ABSTRACT
The field of gene therapy has recently witnessed a number of major conceptual changes. Besides the traditional thinking that comprises the use of viral vectors for the delivery of a given therapeutic gene, a number of original approaches have been recently envisaged, focused on using vectors carrying genes to further modify basal ganglia circuits of interest. It is expected that these approaches will ultimately induce a therapeutic potential being sustained by gene-induced changes in brain circuits. Among others, at present, it is technically feasible to use viral vectors to (1) achieve a controlled release of neurotrophic factors, (2) conduct either a transient or permanent silencing of any given basal ganglia circuit of interest, (3) perform an in vivo cellular reprogramming by promoting the conversion of resident cells into dopaminergic-like neurons, and (4) improving levodopa efficacy over time by targeting aromatic L-amino acid decarboxylase. Furthermore, extensive research efforts based on viral vectors are currently ongoing in an attempt to better replicate the dopaminergic neurodegeneration phenomena inherent to the progressive intraneuronal aggregation of alpha-synuclein. Finally, a number of incoming strategies will soon emerge over the horizon, these being sustained by the underlying goal of promoting alpha-synuclein clearance, such as, for instance, gene therapy initiatives based on increasing the activity of glucocerebrosidase. To provide adequate proof-of-concept on safety and efficacy and to push forward true translational initiatives based on these different types of gene therapies before entering into clinical trials, the use of non-human primate models undoubtedly plays an instrumental role.
Subject(s)
Genetic Therapy , Genetic Vectors , Parkinson Disease/therapy , alpha-Synuclein/genetics , Animals , Disease Models, Animal , Parkinson Disease/genetics , PrimatesABSTRACT
Skeletal muscle fibers form in overlapping, but distinct phases that depend on the generation of temporally different lineages of myogenic cells. During primary myogenesis (E10.5-E12.5 in the mouse), embryonic myoblasts fuse homotypically to generate primary fibers, whereas during later development (E14.5-E17.5), fetal myoblasts differentiate into secondary fibers. How these myogenic waves are regulated remains largely unknown. Studies have been hampered by the lack of markers which would distinguish embryonic from fetal myoblast populations. We show here that the homeobox gene Arx is strongly expressed in differentiating embryonic muscle, downstream of myogenic basic helix-loop-helix (bHLH) genes. Its expression progressively decreases during development. When overexpressed in the C2C12 myogenic cell line, Arx enhances differentiation. Accordingly, it stimulates the transcriptional activity from the Myogenin promoter and from multimerized E-boxes when co-expressed with MyoD and Mef2C in CH310T1/2. Furthermore, Arx co-immunoprecipitates with Mef2C, suggesting that it participates in the transcriptional regulatory network acting in embryonic muscle. Finally, embryonic myoblasts isolated from Arx-deficient embryos show a delayed differentiation in vivo together with an enhanced clonogenic capacity in vitro. We propose here that Arx acts as a novel positive regulator of embryonic myogenesis by synergizing with Mef2C and MyoD and by establishing an activating loop with Myogenin.
Subject(s)
Genes, Homeobox , Homeodomain Proteins/metabolism , Muscle Development , Muscle, Skeletal/embryology , Myoblasts, Skeletal/metabolism , Transcription Factors/metabolism , Animals , Cell Differentiation , Cell Line , Gene Expression Regulation, Developmental , Homeodomain Proteins/genetics , MEF2 Transcription Factors , Mice , Mice, Mutant Strains , Muscle Fibers, Skeletal/cytology , Muscle Fibers, Skeletal/physiology , Muscle, Skeletal/cytology , Muscle, Skeletal/metabolism , MyoD Protein/metabolism , Myoblasts, Skeletal/cytology , Myogenic Regulatory Factors/metabolism , Myogenin/metabolism , Transcription Factors/geneticsABSTRACT
Aristaless-related homeobox gene (ARX) is an important paired-type homeobox gene involved in the development of human brain. The ARX gene mutations are a significant contributor to various forms of X-chromosome-linked mental retardation with and without additional features including epilepsy, lissencephaly with abnormal genitalia, hand dystonia or autism. Here we demonstrate that the human ARX protein is a potent transcriptional repressor, which binds to Groucho/transducin-like enhancer of split (TLE) co-factor proteins and the TLE1 in particular through its octapeptide (Engrailed homology repressor domain (eh-1) homology) domain. We show that the transcription repression activity of ARX is modulated by two strong repression domains, one located within the octapeptide domain and the second in the region of the polyalanine tract 4, and one activator domain, the aristaless domain. Importantly, we show that the transcription repression activity of ARX is affected by various naturally occurring mutations. The introduction of the c.98T>C (p.L33P) mutation results in the lack of binding to TLE1 protein and relaxed transcription repression. The introduction of the two most frequent ARX polyalanine tract expansion mutations increases the repression activity in a manner dependent on the number of extra alanines. Interestingly, deletions of alanine residues within polyalanine tracts 1 and 2 show low or no effect. In summary we demonstrate that the ARX protein is a strong transcription repressor, we identify novel ARX interacting proteins (TLE) and offer an explanation of a molecular pathogenesis of some ARX mutations, including the most frequent ARX mutations, the polyalanine tract expansion mutations, c.304ins(GCG)7 and c.428_451dup.
