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2.
Proc Natl Acad Sci U S A ; 97(17): 9514-8, 2000 Aug 15.
Artículo en Inglés | MEDLINE | ID: mdl-10920195

RESUMEN

Results of a first-stage Sea Urchin Genome Project are summarized here. The species chosen was Strongylocentrotus purpuratus, a research model of major importance in developmental and molecular biology. A virtual map of the genome was constructed by sequencing the ends of 76,020 bacterial artificial chromosome (BAC) recombinants (average length, 125 kb). The BAC-end sequence tag connectors (STCs) occur an average of 10 kb apart, and, together with restriction digest patterns recorded for the same BAC clones, they provide immediate access to contigs of several hundred kilobases surrounding any gene of interest. The STCs survey >5% of the genome and provide the estimate that this genome contains approximately 27,350 protein-coding genes. The frequency distribution and canonical sequences of all middle and highly repetitive sequence families in the genome were obtained from the STCs as well. The 500-kb Hox gene complex of this species is being sequenced in its entirety. In addition, arrayed cDNA libraries of >10(5) clones each were constructed from every major stage of embryogenesis, several individual cell types, and adult tissues and are available to the community. The accumulated STC data and an expanding expressed sequence tag database (at present including >12, 000 sequences) have been reported to GenBank and are accessible on public web sites.


Asunto(s)
Genoma , Mapeo Físico de Cromosoma , Erizos de Mar/genética , Envejecimiento/genética , Animales , Clonación Molecular , Mapeo Contig , ADN Complementario/genética , Bases de Datos Factuales , Etiquetas de Secuencia Expresada , Regulación del Desarrollo de la Expresión Génica , Genes Homeobox/genética , Internet , Datos de Secuencia Molecular , Familia de Multigenes/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Secuencias Repetitivas de Ácidos Nucleicos/genética , Erizos de Mar/citología , Erizos de Mar/embriología
4.
Development ; 126(17): 3857-67, 1999 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-10433914

RESUMEN

A set of 956 expressed sequence tags derived from 7-hour (mid-cleavage) sea urchin embryos was analyzed to assess biosynthetic functions and to illuminate the structure of the message population at this stage. About a quarter of the expressed sequence tags represented repetitive sequence transcripts typical of early embryos, or ribosomal and mitochondrial RNAs, while a majority of the remainder contained significant open reading frames. A total of 232 sequences, including 153 different proteins, produced significant matches when compared against GenBank. The majority of these identified sequences represented 'housekeeping' proteins, i.e., cytoskeletal proteins, metabolic enzymes, transporters and proteins involved in cell division. The most interesting finds were components of signaling systems and transcription factors not previously reported in early sea urchin embryos, including components of Notch and TGF signal transduction pathways. As expected from earlier kinetic analyses of the embryo mRNA populations, no very prevalent protein-coding species were encountered; the most highly represented such sequences were cDNAs encoding cyclins A and B. The frequency of occurrence of all sequences within the database was used to construct a sequence prevalence distribution. The result, confirming earlier mRNA population analyses, indicated that the poly(A) RNA of the early embryo consists mainly of a very complex set of low-copy-number transcripts.


Asunto(s)
Etiquetas de Secuencia Expresada , Erizos de Mar/embriología , Erizos de Mar/genética , Animales , Secuencia de Bases , Fase de Segmentación del Huevo/metabolismo , ADN Complementario/genética , Regulación del Desarrollo de la Expresión Génica , Humanos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Erizos de Mar/metabolismo , Homología de Secuencia de Ácido Nucleico
5.
Dev Growth Differ ; 41(3): 303-12, 1999 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-10400392

RESUMEN

The genomic DNA region that contains the SM50 gene also included a second gene that appeared to encode another skeletogenic matrix protein. The two genes were linked at a distance of about 12 kb. Based on the molecular weight of the implied protein the gene was termed SM37. The SM37 protein included a long tandem sequence of short glycine-rich repeats that was similar to the glycine-rich repeats included in the SM50 protein and several other skeletal matrix proteins, although the sequence of SM37 repeat was distinct. The overall structure of the SM37 protein was similar to SM50 as well. However, SM37 was only about 30% identical in amino acid sequence to SM50, and coding region probes behaved as a single copy sequence under standard conditions. The SM37 gene included the same cis-regulatory elements as the SM50 gene, although in a different order with respect to transcription. This gene was regulated coordinately with SM50 during development, and like SM50 was expressed exclusively in skeletogenic mesenchyme lineages.


