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Edoardo Boncinelli - One of the best experts on this subject based on the ideXlab platform.
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Ciba Foundation Symposium 193 - Development of the Cerebral Cortex - Emx and Otx Gene Expression in the Developing Mouse Brain
Ciba Foundation symposium, 2007Co-Authors: Edoardo Boncinelli, Massimo Gulisano, Fabio Spada, Vania BroccoliAbstract:The homeobox genes Emx1, EMX2, Otx1 and Otx2 are all expressed in the rostral brain of embryos at E10. Their expression domains are continuous regions of the developing brain contained within each other, such that the expression domain of Otx2 contains that of the other three genes, the expression domain of Otx1 contains that of Emx1 and EMX2, and the expression domain of EMX2 contains that of Emx1. The Emx1 expression domain includes the dorsal telencephalon and it has a posterior boundary slightly anterior to that between the presumptive diencephalon and telencephalon, whereas the Otx2 expression domain covers almost the entire forebrain and midbrain. Starting from E10.75, Otx2 expression disappears progressively from the presumptive cerebral cortex, whereas Emx1, EMX2 and Otx1 are expressed in this structure until late gestation. In particular, EMX2 appears to be expressed exclusively in the germinal ventricular zone of the developing cerebral cortex.
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EMX2 and Pax6 Control Regionalization of the Pre-neuronogenic Cortical Primordium
Cerebral cortex (New York N.Y. : 1991), 2002Co-Authors: Luca Muzio, Edoardo Boncinelli, Peter Gruss, Barbara Dibenedetto, Anastassia Stoykova, Antonello MallamaciAbstract:It has recently been demonstrated that the transcription factor genes EMX2 and Pax6, expressed in the developing cerebral cortex along two complementary tangential gradients, are essential for the shaping of the cortical areal profile at late developmental ages, when cortical neuronogenesis is almost completed. In this study we addressed the question of whether cortical regionalization is already affected in EMX2 and Pax6 loss of function mutants at the beginning of neuronogenesis. By comparing expression patterns of selected molecular markers in these mutants at this age, we found that: (i) EMX2 and Pax6 are necessary for the establishment of their own specific expression profiles and are able to down-regulate each other; and (ii) absence of functional EMX2 or PAX6 proteins results in reduction of caudal-medial and rostral-lateral cortical regions, respectively, as well as in impairment of the WNT signalling center at the medial-caudal edge of the cortical field, crucial for cortical growth. These results suggest that pre-neuronogenic cortical regionalization may rely on mutual interactions between these two transcription factors and that the late areal phenotype of EMX2(-/-) and Pax6(-/-) mutants may possibly arise from both misconfiguration of the cortical molecular protomap and distortion of the cortical growth profile.
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Emx homeogenes and mouse brain development
Trends in neurosciences, 2000Co-Authors: Chiara Cecchi, Edoardo BoncinelliAbstract:Abstract Mammalian homeogenes of the Emx family, Emx1 and EMX2 , are expressed in the developing cerebral cortex and are involved in the patterning of the rostral brain. Although very little is known about the role of Emx1 , details of the function of EMX2 are emerging from the observation of cortical phenotypes in normal and mutant mice. EMX2 is expressed in proliferating neuroblasts and in the so-called postmitotic Cajal–Retzius cells, known to control migration of cortical neurons. The graded distribution of EMX2 homeoprotein suggests a potential role for EMX2 in the subdivision of the cortex into territories and possibly areas.
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Mouse forebrain development. The role of EMX2 homeobox gene.
Comptes rendus de l'Academie des sciences. Serie III Sciences de la vie, 1999Co-Authors: Chiara Cecchi, Antonello Mallamaci, Edoardo BoncinelliAbstract:Over the last few years great progress has been made in the understanding of the formation of the mouse forebrain. Among the genes involved in this process, the mouse Emx homeobox genes Emx1 and particularly EMX2 play a primary role. Here we describe the mRNA and protein expression related to EMX2 in the developing mouse telencephalon, as well as the results obtained studying the corresponding knock-out mice. Our findings indicate a role for this gene in the specification of the forebrain via the control of cell proliferation, as well as in guiding neuronal migration during development through the cortical plate. These studies will hopefully enable us to better understand the molecular mechanisms underlying the formation of the mouse cerebral cortex as well as to establish relevant interactions between the various proteins present in this region of the brain.
