RESUMEN
Runt-homologous molecules are characterized by their DNA binding runt-domain which is highly conserved within bilaterians. The three mammalian runt-genes are master regulators in cartilage/bone formation and hematopoiesis. Historically these features evolved in Craniota and might have been promoted by runt-gene duplication events. The purpose of this study was therefore to investigate how many runt-genes exist in the stem species of chordates, by analyzing the number of runt-genes in what is likely to be the closest living relative of Craniota-amphioxus. To acquire further insight into the possible role of runt-genes in early chordate evolution we have determined the number of runt-genes in sea urchins and have analyzed the runt-expression pattern in this species. Our findings demonstrate the presence of a single runt-gene in amphioxus and sea urchin, which makes it highly likely that the stem species of chordates harbored only a single runt-gene. This suggests that runt-gene duplications occurred later in chordate phylogeny, and are possibly also associated with the evolution of features such as hematopoiesis, cartilage and bone development. In sea urchin embryos runt-expression involves cells of endodermal, mesodermal and ectodermal origin. This complex pattern of expression might reflect the multiple roles played by runt-genes in mammals. A strong runt-signal in the gastrointestinal tract of the sea urchin is in line with runt-expression in the intestine of nematodes and in the murine gastrointestinal tract, and seems to be one of the phylogenetically ancient runt-expression domains.
Asunto(s)
Cordados no Vertebrados/genética , Evolución Molecular , Duplicación de Gen , Proteínas de Neoplasias , Erizos de Mar/genética , Factores de Transcripción/genética , Secuencia de Aminoácidos , Animales , Southern Blotting , Expresión Génica , Humanos , Hibridación in Situ , Datos de Secuencia Molecular , Filogenia , Homología de Secuencia de AminoácidoRESUMEN
The HB9 homeobox gene has been cloned from several vertebrates and is implicated in motor neuron differentiation. In the chick, a related gene, MNR2, acts upstream of HB9 in this process. Here we report an amphioxus homologue of these genes and show that it diverged before the gene duplication yielding HB9 and MNR2. AmphiMnx RNA is detected in two irregular punctate stripes along the developing neural tube, comparable to the distribution of 'dorsal compartment' motor neurons, and also in dorsal endoderm and posterior mesoderm. We propose a new homeobox class, Mnx, to include AmphiMnx, HB9, MNR2 and their Drosophila and echinoderm orthologues; we suggest that vertebrate HB9 is renamed Mnx1 and MNR2 be renamed Mnx2.
Asunto(s)
Cordados no Vertebrados/genética , Proteínas de Drosophila , Genes Homeobox/genética , Proteínas de Homeodominio/clasificación , Proteínas de Homeodominio/genética , Secuencia de Aminoácidos , Animales , Embrión de Pollo , Cordados no Vertebrados/clasificación , Clonación Molecular , Evolución Molecular , Regulación del Desarrollo de la Expresión Génica , Proteínas de Homeodominio/química , Hibridación in Situ , Datos de Secuencia Molecular , Filogenia , ARN Mensajero/análisis , ARN Mensajero/genética , Homología de Secuencia de Aminoácido , Terminología como Asunto , Factores de Transcripción/química , Factores de Transcripción/clasificación , Factores de Transcripción/genéticaRESUMEN
Expression of cytoplasmic intermediate filament (IF) proteins starts in the gastrula with three keratins (k1, Y1, D1) and protein X1. The number of IF proteins expressed increases at the neurula and early larval stages to seven and 11, respectively, and reaches 13 in the adult. Using antibodies specific for a single IF protein the expression patterns of nine of the 13 IF proteins were analyzed at different developmental stages. Keratin k1 of the larval epidermis is replaced in the juvenile by keratin E1. Protein C1 of the larval epidermis persists only weakly and only in the most ventral part of the adult. While down-regulated in the adult epidermis k1 and C1 are major proteins in the atrial epithelium which forms in the later larva. B1 is currently the only IF protein expressed in mesodermally derived tissues such as the muscle tails and some coelomic epithelia. Two-dimensional gels confirm that keratins are the major IF proteins in the nerve cord. Immunogold electronmicroscopy shows that proteins X1 and C2 are present in epidermis and nerve cord in keratin IF.