Subject(s)
Enhancer Elements, Genetic/physiology , Homeodomain Proteins/genetics , Mutation , Repressor Proteins/metabolism , Transcription Factors/genetics , Transcription, Genetic/physiology , Age Factors , Alanine/genetics , Animals , Brain/cytology , Cells, Cultured , Embryo, Mammalian , Gene Expression Regulation, Developmental/physiology , Humans , Immunoprecipitation/methods , In Situ Hybridization/methods , Mice , Neurons/metabolism , Transducin/metabolism , Transfection/methodsABSTRACT
Homeobox genes encode a particular class of transcription factors that are involved in several different developmental processes such as specification of regional identity, cell determination and proliferation. In particular, during early brain morphogenesis, they provide a genetic code, which generates single rhombomere identity in the hindbrain (Science 284 (1999) 2168) and interneurons specification in the ventral neural tube (Nat. Rev. Genet. 1 (2000) 20). We have isolated a paired homeobox containing gene, which has been recently named Dmbx1 (Mech. Dev. 110 (2002) 241). Dmbx1 protein can be listed into the paired-like class, due to the highest homology in its homeodomain, with several other members of this family. With the exception of olfactory neurons, Dmbx1 is expressed only in the developing central nervous system and in particular during early determination and successive differentiation of the midbrain and caudal diencephalon. Interestingly, Dmbx1 expression labels cerebellar granule progenitors at the onset of differentiation and spinal cord V0 interneurons.
Subject(s)
Brain/embryology , Gene Expression , Homeodomain Proteins/biosynthesis , Homeodomain Proteins/metabolism , Nerve Tissue Proteins/biosynthesis , Amino Acid Sequence , Animals , Base Sequence , Cell Differentiation , Cell Division , Cell Lineage , Central Nervous System/embryology , Diencephalon/embryology , Homeodomain Proteins/chemistry , Homeodomain Proteins/genetics , In Situ Hybridization , Mesencephalon/embryology , Mice , Models, Genetic , Molecular Sequence Data , Nerve Tissue Proteins/chemistry , Nerve Tissue Proteins/genetics , Neurons/metabolism , Otx Transcription Factors , Protein Structure, Tertiary , Sequence Homology, Amino Acid , Time FactorsABSTRACT
We report the isolation of noz1, a novel zebrafish zinc finger gene which displays sequence similarity to Drosophila nocA. noz1 transcripts are detected at the shield stage within the germ ring and excluded from the most dorsal region. By the end of gastrulation, noz1 is expressed in the presumptive hindbrain and spinal cord as well as in the forming tailbud. During somitogenesis noz1 shows a dynamic expression in the midbrain-hindbrain boundary, hindbrain and spinal cord. This results, at 24 hpf, in a graded expression with the highest level in rhombomeres 2 and 3, and the lowest in the spinal cord. Expression analysis in swirl and chordino mutants as well as in retinoic acid treated embryos indicate that noz1 is activated by BMP antagonists and neural posteriorizing signals.