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EMX2 protein in the developing mouse brain and olfactory area
Mechanisms of development, 1998Co-Authors: Antonello Mallamaci, Edoardo Boncinelli, Raffaella Iannone, Paola Briata, Luisa Pintonello, Sara Mercurio, Giorgio CorteAbstract:Abstract The distribution of EMX2, the protein product of the homeobox gene EMX2, was analyzed in the developing mouse CNS by means of a polyclonal antibody we raised against it. The protein is present in the rostral brain, the olfactory area and a set of scattered cells lying between the nasal pits and the telencephalon. In the cortical neuroepithelium EMX2 is expressed all along the rostro-caudal axis in a graded distribution with a caudal-medial maximum and a rostral-lateral minimum. Anti-EMX2 immunoreactivity is also detectable in Cajal–Retzius cells as well as in apical dendrites of marginal neurons of the cortical plate. We also observe that the EMX2 and EMX1 homeoproteins display complementary expression patterns in olfactory bulbs and amygdaloid complex. Here, they demarcate different neuronal populations, involved in processing olfactory information coming from the vomero-nasal organ and from the main olfactory epithelium, respectively. EMX2 is also detectable in mesencephalic structures, such as the optic tectum and tegmentum. The graded distribution of EMX2 along antero-posterior and medial-lateral axes of the primitive cortex prefigures a role of this protein in the subdivision of the cortex in cytoarchitectonic regions and possibly functional areas, whereas its presence in Cajal–Retzius cells suggests a role in the process of cortical lamination.
Antonello Mallamaci - One of the best experts on this subject based on the ideXlab platform.
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EMX2 as a novel tool to suppress glioblastoma
Oncotarget, 2016Co-Authors: Carmen Falcone, Antonio Daga, Giampiero Leanza, Antonello MallamaciAbstract:// Carmen Falcone 1 , Antonio Daga 2 , Giampiero Leanza 3 , Antonello Mallamaci 1 1 Department of Neuroscience, SISSA, 34136 Trieste, Italy 2 DIPOE, IRCCS AOU San Martino IST, 16132 Genoa, Italy 3 Department of Life Sciences, University of Trieste, 34127 Trieste, Italy Correspondence to: Antonello Mallamaci, email: amallama@sissa.it Keywords: EMX2, glioblastoma, gene therapy, EGFR, SOX2 Received: February 28, 2016 Accepted: April 26, 2016 Published: May 13, 2016 ABSTRACT Glioblastoma is a devastating CNS tumour for which no cure is presently available. We wondered if manipulation of EMX2 , which normally antagonizes cortico-cerebral astrogenesis by inhibiting proliferation of astrocyte progenitors, may be employed to counteract it. We found that EMX2 overexpression induced the collapse of seven out of seven in vitro tested glioblastoma cell lines. Moreover, it suppressed four out of four of these lines in vivo . As proven by dedicated rescue assays, the antioncogenic activity of EMX2 originated from its impact on at least six metabolic nodes, which accounts for the robustness of its effect. Finally, in two out of two tested lines, the tumor culture collapse was also achieved when EMX2 was driven by a neural stem cell-specific promoter, likely active within tumor-initiating cells. All that points to EMX2 as a novel, promising tool for therapy of glioblastoma and prevention of its recurrencies.
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Tuning of neocortical astrogenesis rates by EMX2 in neural stem cells.