Asunto(s)
Cordados no Vertebrados/metabolismo , Regulación del Desarrollo de la Expresión Génica , Proteínas de Filamentos Intermediarios/biosíntesis , Animales , Regulación hacia Abajo , Electroforesis en Gel Bidimensional , Epidermis/metabolismo , Epidermis/ultraestructura , Inmunohistoquímica , Queratinas/biosíntesis , Microscopía Fluorescente , Neuronas/metabolismo , Neuronas/ultraestructura , Factores de TiempoRESUMEN
The cloning of three intermediate filament proteins expressed at the gastrula stage (kl, Y1, X1) extends the size of the IF multigene family of Branchiostoma to at least 13 members. This is one of the largest protein families established for the lancelet. Sequence comparisons indicate five keratin orthologs, three of type I (E1, k1, Y1) and two of type II (E2, D1). This assignment is confirmed by the obligatory heteropolymeric polymerisation behaviour of the recombinant proteins. In line with the hetero-coiled-coil principle IF are formed by any stoichiometric mixture of type I and II keratin orthologs. In spite of the strong sequence drift chimeric IF are formed between K8, a human keratin II, and two of the lancelet type I keratins. We discuss whether the remaining 8 IF proteins reflect three additional and potentially cephalochordate-specific subfamilies. The tissue-specific expression patterns of the 5 keratins and some other IF proteins were analysed by immunofluorescence in the adult. Keratins are primarily present in ectodermally derived tissues. Developmental control of the expression of some IF proteins is observed, but three keratins (k1, Y1, D1) and an additional IF protein (X1) detected at the gastrula stage are expressed throughout the life cycle.
Asunto(s)
Cordados no Vertebrados/genética , Queratinas/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Clonación Molecular , Citoplasma/metabolismo , ADN Complementario , Evolución Molecular , Gástrula , Humanos , Queratinas/biosíntesis , Queratinas/clasificación , Datos de Secuencia Molecular , Polímeros , Homología de Secuencia de Aminoácido , Distribución TisularRESUMEN
The full-length sequence and developmental expression of an amphioxus Wnt gene (AmphiWnt8) are described. In amphioxus embryos, the expression patterns of AmphiWnt8 suggest patterning roles in the forebrain, in the hindgut, and in the paraxial mesoderm that gives rise to the muscular somites. Phylogenetic analysis indicates that a single Wnt8 subfamily gene in an ancestral chordate duplicated early in vertebrate evolution into a Wnt8 clade and a Wnt8b clade. Coincident with this gene duplication, the functions of the ancestral AmphiWnt8-like gene appear to have been divided between vertebrate Wnt8b (exclusively neurogenic, especially in the forebrain) and vertebrate Wnt8 (miscellaneous, especially in early somitogenesis). Amphioxus AmphiWnt8 and its vertebrate Wnt8 homologs probably play comparable roles in the early dorsoventral patterning of the embryonic body axis.
Asunto(s)
Tipificación del Cuerpo , Cordados no Vertebrados/genética , Evolución Molecular , Proteínas/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Southern Blotting , Cordados no Vertebrados/embriología , Proteínas del Citoesqueleto , Femenino , Masculino , Datos de Secuencia Molecular , Filogenia , Proteínas/química , Proteínas Wnt , Proteínas de Pez CebraRESUMEN
The organizer is a central feature of vertebrate embryogenesis and is functionally subdivided into the head organizer that gives rise primarily to the prechordal plate and induces forebrain structures, and the trunk/tail organizer that gives rise primarily to the notochord and induces more posterior structures. Goosecoid(gsc) encodes a homeodomain-containing transcription factor that is expressed in the vertebrate head organizer and prechordal plate, and can induce a secondary axis when expressed ectopically. To investigate the evolution of the vertebrate head organizer and prechordal plate, we have cloned and characterized a gsc homolog from the cephalochordate amphioxus. Amphioxus, it is important to note, lacks a prechordal plate in that the notochord extends to the extreme anterior end of the animal, and lacks elaborate differentiation of its forebrain. Gsc expression in amphioxus is initially localized during gastrulation to the mesendodermal layer of the dorsal lip of the blastopore. However, gsc expression in amphioxus is not maintained in anterior axial mesoderm, as is the case with the vertebrate prechordal plate. Rather, gsc is expressed in the dorsal axial mesoderm of the blastopore lip throughout gastrulation, appearing transiently in the presumptive notochord that underlies all regions of the amphioxus brain. The similarities in gsc expression in amphioxus and vertebrates suggest that a primitive version of the head organizer evolved prior to the origin of the vertebrates. The differences in gsc expression can be interpreted either as the loss of the prechordal plate domain in the cephalochordate lineage, or the gain of a distinct gsc-expressing prechordal plate that plays a role in forebrain induction in the vertebrate lineage.