Subject(s)
Brain/embryology , DNA-Binding Proteins/biosynthesis , DNA-Binding Proteins/genetics , Drosophila Proteins , Spinal Cord/embryology , Transcription Factors , Zebrafish Proteins , Zinc Fingers , Amino Acid Motifs , Amino Acid Sequence , Animals , Cloning, Molecular , DNA, Complementary/metabolism , DNA-Binding Proteins/chemistry , Drosophila , In Situ Hybridization , Insect Proteins/biosynthesis , Insect Proteins/chemistry , Molecular Sequence Data , Mutation , Sequence Homology, Amino Acid , Time Factors , Tretinoin/pharmacology , ZebrafishABSTRACT
Most of the gene candidates for the control of developmental programmes that underlie brain morphogenesis in vertebrates are the homologues of Drosophila genes coding for signalling molecules or transcription factors. Among these, the orthodenticle group includes the Drosophila orthodenticle (otd) and the vertebrate Otx1 and Otx2 genes, which are mostly involved in fundamental processes of anterior neural patterning. These genes encode transcription factors that recognise specific target sequences through the DNA binding properties of the homeodomain. In Drosophila, mutations of otd cause the loss of the anteriormost head neuromere where the gene is transcribed, suggesting that it may act as a segmentation "gap" gene. In mouse embryos, the expression patterns of Otx1 and Otx2 have shown a remarkable similarity with the Drosophila counterpart. This suggested that they could be part of a conserved control system operating in the brain and different from that coded by the HOX complexes controlling the hindbrain and spinal cord. To verify this hypothesis a series of mouse models have been generated in which the functions of the murine genes were: (i) fully inactivated, (ii) replaced with each others, (iii) replaced with the Drosophila otd gene. Otx1-/- mutants suffer from epilepsy and are affected by neurological, hormonal, and sense organ defects. Otx2-/- mice are embryonically lethal, they show gastrulation impairments and fail in specifying anterior neural plate. Analysis of the Otx1-/-; Otx2+/- double mutants has shown that a minimal threshold level of the proteins they encode is required for the correct positioning of the midbrain-hindbrain boundary (MHB). In vivo otd/Otx reciprocal gene replacement experiments have provided evidence of a general functional equivalence among otd, Otx1 and Otx2 in fly and mouse. Altogether these data highlight a crucial role for the Otx genes in specification, regionalization and terminal differentiation of rostral central nervous system (CNS) and lead to hypothesize that modification of their regulatory control may have influenced morphogenesis and evolution of the brain.
Subject(s)
Brain/physiology , Gene Expression Regulation, Developmental/physiology , Homeodomain Proteins/genetics , Nerve Tissue Proteins/genetics , Trans-Activators/genetics , Transcription Factors , Animals , Biological Evolution , Brain/metabolism , Drosophila Proteins , Humans , Otx Transcription FactorsABSTRACT
Citron-kinase (Citron-K) has been proposed by in vitro studies as a crucial effector of Rho in regulation of cytokinesis. To further investigate in vivo its biologic functions, we have inactivated Citron-K gene in mice by homologous recombination. Citron-K-/- mice grow at slower rates, are severely ataxic, and die before adulthood as a consequence of fatal seizures. Their brains display defective neurogenesis, with depletion of specific neuronal populations. These abnormalities arise during development of the central nervous system due to altered cytokinesis and massive apoptosis. Our results indicate that Citron-K is essential for cytokinesis in vivo but only in specific neuronal precursors. Moreover, they suggest a novel molecular mechanism for a subset of human malformative syndromes of the CNS.
Subject(s)
Apoptosis/genetics , Cell Division/genetics , Neurodegenerative Diseases/genetics , Neurons/metabolism , Protein Serine-Threonine Kinases/genetics , Animals , Ataxia/etiology , Brain/embryology , Brain/pathology , Cyclin D1/metabolism , DNA/biosynthesis , Intracellular Signaling Peptides and Proteins , Mice , Mice, Knockout , Neurodegenerative Diseases/complications , Neurodegenerative Diseases/pathology , Neurons/pathology , Polyploidy , Protein Serine-Threonine Kinases/biosynthesis , Protein Serine-Threonine Kinases/deficiency , Seizures/etiology , Stem Cells/metabolism , Stem Cells/pathology , rho-Associated KinasesABSTRACT
We report on the expression pattern of a novel EGF- containing gene named Maeg. RNA in situ studies indicate that Maeg is first activated during specification of the early lateral dermatome, and continues to be expressed in all the dermatome derivatives as the dermis of the trunk, the hair follicles, and the mesenchyme of the cranio-facial region.