Neural regeneration research, 2015Co-Authors: Carmen Falcone, Antonello MallamaciAbstract:Generation of astrocytes within the murine developing cerebral cortex mainly takes place during the first postnatal week, after neuronogenesis and prior to the bulk of oligogenesis. This process involves a great variety of highly complex regulatory mechanisms. Astrocytic outputs depend on two primary factors: progressive commitment of multipotent precursors to astroglial fates and proper tuning of proliferation of astrocyte-committed progenitors. To date, several regulatory mechanisms have been identified for the former process, while very little is known about modulation of astroblast proliferation (reviewed in Mallamaci, 2013). Intriguingly, astrogenic rates remain very low during the whole neuronogenic phase, although the mouse cortex is already able to generate astrocytes at E14.5–E15.5, thanks to specific chromatin reconfiguration (Fan et al., 2005). Poor proliferation of astroblasts may contribute to this effect (Seuntjens et al., 2009). Among different factors modulating astrocyte-committed proliferation, the Egf-receptor (EgfR) and the secreted ligand Fgf9 both specifically promote it (Viti et al., 2003; Lum et al., 2009). EMX2, a pleiotropic hub (Gangemi et al., 2006) controlling a variety of neurodevelopmental processes, is highly expressed in the early neuronogenic pallium, while it fades out together with neuronogenesis ending. This temporal progression is possibly linked to the progressive decline of Wnt signals supporting EMX2 expression (Theil et al, 2002) and late arousal of Fgf8 (http://developingmouse.brain-map.org/) antagonizing it (Garel et al., 2003). In a previous in vitro study (Brancaccio et al., 2010), we reported that EMX2 overexpression in neural stem cells (NSCs) leads to a reduction of their astrocytic outputs, due to unknown mechanisms. In the paper highlighted here (Falcone et al., 2014), we showed that this phenomenon occurs also in vivo and dissected its cellular and molecular mechanisms. At the beginning of our study, we verified that the decrease of the ultimate glial output of NSCs induced by EMX2 overexpression takes place also in vivo and it is due to a shrinkage of the proliferating astrogenic pool. We injected a plasmid expressing EMX2 into the lateral ventricular cavity of P0 pups and electroporated it into the cortex. The analysis of P4 mice electroporated cortices revealed a decrease of S100β+ astrocytes and S100β+Ki67+ astroglial proliferating progenitors in EMX2-gain of function (GOF) samples, by about 30% and 50%, respectively (Figure 1A). [Frequencies of these cell types were conversely upregulated in the posterior cortex of E17.5 EMX2+/- embryos, suggesting that the inhibition of astrogenesis elicited by gain-of-function manipulations was not due to a dominant negative effect (Figure 1B)]. Then, these results were replicated in an in vitro model, set up to dissect molecular mechanisms involved in EMX2 antiastrogenic function. E12.5 cortico-cerebral precursors were engineered for conditional EMX2 overexpression, which was activated at the in vitro equivalent of P0. Following this manipulation, the final astroglial output was reduced approximately as much as in vivo. Moreover, it was associated to a prominent shrinkage of the astrogenic proliferating pool. Interestingly, EMX2 impact on astrogenesis depended mainly on cell-autonomous mechanisms. This was verified by mixing a small amount of lentivirus-engineered precursors with an excess of isochronic wild type precursors, conditioning the medium. Even in this situation, EMX2-engineered cells expressing S100β were significantly reduced upon transgene activation. Figure 1 Altered astrocytogenesis upon EMX2 manipulation in vivo. To cast light on molecular mechanisms mediating EMX2 anti-astrogenic effect, we looked at a few well-known genes promoting the expansion of the astrogenic proliferating pool, including EgfR and Fgf9. We found that EMX2 overexpression downregulates both EgfR and Fgf9. Consistently, the same genes were upregulated in EMX2+/- cultures. Functional relevance of EgfR and Fgf9 to EMX2 action was tested by delivering a lentivector driving EgfR expression and, alternatively, the Fgf9 ligand to EMX2-GOF cultures at in vitro equivalent of P0. Both EgfR and Fgf9 were able not only to rescue the normal astroglial output, but to restore wild type astrogenic proliferating rates too. The reconstruction of the pathways leading to EgfR and Fgf9 downregulation represented a step forward in the understanding of EMX2 anti-astrogenic activity. Both EgfR and Fgf9 levels showed scarce sensitivity to exogenous Fgf9 addition and EgfR overexpression, respectively, thus suggesting that EMX2 regulation of astrogenesis may occur along two separated pathways. Regarding EgfR regulation, we suspected that it could be