 (Gas6) are expressed on multiple cell types. Axl functions in dampening the immune response, regulating cytokine secretion, clearing apoptotic cells and debris, and maintaining cell survival. Axl is upregulated in various disease states, such as in the cuprizone toxicity-induced model of demyelination and in multiple sclerosis (MS) lesions, suggesting that it plays a role in disease pathogenesis. To test for this, we studied the susceptibility of Axl-/-mice to experimental autoimmune encephalomyelitis (EAE), an animal model for multiple sclerosis. Methods: WT and Axl-/-mice were immunized with myelin oligodendrocyte glycoprotein (MOG) 35-55 peptide emulsified in complete Freund's adjuvant and injected with pertussis toxin on day 0 and day 2. Mice were monitored daily for clinical signs of disease and analyzed for pathology during the acute phase of disease. Immunological responses were monitored by flow cytometry, cytokine analysis and proliferation assays. Results: Axl-/-mice had a significantly more severe acute phase of EAE than WT mice. Axl-/-mice had more spinal cord lesions with larger inflammatory cuffs, more demyelination, and more axonal damage than WT mice during EAE. Strikingly, lesions in Axl-/-mice had more intense Oil-RedO staining indicative of inefficient clearance of myelin debris. Fewer activated microglia/macrophages (Iba1+) were found in and/or surrounding lesions in Axl-/-mice relative to WT mice. In contrast, no significant differences were noted in immune cell responses between naïve and sensitized animals. Conclusions: These data show that Axl alleviates EAE disease progression and suggests that in EAE Axl functions in the recruitment of microglia/macrophages and in the clearance of debris following demyelination. In addition, these data provide further support that administration of the Axl ligand Gas6 could be therapeutic for immunemediated demyelinating diseases.