Subject(s)
Embryonic and Fetal Development/genetics , Epidermal Growth Factor/genetics , Glycoproteins , Growth Substances , Neoplasm Proteins , Peptides , Animals , Base Sequence , Calcium-Binding Proteins , Cell Adhesion Molecules , DNA Primers/genetics , Gene Expression Regulation, Developmental , Genetic Markers , In Situ Hybridization , Mice , RNA, Messenger/genetics , RNA, Messenger/metabolism , Somites/metabolismABSTRACT
The BarH1 and BarH2 ( Bar ) Drosophila genes are homeobox-containing genes, which are required for the fate determination of external sensory organs in the fly. By means of a bioinformatic approach, we have identified in mouse and human two homeobox genes highly related to the Bar Drosophila genes, Barhl1 and Barhl2. While Barhl1 represents a novel gene, Barhl2 turned out to correspond to the mBH1 cDNA recently described in rat. We isolated and sequenced the full-length mouse Barhl1 and mapped both the human BARHL1 and BARHL2 genes to chromosomes 9q34 and 1p22, respectively. Detailed analysis of the murine Barhl1 expression pattern by in situ hybridization revealed that this transcript is exclusively expressed in restricted domains of the developing CNS, which suggests that this gene, similar to its Drosophila counterparts BarH1 and BarH2, may play a crucial role in cell fate determination of neural structures. In particular, Barhl1 showed specific domains of expression in the diencephalon and in the rhombencephalon where it was found to be expressed in migrating cells giving rise to the cerebellar external granular layer and to specific populations of dorsal sensory interneurons of the spinal cord. Thus, Barhl1 function may be required for the generation of these specific subtypes of neuronal progenitors. Furthermore, the mapping assignment and the expression pattern make BARHL1 an attractive positional candidate gene for a form of Joubert syndrome, a rare developmental anomaly of the cerebellum in humans.
Subject(s)
Central Nervous System/metabolism , Genes, Homeobox , Homeodomain Proteins/biosynthesis , Homeodomain Proteins/genetics , Nerve Tissue Proteins/biosynthesis , Nerve Tissue Proteins/genetics , Neurons/metabolism , Amino Acid Sequence , Animals , Central Nervous System/embryology , Cerebellum/abnormalities , Cerebellum/metabolism , Chromosomes, Human, Pair 1 , Chromosomes, Human, Pair 9 , DNA, Complementary/metabolism , Embryo, Mammalian/metabolism , Gene Library , Humans , In Situ Hybridization , Mice , Molecular Sequence Data , Repressor Proteins , Sequence Analysis, DNA , Sequence Homology, Amino Acid , Spinal Cord/metabolismABSTRACT
The homeobox gene Otx2 is expressed in the anterior neural tube with a sharp limit at the midbrain/hindbrain junction (the isthmic organizer). Otx2 inactivation experiments have shown that this gene is essential for the development of its expression domain. Here we investigate whether the caudal limit of Otx2 expression is instrumental in positioning the isthmic organizer and in specifying midbrain versus hindbrain fate, by ectopically expressing Otx2 in the presumptive anterior hindbrain using a knock-in strategy into the En1 locus. Transgenic offspring display a cerebellar ataxia. Morphological and histological studies of adult transgenic brains reveal that most of the anterior cerebellar vermis is missing, whereas the inferior colliculus is complementarily enlarged. During early neural pattern formation expression of the midbrain markers Wnt1 and Ephrin-A5, the isthmic organizer markers Pax2 and Fgf-8 and the hindbrain marker Gbx2 are shifted caudally in the presumptive hindbrain territory. These findings show that the caudal limit of Otx2 expression is sufficient for positioning the isthmic organizer and encoding caudal midbrain fate within the mid/hindbrain domain.