mediated by Bmp signaling. Indeed, EMX2 promotes such signaling (Shimogori et al., 2004), which, in turn, inhibits EgfR expression (Lillien and Raphael, 2000). Interestingly, EMX2 was able to upregulate two established endogenous reporters of Bmp signaling, Id3 and Msx1. Moreover Bmp inhibition by LDN193189 rescued EgfR expression levels in EMX2-GOF samples, while not perturbing them in controls. As for Fgf9, we hypothesized that its regulation might depend on Sox2 repression. We found that EMX2 overexpression almost abolishes Sox2 expression in cortico-cerebral precursors at astrogenesis peak time. Moreover, Sox2 overexpression rescued Fgf9-mRNA levels in EMX2-GOF cultures. Interestingly, a sort of upstream crosslink among these two regulatory branches exists. In fact, on one hand Bmp inhibition restored also Fgf9 expression, on the other hand Sox2 overexpression rescued EgfR levels. Besides, EMX2, while downregulating Fgf9 in control conditions, increased Fgf9 upon Bmp signaling inhibition. This suggests that Bmp signaling could inhibit Fgf9 expression by counteracting an EMX2-dependent stimulatory pathway (Figure 2). Figure 2 Epistatic relationships among EMX2 and mediators of its antiastrogenic activity. Finally, we evaluated the physiological relevance of EMX2 to the confinement of the bulk of astrogenesis to postnatal life. First, we rigorously documented that both EMX2 mRNA and protein levels progressively decrease in cortico-cerebral stem cells from embryonic towards perinatal stages. High EMX2 levels are associated to the neuronogenic phase, whereas EMX2 is barely detectable concomitantly with the arousal of astrogenesis. Then, we assessed consequences of short-term EMX2 overexpression in embryonic NSCs on the size of the astrogenic lineage, unveiled by a Lif-supplemented, pro-differentiative medium. As expected, EMX2 overexpression led to a reduction of the final output of both S100β+ and GFAP+ cells, a result mirrored by cultures loss-of-function for EMX2. In summary: (1) EMX2 overexpression in cortico-cerebral stem cells inhibits astrogenesis in vivo as well as in vitro, by shrinking the proliferating astrogenic pool and thus provoking a pronounced decrease of its ultimate astroglial output (Figure 1); (2) EMX2 exerts its anti-astrogenic function by downregulating EgfR and Fgf9, via promotion of Bmp signaling and Sox2 suppression (Figure 2). These phenomena may be instrumental to fine tuning of cortico-cerebral histogenesis, i.e., the in vivo temporal progression of EMX2 expression levels can help to hold back astrogliogenesis during the neuronogenic phase of development. On the other side, the sensitivity of astrogenic rates to EMX2 expression levels points to EMX2 as an appealing therapeutic tool, suitable for control of reactive gliosis as well as for selective channelling of neural precursors to neuronogenesis in processes of brain repair. The work highlighted in this manuscript was fully supported by SISSA in tramurary funding. The subject of this “Highlight” has been presented at the ISDN 2014 meeting (19-24 July, 2014, Montreal, Canada).
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EMX2 expression levels in NSCs modulate astrogenesis rates by regulating EgfR and Fgf9
Glia, 2014Co-Authors: Carmen Falcone, Marilena Granzotto, Carol Filippis, Antonello MallamaciAbstract:Generation of astrocytes within the developing cerebral cortex is a tightly regulated process, initiating at low level in the middle of neuronogenesis and peaking up after its completion. Astrocytic outputs depend on two primary factors: progression of multipotent precursors toward the astroglial lineage and sizing of the astrogenic proliferating pool. The aim of this study was to investigate the role of the EMX2 homeobox gene in the latter process. We addressed this issue by combined gain- and loss-of-function methods, in vivo as well as in primary cultures of cortico-cerebral precursors. We found that EMX2 overexpression in cortico-cerebral stem cells shrinked the proliferating astrogenic pool, resulting in a severe reduction of the astroglial outcome. We showed that this was caused by EgfR and Fgf9 downregulation and that both phenomena originated from exaggerated Bmp signaling and Sox2 repression. Finally, we provided evidence that in vivo temporal progression of EMX2 levels in cortico-cerebral multipotent precursors contributes to confine the bulk of astrogenesis to postnatal life. EMX2 regulation of astrogenesis adds to a number of earlier developmental processes mastered by this gene. It points to EMX2 as a new promising tool for controlling reactive astrogliosis and optimizing cell-based designs for brain repair. GLIA 2015;63:412–422
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Promotion of cortico-cerebral precursors expansion by artificial pri-miRNAs targeted against the EMX2 locus.