Asunto(s)
Evolución Biológica , Cordados no Vertebrados/embriología , Cordados no Vertebrados/genética , Cabeza/embriología , Proteínas de Homeodominio/genética , Proteínas Represoras , Factores de Transcripción , Secuencia de Aminoácidos , Animales , Proteína Goosecoide , Proteínas de Homeodominio/química , Humanos , Datos de Secuencia Molecular , Filogenia , Homología de Secuencia de AminoácidoRESUMEN
The transcription factor Krox-20 has roles in the maintenance of segmentation and specification of segment identity in the vertebrate hindbrain. Overt hindbrain segmentation is a vertebrate novelty, and is not seen in invertebrate chordates such as amphioxus and tunicates. To test if the roles of Krox-20 are also derived, we cloned a Krox-20 related gene, AmphiKrox, from amphioxus. AmphiKrox is related to a small family of vertebrate Krox genes and is expressed in the most anterior region of the amphioxus brain and in the club shaped gland, a secretory organ that develops in the anterior pharynx. Neither expression domain overlaps with the expression of AmphiHox-1, -2, -3 or -4, suggesting that the roles of Krox-20 in hindbrain segmentation and in Hox gene regulation were acquired concomitant with the duplication of Krox genes in vertebrate evolution.
Asunto(s)
Cordados no Vertebrados/genética , Proteínas de Unión al ADN/genética , Rombencéfalo/embriología , Factores de Transcripción/genética , Secuencia de Aminoácidos , Animales , ADN Complementario , Proteínas de Unión al ADN/química , Proteína 2 de la Respuesta de Crecimiento Precoz , Humanos , Datos de Secuencia Molecular , Homología de Secuencia de Aminoácido , Factores de Transcripción/químicaRESUMEN
We report sequences for nuclear lamins from the teleost fish Danio and six invertebrates. These include two cnidarians (Hydra and Tealia), one priapulid, two echinoderms, and the cephalochordate Branchiostoma. Combining these results with earlier data on Drosophila, Caenorhabditis elegans, and various vertebrates, the following conclusions on lamin evolution can be drawn. First, all invertebrate lamins resemble in size the vertebrate B-type lamin. Second, all lamins described previously for amphibia, birds and mammals as well as the first lamin of a fish, characterized here, show a cluster of 7 to 12 acidic residues in the tail domain. Since this acidic cluster is absent from all invertebrate lamins including that of the cephalochordate Branchiostoma, it was acquired with the vertebrate lineage. The larger A-type lamin of differentiated cells must have arisen subsequently by gene duplication and insertion of an extra exon. This extra exon of the vertebrate A-lamins is the only major change in domain organization in metazoan lamin evolution. Third, the three introns of the Hydra and Priapulus genes correspond in position to the last three introns of vertebrate B-type lamin genes. Thus the entirely different gene organization of the C. elegans and Drosophila Dmo genes seems to reflect evolutionary drift, which probably also accounts for the fact that C. elegans has the most diverse lamin sequence. Finally we discuss the possibility that two lamin types, a constitutively expressed one and a developmentally regulated one, arose independently on the arthropod and vertebrate lineages.
Asunto(s)
Evolución Molecular , Hydra/genética , Invertebrados/genética , Proteínas Nucleares/genética , Filogenia , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Caenorhabditis elegans/genética , Cnidarios/genética , Drosophila/genética , Equinodermos/genética , Peces/genética , Genes , Intrones , Laminas , Datos de Secuencia Molecular , Proteínas Nucleares/química , Fragmentos de Péptidos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Alineación de Secuencia , Homología de Secuencia de AminoácidoRESUMEN
For any attempt to understand the biology of an organism the incorporation of a cDNA-based approach is unavoidable, because it is a major approach to studying gene function. The complete sequence of the genome alone is not sufficient to understand any organism; its gene regulation, expression, splice variation, posttranslational modifications, and protein-protein interactions all need to be addressed. Because the majority of vertebrate genes have probably been identified as ESTs the next stage of the Human Genome Project is attributing functional information to these sequences. In most cases hybridization-based approaches on arrayed pieces of DNA represent the most efficient way to study the expression level and splicing of a gene in a given tissue. Similar technology, now being applied at the protein level using protein expression libraries, high-density protein membranes, and antibody screening, should allow studies of protein localization and modifications. Coupled to these approaches is the use of technologies, which although lacking the highly parallel nature of hybridization, can potentially characterize large numbers of samples individually and with high accuracy. Automated gel-based DNA sequencing is an example of such a technique; protein sequencing and mass fingerprinting are further examples. In the case of mass spectroscopic analysis, the speed and sensitivity are vastly superior to that of gel-based approaches; however, the preparation of samples is more tedious. Our laboratory is developing a system to characterize DNA samples by mass spectrometry, allowing more rapid genotyping than is currently possible using gel-based technologies ([symbol: see text]. Gut, [symbol: see text]. Berlin and H. Lehrach, personal communication, 1998). Such technology would make information on gene polymorphisms widely accessible. Data generated using all of these techniques at the DNA and protein level will be connected by both protein expression libraries and database comparisons; finally, two hybrid library screens will identify many of the protein-protein interactions, linking genes together. In this way we will start to understand the interplay between genes on a global scale, both at the level of molecular interaction and the biological processes they regulate.