Subject(s)
Mesencephalon/embryology , Nerve Tissue Proteins/physiology , Rhombencephalon/embryology , Trans-Activators/physiology , Animals , Cell Line , Cerebellum/abnormalities , Cerebellum/embryology , Embryo, Mammalian/metabolism , Embryo, Mammalian/physiology , Embryonic Induction , Ephrin-A5 , Homeodomain Proteins/genetics , Membrane Proteins/metabolism , Mice , Mice, Inbred C57BL , Mice, Transgenic , Mutation , Nerve Tissue Proteins/genetics , Otx Transcription Factors , Recombinant Fusion Proteins/genetics , Trans-Activators/geneticsABSTRACT
The A-Myb transcription factor is structurally related to the c-myb proto-oncogene and is involved in the control of proliferation and/or differentiation of mature B lymphocytes. We have shown previously by PCR analysis that A-myb is preferentially expressed in CD38+ CD39- sIgM- mature B cells. We demonstrate here, using in situ hybridization, that A-myb expression is restricted to the dark zone of human tonsils and lymph nodes. Furthermore, we show that A-Myb expression is cell cycle regulated both in tonsillar B cells and in Burkitt's lymphoma cell lines, being detectable only in the S and G2/M phases of the cell cycle and not in G0/G1 phase. Strong proliferation of resting human B cells induced in vitro by a variety of physiologic signals, including anti-mu, CD40 ligand, IL-2, IL-4, IL-6, IL-13, IFN-gamma, TNF-alpha, anti-CD19, and anti-CD20, failed to induce A-myb expression, suggesting that proliferation alone is not sufficient for A-myb expression in the absence of induction of a true centroblast phenotype. Finally, we show that differentiation of germinal center B cells in vitro toward either memory or plasma cells is accompanied by rapid down-regulation of A-myb expression. We conclude that A-myb is a marker of centroblasts generated in vivo.
Subject(s)
B-Lymphocytes/physiology , Germinal Center/physiology , Proto-Oncogene Proteins/genetics , Trans-Activators/genetics , Biomarkers , Cell Cycle , Cell Differentiation , Cell Division , Gene Expression Regulation , Humans , Lymphocyte Activation , Proto-Oncogene MasABSTRACT
We show in this report that the human myeloid leukemia cell line GFD8 is a useful model to compare the biological function of the structurally related c-Myb and B-Myb proto-oncogenes and to investigate the c-myb domains required for this function. GFD8 cells are dependent for growth on granulocyte-macrophage colony-stimulating factor and differentiate in response to phorbol myristate acetate (PMA). We have stably transfected this cell line with constructs constitutively expressing c-Myb or B-Myb. Deregulated expression of both c-Myb and B-Myb inhibited the differentiation observed in response to PMA and, in particular, the induction of the CD11b and CD11c antigens on the cell surface, and the induction of adherence. Furthermore, c-Myb and B-Myb enhanced expression of CD13 upon PMA treatment. Although deregulated Myb expression did not alter the growth factor dependence of the cells, it led to an increase in G2 relative to G1 arrest in cells induced to differentiate in response to PMA, whereas control vector-transfected cells were blocked mostly in G1. This decrease in G1 block took place despite normal induction of the cyclin-dependent kinase inhibitor protein p21 (CIP1/WAF1). Thus, GFD8 cells stably expressing the human B-Myb protein behaved in a manner indistinguishable from those stably expressing C-Myb for both differentiation and cell cycle parameters. In agreement with these findings and differently from most previous reports, transactivation assays show that B-myb can indeed act as a strong activator of transcription. Finally, we demonstrated that although the DNA-binding domain of c-myb is required for both the differentiation block and the shift in cell cycle after PMA treatment, phosphorylation by casein kinase II and mitogen-activated protein kinase at positions 11 and 12 or 532 of c-myb, respectively, are not. We conclude that c-Myb and B-Myb may activate a common cellular program in the GFD8 cell line involved in both differentiation and cell cycle control.