Current gene therapy, 2013Co-Authors: Assunta Diodato, Moira Pinzan, Marilena Granzotto, Antonello MallamaciAbstract:EMX2 encodes for a transcription factor controlling several aspects of cerebral cortex development. Its overexpression promotes self-renewal of young cortico-cerebral precursors, it promotes neuronal rather than gliogenic fates and it protects neuronal progenitors from cell death. These are all key activities for purposes of gene-promoted brain repair. Artificial pri-miRNAs targeting non-coding cis-active modules and/or conserved sequences of the EMX2 locus were delivered to embryonic cortico-cerebral precursors, by lentiviral vectors. A subset of these pri-miRNAs upregulated EMX2, possibly stimulating its transcription. That led to enhanced self-renewal, delayed differentiation and reduced death of neuronally committed precursors, resulting in an appreciable expansion of the neuronogenic precursors pool. This method makes EMX2 overexpression for purposes of brain repair a more feasible goal, avoiding the drawbacks of exogenous gene copies introduction. Interestingly, the two genomic enhancers targeted by these pri-miRNAs were discovered to be naturally transcribed. Their expression profile suggests their possible involvement in regulation of EMX2 transcription.
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Regulation of EMX2 Expression by Antisense Transcripts in Murine Cortico-Cerebral Precursors
PloS one, 2010Co-Authors: Giulia Spigoni, Chiara Gedressi, Antonello MallamaciAbstract:Background EMX2 encodes for a transcription factor expressed in the embryonic intermediate mesoderm and central nervous system (CNS). It is implicated in several aspects of cerebral cortex development, including morphogenetic field specification, arealization, precursor proliferation and lamination. Four EMX2-associated antisense transcripts have been found in the urogenital system; one of them, EMX2OS, has been also detected in the adult brain. Until now, however, nothing is known about expression and function of EMX2OS in the developing CNS. Methodology/Principal Findings By quantitative RT-PCR and in situ hybridization, we reconstructed the EMX2OS expression profile in the embryonic CNS, paying special attention to the developing cerebral cortex. EMX2OS was observed in a number of CNS structures expressing also EMX2. Within the cortex, EMX2OS was detectable in periventricular precursors, expressing the sense transcript, and peaked in newly born post-mitotic neurons not expressing such transcript. By integrating lentiviral gene delivery, RNAi, TetON technology, morpholino-mediated gene knock-down, drug-induced perturbation of gene expression, and quantitative RT-PCR, we addressed possible roles of Ex2 antisense RNA in EMX2 regulation, in primary CNS precursor cultures. We found that, in both cortical precursors and their neuronal progenies, EMX2 antisense RNA contributes to post-transcriptional down-regulation of its sense partner, possibly by a Dicer-promoted mechanism. The same RNA, when delivered to rhombo-spinal precursors, stimulates ectopic expression of EMX2, whereas EMX2 knock-out dramatically impairs EMX2OS transcription. This suggests that, within the developing CNS, a reciprocal EMX2/EMX2OS regulatory loop may normally sustain transcription at the EMX2 locus. Conclusions/Significance This study shows that antisense transcripts may contribute to developmental regulation of a key transcription factor gene implicated in CNS patterning, possibly by complex and multilevel mechanisms. The activation of EMX2 by a short antisense transcript may be a prototype of a method for overexpressing single specific genes, without introducing additional copies of them into the genome.
Joseph X. Dimario - One of the best experts on this subject based on the ideXlab platform.
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EMX2 activates slow myosin heavy chain 2 gene expression in embryonic muscle fibers.
Mechanisms of development, 2017Co-Authors: Kristina Hatch, Amanda Pabon, Joseph X. DimarioAbstract:Avian myogenesis is partly characterized by commitment of distinct myoblast cell lineages to the formation of specific muscle fiber types. Previous studies have identified the transcription factor EMX2 as a regulator of slow myosin heavy chain 2 (MyHC2) gene expression in fast/slow primary muscle fibers. We report here the interaction of EMX2 with the slow MyHC2 transcriptional regulatory region in fast/slow embryonic muscle fibers. Promoter activity and electromobility shift assays localized the site of interaction of EMX2 with the slow MyHC2 gene within a defined binding site located between 3336 and 3326bp from the 3' end of the cloned slow MyHC2 DNA containing the transcriptional regulatory region. Using clonally-derived myoblasts stably committed to the formation of fast/slow muscle fibers, we also report the effect of altered EMX2 gene expression on genome-wide gene expression within these myoblasts. Increased EMX2 gene expression in fast/slow myoblasts caused altered gene expression of 1185 genes, indicating that EMX2 plays a central role in the gene expression profile of embryonic myoblasts.