Asunto(s)
ADN Complementario , Biblioteca de Genes , ARN Mensajero/genética , Empalme Alternativo , Mapeo Cromosómico/métodos , ADN/química , ADN/genética , Dermatoglifia del ADN/instrumentación , Dermatoglifia del ADN/métodos , Regulación de la Expresión Génica , Variación Genética , Proyecto Genoma Humano , Humanos , Espectrometría de Masas/métodos , Robótica/instrumentación , Robótica/métodosRESUMEN
To gain an insight into vertebrate genome evolution, we have analysed the organization of an approximately 40-kb genomic clone of an amphioxus (Branchiostoma floridae) cosmid library. Amphioxus is considered as being the last non-vertebrate relative to vertebrates. Sequencing and analysis of the above clone using three different exon prediction programs (Grail, GenScan, Mzef) have led to the identification of a gene of the aldo-keto reductase family as well as further exons that gave a significant database match to known genes.
Asunto(s)
Oxidorreductasas de Alcohol/genética , Cordados no Vertebrados/genética , Aldehído Reductasa , Aldo-Ceto Reductasas , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Mapeo Cromosómico , Clonación Molecular , Cósmidos/genética , Datos de Secuencia Molecular , Secuencias Repetitivas de Ácidos Nucleicos , Alineación de SecuenciaRESUMEN
The paper presents a theoretical background for the analysis of the relationship between fertility and a number of socioeconomic factors associated with the process of economic development and analyses empirically this relationship within a cross-country framework. Fertility is found to be negatively related with female education, urbanization, and family planning and positively related with the levels of infant mortality and economic development, whereas no significant relationship between fertility and female labor force participation is established. Sensitivity analysis is performed and the policy implications of the empirical findings are briefly discussed.
Asunto(s)
Países en Desarrollo , Economía , Fertilidad , Modelos Teóricos , Factores Socioeconómicos , Demografía , Población , Dinámica Poblacional , InvestigaciónRESUMEN
Amphioxus AmphiBMP2/4 appears to be a single gene closely related to vertebrate BMP2 and BMP4. In amphioxus embryos, the expression patterns of AmphiBMP2/4 suggest patterning roles in the ectodermal dorsoventral axis (comparable to dorsoventral axis establishment in the ectoderm by Drosophila decapentaplegic and vertebrate BMP4). In addition AmphiBMP2/4 may be involved in somite evagination, tail bud growth, pharyngeal differentiation (resulting in club-shaped gland morphogenesis), hindgut regionalization, differentiation of olfactory epithelium, patterning of the anterior central nervous system, and establishment of the heart primordium. One difference between the developmental role of amphioxus AmphiBMP2/4 and vertebrate BMP4 is that the former does not appear to be involved in the initial establishment of the dorsoventral polarity of the mesoderm.
Asunto(s)
Proteínas Morfogenéticas Óseas/genética , Proteínas de Drosophila , Proteínas de Insectos/genética , Factor de Crecimiento Transformador beta , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Evolución Biológica , Southern Blotting , Proteína Morfogenética Ósea 2 , Proteína Morfogenética Ósea 4 , Cordados no Vertebrados/embriología , Cordados no Vertebrados/genética , ADN Complementario , Drosophila , Regulación del Desarrollo de la Expresión Génica , Ratones , Datos de Secuencia Molecular , Filogenia , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , VertebradosRESUMEN
Tob is a member of the PC3/ BTG1/Tob family of vertebrate tumor suppressor genes; its expression is known to inhibit proliferation of cells in vitro, but its possible roles during normal development have not been investigated previously. The present study concerns the structure and developmental expression of AmphiTob in an invertebrate chordate, amphioxus. This is the first investigation of any Tob gene during embryological development. The 311 amino acid AmphiTob protein is similar to vertebrate Tob but lacks the C-terminal PQ-rich domain of the latter. In early embryos of amphioxus, in situ hybridization first reveals AmphiTob expression in the hypoblast at the gastrula stage on the likely dorsal side of the embryo. During subsequent development, expression is seen in several tissues of the ectoderm, mesoderm, and endoderm. The most striking expression domains are in the developing somitic musculature and dorsal nerve cord. In the medial wall of each somite, AmphiTob is expressed strongly by cells destined to differentiate into the axial trunk muscles; this pattern persists until late in the larval stage, evidently because undifferentiated cells are continually becoming myogenic as the muscles grow. Nerve cord cells conspicuously transcribe AmphiTob from the late neurula until the early larval stage: Expression occurs in a few cells scattered along the nerve cord and in a group of cells located in the cerebral vesicle (in a region presumably homologous to the vertebrate diencephalic forebrain). During development, an intense and transitory transcription of AmphiTob may be an early event in cells exiting the cell cycle in preparation for differentiation.