Subject(s)
Cell Cycle Proteins , Cell Cycle/genetics , Cell Differentiation/genetics , DNA-Binding Proteins/genetics , Leukemia, Myeloid/pathology , Proto-Oncogene Proteins/genetics , Trans-Activators/genetics , Transcription Factors/genetics , Antigens, Differentiation/biosynthesis , Blotting, Western , Cell Cycle/drug effects , Cell Differentiation/drug effects , Cell Division/drug effects , Cell Division/genetics , Cyclin-Dependent Kinase Inhibitor p21 , Cyclins/metabolism , DNA-Binding Proteins/drug effects , Gene Expression Regulation , Granulocyte-Macrophage Colony-Stimulating Factor/metabolism , Humans , Leukemia, Myeloid/genetics , Leukemia, Myeloid/metabolism , Proto-Oncogene Proteins/drug effects , Proto-Oncogene Proteins c-myb , Tetradecanoylphorbol Acetate/pharmacology , Trans-Activators/drug effects , Transcription Factors/drug effects , Transfection , Tumor Cells, CulturedABSTRACT
Emx1 and Emx2 genes are known to be involved in mammalian forebrain development. In order to investigate the evolution of the Emx gene family in vertebrates, a phylogenetic analysis was carried out on the Emx genes sequenced in man, mice, frogs, coelacanths and zebrafish. The results demonstrated the existence of two clades (Emx1 and Emx2), each grouping one of the two genes of the investigated taxa. The only exception was the zebrafish Emx1-like gene which turned out to be a sister group to both the Emx1 and Emx2 clusters. Such striking sequence divergence observed for the zebrafish Emx1-like gene could indicate that it is not orthologous to the other Emx1 genes, and therefore, in vertebrates there must be three Emx genes. Alternatively, if the zebrafish emx1 gene is orthologous to the tetrapod one, it must have undergone to strong diversifying selection.
Subject(s)
Evolution, Molecular , Homeodomain Proteins/genetics , Nerve Tissue Proteins/genetics , Animals , Humans , Mice , Phylogeny , Prosencephalon/growth & development , Transcription Factors , Vertebrates , Xenopus laevis , ZebrafishABSTRACT
Vascular endothelial growth factor (VEGF) is a previously discovered angiogenic factor that seems to influence the neoangiogenesis of neoplastic and non-neoplastic tissues. Substantial experimental evidence links tumor growth and metastasis with blood vessel formation. Tumor angiogenesis can be induced by factors released by the tumor cells themselves. A variety of transformed cell lines expresses the VEGF transcript and secretes an EGF-like protein, suggesting that this angiogenic factor may be one of the mediators of tumor angiogenesis. By Northern blot analysis and in situ hybridization, we investigated the expression of VEGF transcript in human ovarian and endometrial neoplasms. Messenger RNA encoding VEGF was detected in all tissues studied and was more densely expressed in endometrial carcinoma. VEGF expression was also identified in cells obtained from ovarian and endometrial ascitic fluid. VEGF mRNA, detected by in situ hybridization, was identified in the epithelial cells of endometrial adenocarcinoma. This distribution was localized primarily in the apices of the papillae. The prominence of VEGF mRNA levels in human ovarian and endometrial tumors demonstrates that VEGF may be involved in promoting tumor angiogenesis and stroma generation, acting as an endothelial cell mitogen.
Subject(s)
Endometrial Neoplasms/genetics , Endothelial Growth Factors/genetics , Gene Expression , Lymphokines/genetics , Ovarian Neoplasms/genetics , RNA, Messenger/analysis , Adenocarcinoma/genetics , Blotting, Northern , Epithelium/chemistry , Female , Humans , In Situ Hybridization , Tissue Distribution , Vascular Endothelial Growth Factor A , Vascular Endothelial Growth FactorsABSTRACT
Insights into the complex structure of the forebrain and its regulation have recently come from the analysis of the expression of genes that are likely to be involved in regionalization of this structure. We cloned four new homeo box genes, Emx1, Emx2, Otx1 and Otx2, and showed that in day 10 mouse embryos their expression domains are continuous regions of the developing brain contained within each other in the sequence Emx1 < Emx2 < Otx1 < Otx2. Recently Otx1 has been found to be specifically expressed during neurogenesis of layer 5 and 6 in the developing cerebral cortex. In order to better understand the role of Emx1 and Emx2 in the maturation of the cortex we analysed by in situ hybridization their expression patterns in the developing mouse cerebral cortex, from embryonic day 12.5 to adulthood. We found that Emx2 is expressed exclusively in proliferating cells of the ventricular zone whereas Emx1 is expressed in both proliferating and differentiated neurons, throughout the cortical layers and during all the developmental stages examined. Therefore, Emx2 gene products might control some biological parameters of the proliferation of cortical neuroblasts or of the subsequent cell migration of postmitotic neurons, leaving the cortical germinal zone. Conversely, Emx1 expression, which is confined exclusively to the dorsal telencephalon, characterizes most cortical neurons during proliferation, differentiation, migration and postnatal development and maturation.