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genome wide expression analysis and EMX2 gene expression in embryonic myoblasts committed to diverse skeletal muscle fiber type fates
Developmental Dynamics, 2013Co-Authors: Kristina Weimer, Jillian Theobald, Kenneth S. Campbell, Karyn A. Esser, Joseph X. DimarioAbstract:Background: Primary skeletal muscle fibers form during embryonic development and are characterized as fast or slow fibers based on contractile protein gene expression. Different avian primary muscle fiber types arise from myoblast lineages committed to formation of diverse fiber types. To understand the basis of embryonic muscle fiber type diversity and the distinct myoblast lineages that generate this diversity, gene expression analyses were conducted on differentiated muscle fiber types and their respective myoblast precursor lineages. Results: Embryonic fast muscle fibers preferentially expressed 718 genes, and embryonic fast/slow muscle fibers differentially expressed 799 genes. Fast and fast/slow myoblast lineages displayed appreciable diversity in their gene expression profiles, indicating diversity of precursor myoblasts. Several genes, including the transcriptional regulator EMX2, were differentially expressed in both fast/slow myoblasts and muscle fibers vs. fast myoblasts and muscle fibers. EMX2 was localized to nuclei of fast/slow myoblasts and muscle fibers and was not detected in fast lineage cells. Furthermore, EMX2 overexpression and knockdown studies indicated that EMX2 is a positive transcriptional regulator of the slow myosin heavy chain 2 (MyHC2) gene promoter activity in fast/slow muscle fibers. Conclusions: These results indicate the presence of distinct molecular signatures that characterize diverse embryonic myoblast lineages before differentiation. Developmental Dynamics 242:1001–1020, 2013. © 2013 Wiley Periodicals, Inc.
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Genome‐wide expression analysis and EMX2 gene expression in embryonic myoblasts committed to diverse skeletal muscle fiber type fates
Developmental dynamics : an official publication of the American Association of Anatomists, 2013Co-Authors: Kristina Weimer, Jillian Theobald, Kenneth S. Campbell, Karyn A. Esser, Joseph X. DimarioAbstract:Background: Primary skeletal muscle fibers form during embryonic development and are characterized as fast or slow fibers based on contractile protein gene expression. Different avian primary muscle fiber types arise from myoblast lineages committed to formation of diverse fiber types. To understand the basis of embryonic muscle fiber type diversity and the distinct myoblast lineages that generate this diversity, gene expression analyses were conducted on differentiated muscle fiber types and their respective myoblast precursor lineages. Results: Embryonic fast muscle fibers preferentially expressed 718 genes, and embryonic fast/slow muscle fibers differentially expressed 799 genes. Fast and fast/slow myoblast lineages displayed appreciable diversity in their gene expression profiles, indicating diversity of precursor myoblasts. Several genes, including the transcriptional regulator EMX2, were differentially expressed in both fast/slow myoblasts and muscle fibers vs. fast myoblasts and muscle fibers. EMX2 was localized to nuclei of fast/slow myoblasts and muscle fibers and was not detected in fast lineage cells. Furthermore, EMX2 overexpression and knockdown studies indicated that EMX2 is a positive transcriptional regulator of the slow myosin heavy chain 2 (MyHC2) gene promoter activity in fast/slow muscle fibers. Conclusions: These results indicate the presence of distinct molecular signatures that characterize diverse embryonic myoblast lineages before differentiation. Developmental Dynamics 242:1001–1020, 2013. © 2013 Wiley Periodicals, Inc.
Hugh S. Taylor - One of the best experts on this subject based on the ideXlab platform.
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A novel role for the AAA ATPase spastin as a HOXA10 transcriptional corepressor in Ishikawa endometrial cells.
Molecular endocrinology (Baltimore Md.), 2011Co-Authors: Gaurang S. Daftary, Amy M. Tetrault, Elisa M. Jorgensen, Jennifer L. Sarno, Hugh S. TaylorAbstract:Homeobox A10 (HOXA10), a transcription factor required for uterine development and embryo receptivity, functions downstream of estrogen and progesterone in uterine endometrium. HOXA10 represses endometrial expression of empty spiracles homeobox 2 (EMX2), the human ortholog of Drosophila empty spiracles. The ATPases associated with various cellular activities (AAA) ATPase spastin has a well-characterized role in neurotransmitter trafficking. In this study, we characterize a novel role of spastin in transcriptional regulation. We identified spastin as a novel component of the HOXA10 transcriptional complex in Ishikawa nuclear extracts by immunoprecipitation and mass spectrophotometry. Using EMX2 as a model endometrial HOXA10 target gene, we show that the HOXA10-spastin corepressor complex bound the EMX2 promoter in chromatin immunoprecipitation assays. HOXA10 has been previously shown to repress endometrial EMX2 expression. We further observed that, although cotransfection of HOXA10 and spastin continued to repress endometrial EMX2-luciferase expression, the repression was reversed when spastin small interfering RNA was cotransfected with HOXA10. Mutations in the nuclear localization signal sequences of spastin abrogated not only its nuclear translocation but also its colocalization with HOXA10 as well as reversed EMX2-luciferase repression. Here, we describe a novel role for the AAA ATPase spastin in Ishikawa cells as a HOXA10 corepressor of EMX2. Uterine EMX2 levels are inversely related to embryo implantation rates. HOXA10 acts downstream of progesterone and has been shown to facilitate embryo implantation through regulation of endometrial EMX2 expression. Endometrial spastin, therefore, likely has a novel function downstream of estrogen and progesterone in implantation biology as a cofactor of HOXA10.
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EMX2 regulates mammalian reproduction by altering endometrial cell proliferation.
Molecular endocrinology (Baltimore Md.), 2005Co-Authors: Hugh S. Taylor, Xiaolan FeiAbstract:The molecular mechanisms that underlie embryo implantation are poorly understood. Under the control of sex steroids, uterine endometrium undergoes tremendous, yet tightly controlled, proliferation in each estrous cycle to facilitate implantation; disorders of endometrial proliferation underlie several uterine diseases. We have previously identified the EMX2 gene as a transcriptional target of HOXA10 regulation in the reproductive tract. Here we report the function of EMX2 in murine implantation and regulation of endometrial proliferation. We transfected mice on d 2 post coitus with pcDNA3.1/EMX2, EMX2 antisense, or respective controls consisting of empty pcDNA3.1 or a random order oligonucleotide by intrauterine lipofection. Increased expression of EMX2 reduced average implantation rate by approximately 40% (P = 0.00006) resulting in an average number of implanted embryos per litter of 13.7 in the control group to 8.2 in the pcDNA3.1/EMX2-treated group. Neither treatment altered the number of mice attaini...
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EMX2 gene expression in the female reproductive tract and aberrant expression in the endometrium of patients with endometriosis.
The Journal of clinical endocrinology and metabolism, 2004Co-Authors: Gaurang S. Daftary, Hugh S. TaylorAbstract:EMX2 is a transcription factor necessary for reproductive tract development. Sex steroids regulate endometrial HOXA10 expression, which in turn negatively regulates EMX2. In this study, we characterize menstrual cycle-dependent expression of EMX2 in endometrium from women with and without endometriosis. In the absence of endometriosis, EMX2 mRNA levels declined 50% in periimplantation endometrium compared with levels in the proliferative phase. To determine whether the decrease in endometrial EMX2 expression was regulated by endogenous endometrial HOXA10, primary endometrial stromal cells were transfected with an EMX2-reporter construct containing a HOXA10 binding site. Acting via this site, we observed HOXA10-mediated repression of reporter expression. In the endometrium of patients with endometriosis, unlike normal endometrium, EMX2 levels were not decreased in the periimplantation period. We have previously shown that up-regulation of HOXA10 in periimplantation endometrium fails to occur in women with ...
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Transcriptional repression of peri-implantation EMX2 expression in mammalian reproduction by HOXA10.