Subject(s)
Cerebral Cortex/metabolism , Gene Expression Regulation, Developmental/physiology , Genes, Homeobox , Animals , Cell Differentiation/physiology , Cell Division/physiology , Cell Movement/physiology , Cerebral Cortex/embryology , Cerebral Cortex/growth & development , Epithelial Cells , MiceABSTRACT
The myb family of genes include the virally encoded v-myb oncogene, its normal cellular equivalent c-myb and two related members called A-myb and B-myb. They are all transcription factors that recognize the same DNA sequence (PyAACG/TG) and are all involved in the regulation of proliferation and differentiation in different cell types, including hematopoietic cells. C-myb is most highly expressed in hematopoietic cells and its oncogenic activation leads to transformation of these cells. Several lines of evidence have demonstrated that c-myb regulates both the proliferation and differentiation of hematopoietic cells of different lineages. The mechanisms of action of c-myb and v-myb are becoming clearer, mostly through the study of the different genes that are regulated by these transcription factors and the cofactors with which c-myb and v-myb co-operate. More recently the biological and biochemical functions of the B-myb and A-myb gene products have been investigated. Evidence for the function of the different members of the myb family in relation to hematopoietic proliferation and differentiation is presented, and the different roles of the myb genes are discussed.
Subject(s)
Blood Cells/cytology , DNA-Binding Proteins/physiology , Multigene Family , Oncogenes , Transcription Factors/physiology , Animals , Cell Differentiation/physiology , Cell Division/physiology , Oncogene Proteins v-myb , Proto-Oncogene Proteins/physiology , Proto-Oncogene Proteins c-myb , Retroviridae Proteins, Oncogenic/physiology , Trans-Activators/physiology , Transcription Factors/geneticsABSTRACT
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.
Subject(s)
Brain/embryology , Brain/physiology , Genes, Homeobox/genetics , Animals , Gene Expression Regulation, Developmental/physiology , MiceABSTRACT
We cloned the chick homologue of the mouse Otx2 gene, c-otx2, and analyzed its expression pattern during gastrulation. During mouse embryogenesis, Otx2 expression is first detected in the entire epiblast and after the formation of the primitive streak becomes confined to the most anterior region of the embryo corresponding to presumptive fore- and mid-brain. Similarly, two distinct phases of c-otx2 expression were observed in the chick. c-otx2 transcripts were first detected in the unincubated egg and up to stage XIII, in all epiblast, and forming hypoblast and mesoblast cells. During primitive streak progression, c-otx2 expression becomes progressively restricted to anterior regions and is mainly associated with Hensen's node. When the extension of the streak is maximal, transcripts are only found in Hensen's node. A second phase of c-otx2 expression starts during streak regression. c-otx2 transcripts are lost from the node and present in higher abundance in anterior neuroectoderm and mesendoderm, with the exception of forming notochord and floor plate. The first phase of expression bears strong similarity with that of c-gsc, a gene shown to be a marker for cells that have organizer activity in the chick. Therefore, we compared the expression of the two genes by double staining on the same embryo. This analysis demonstrated that c-otx2 is transcribed first and its expression in the hypoblast precedes that of c-gsc. On the other hand, c-gsc is an earlier marker of primitive streak cells. The expression domains of the two genes transiently overlap in Hensen's node and anterior mesendoderm, whereas only c-otx2 is expressed in neuroectodermal areas. The second phase of c-otx2 expression is sensitive to an early treatment with retinoic acid. This treatment abolishes c-otx2 expression in mesendoderm and restricts it to most anterior regions in the forming neural plate. In conclusion, our results suggest that c-otx2 expression is first associated with cells with an anterior mesendoderm fate and subsequently extends to anterior neuroectoderm.