Molecular and cellular biology, 2003Co-Authors: Patrick J. Troy, Gaurang S. Daftary, Catherine N. Bagot, Hugh S. TaylorAbstract:HOXA10 is necessary for mammalian reproduction; however, its transcriptional targets are not completely defined. EMX2, a divergent homeobox gene, is necessary for urogenital tract development. In these studies we identify and characterize the regulation of EMX2 by HOXA10. By using Northern analysis and in situ hybridization, we found that EMX2 is expressed in the adult urogenital tract in an inverse temporal pattern from HOXA10, suggestive of a negative regulatory relationship. Constitutive expression of HOXA10 diminished EMX2 mRNA, whereas blocking HOXA10 through the use of antisense resulted in high EMX2 mRNA expression. Deletional analysis of the EMX2 5′ regulatory region revealed that a 150-bp element mediated transcriptional repression when cotransfected with pcDNA3.1/HOXA10 in transient-transfection assays. Binding of HOXA10 protein to this element was demonstrated by electrophoretic mobility shift assay and further localized to a consensus HOXA10 binding site within this element by DNase I footprinting. Site-directed mutagenesis abolished binding, as well as the negative transcriptional regulation. Transcriptional activation of empty spiracles, the Drosophila ortholog of EMX2, by Abdominal-B (HOXA10 ortholog) has been previously demonstrated. These findings demonstrate conservation of the transcription factor-target gene relationship, although the direction of regulation is reversed with possible evolutionary implications.
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Mutation analysis of the EMX2 gene in Kallmann’s syndrome ☆
Fertility and Sterility, 1999Co-Authors: Hugh S. Taylor, Karen Block, David P Bick, Richard J Shering, Lawrence C. LaymanAbstract:Abstract Objective: To investigate the possibility that a mutation in the human EMX2 gene may be involved in Kallmann's syndrome. Design: In vitro experiment. Setting: Academic Medical Center. Patients: One hundred and twenty patients with Kallman's syndrome or idiopathic hypogonadotrophic hypogonadism (IHH). Intervention: Peripheral blood leukocytes were used to obtain DNA. Main outcomes measures: Single-stranded conformational polymorphism (SSCP) analysis was used to identify possible mutations of the EMX2 gene. Results: One hundred and twenty patients with Kallmann's syndrome or IHH, had no mutations noted in this gene. Conclusion: It is unlikely that EMX2 mutations are a clinically significant cause of IHH or Kallman's syndrome.
Katie S Kindt - One of the best experts on this subject based on the ideXlab platform.
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directional selectivity of afferent neurons in zebrafish neuromasts is regulated by EMX2 in presynaptic hair cells
eLife, 2018Co-Authors: Young Rae Ji, Doris K Wu, Sunita Warrier, Katie S KindtAbstract:: The orientation of hair bundles on top of sensory hair cells (HCs) in neuromasts of the lateral line system allows fish to detect direction of water flow. Each neuromast shows hair bundles arranged in two opposing directions and each afferent neuron innervates only HCs of the same orientation. Previously, we showed that this opposition is established by expression of EMX2 in half of the HCs, where it mediates hair bundle reversal ( Jiang et al., 2017 ). Here, we show that EMX2 also regulates neuronal selection: afferent neurons innervate either EMX2-positive or negative HCs. In EMX2 knockout and gain-of-function neuromasts, all HCs are unidirectional and the innervation patterns and physiological responses of the afferent neurons are dependent on the presence or absence of EMX2. Our results indicate that EMX2 mediates the directional selectivity of neuromasts by two distinct processes: regulating hair bundle orientation in HCs and selecting afferent neuronal targets.
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Transcription factor EMX2 controls stereociliary bundle orientation of sensory hair cells
eLife, 2017Co-Authors: Tao Jiang, Katie S KindtAbstract:The asymmetric location of stereociliary bundle (hair bundle) on the apical surface of mechanosensory hair cells (HCs) dictates the direction in which a given HC can respond to cues such as sound, head movements, and water pressure. Notably, vestibular sensory organs of the inner ear, the maculae, exhibit a line of polarity reversal (LPR) across which, hair bundles are polarized in a mirror-image pattern. Similarly, HCs in neuromasts of the zebrafish lateral line system are generated as pairs, and two sibling HCs develop opposite hair bundle orientations. Within these sensory organs, expression of the transcription factor EMX2 is restricted to only one side of the LPR in the maculae or one of the two sibling HCs in neuromasts. EMX2 mediates hair bundle polarity reversal in these restricted subsets of HCs and generates the mirror-image pattern of the sensory organs. Downstream effectors of EMX2 control bundle polarity cell-autonomously via heterotrimeric G proteins.