Asunto(s)
Huesos/embriología , Proteínas del Citoesqueleto/genética , Proteínas de la Matriz Extracelular , Proteínas/genética , Erizos de Mar/embriología , Secuencia de Aminoácidos , Animales , Secuencia de Bases , ADN Complementario , Regulación del Desarrollo de la Expresión Génica , Ligamiento Genético , Datos de Secuencia Molecular , Homología de Secuencia de Aminoácido
6.
Dev Biol ; 208(2): 255-64, 1999 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-10191043

RESUMEN

Strongylocentrotus purpuratus, a major research model in developmental molecular biology, has been inbred through six generations of sibling matings. Though viability initially decreased, as described earlier, the inbred line now consists of healthy, fertile animals. These are intended to serve as a genomic resource in which the level of polymorphism is decreased with respect to wild S. purpuratus. To genotype the inbred animals eight simple sequence genomic repeats were isolated, in context, and PCR primers were generated against the flanking single-copy sequences. Distribution and polymorphism of these regions of the genome were studied in the genomes of 27 wild individuals and in a sample of the inbred animals at F2 and F3 generations. All eight regions were polymorphic, though to different extents, and their homozygosity was increased by inbreeding as expected. The eight markers suffice to identify unambiguously the cellular DNA of any wild or F3 S. purpuratus individual.


Asunto(s)
Animales de Laboratorio/genética , Repeticiones de Microsatélite , Erizos de Mar/genética , Animales , Secuencia de Bases , Cruzamientos Genéticos , Femenino , Frecuencia de los Genes , Biblioteca Genómica , Genotipo , Heterocigoto , Homocigoto , Endogamia , Masculino , Datos de Secuencia Molecular , Sondas de Oligonucleótidos
7.
Proc Natl Acad Sci U S A ; 95(16): 9372-7, 1998 Aug 04.
Artículo en Inglés | MEDLINE | ID: mdl-9689087

RESUMEN

These 10 obvious propositions make a model of the specification of form, intended to expose underlying assumptions of developmental biology for examination and future experimentation. (I) The control of development is by means of local interactions, rather than global control mechanisms. (II) A macromolecule near a specific site will bind by mass action. (III) Starting with a precursor cell, all cells are assembled automatically by specifically binding macromolecules. (IV) At the surface of cells are specific adhesion sites that determine how all cells bind to each other. (V) An organism will assemble automatically from parts (macromolecules, structures, and cells) specified by nuclear control factors. (VI) The nuclear control factors in each cell are from precursor cells and factors derived by signaling from other cells. (VII) The macromolecules that determine specific binding, cell adhesion, and signaling are controlled by nuclear control factors, and in a grand feedback the cell adhesion and signaling systems determine the nuclear factor patterns. (VIII) The embryonic precursor cells for organs, termed "precursor groups," are linked by adhesion and signaling relationships. (IX) The precursor groups include precursors for regions of an organ and boundary cells between regions having few cell types, growing without additional specific cell-to-cell relationships. (X) Organs are held together by cell adhesion in functional relationships. Thus the form and function of the organism is specified entirely by local control mechanisms. Without global control systems, information for form is in the genes for structural proteins, adhesion molecules, control factors, signaling molecules, and their control regions.


Asunto(s)
Desarrollo Embrionario y Fetal , Modelos Biológicos , Animales , Humanos
8.
Proc Natl Acad Sci U S A ; 95(11): 5906-12, 1998 May 26.
Artículo en Inglés | MEDLINE | ID: mdl-9600891

RESUMEN

The terminal regions (last 20 kb) of Saccharomyces cerevisiae chromosomes universally contain blocks of precise sequence similarity to other chromosome terminal regions. The left and right terminal regions are distinct in the sense that the sequence similarities between them are reverse complements. Direct sequence similarity occurs between the left terminal regions and also between the right terminal regions, but not between any left ends and right ends. With minor exceptions the relationships range from 80% to 100% match within blocks. The regions of similarity are composites of familiar and unfamiliar repeated sequences as well as what could be considered "single-copy" (or better "two-copy") sequences. All terminal regions were compared with all other chromosomes, forward and reverse complement, and 768 comparisons are diagrammed. It appears there has been an extensive history of sequence exchange or copying between terminal regions. The subtelomeric sequences fall into two classes. Seventeen of the chromosome ends terminate with the Y' repeat, while 15 end with the 800-nt "X2" repeats just adjacent to the telomerase simple repeats. The just-subterminal repeats are very similar to each other except that chromosome 1 right end is more divergent.


Asunto(s)
Cromosomas Fúngicos , ADN Complementario , Modelos Moleculares , Modelos Teóricos , Saccharomyces cerevisiae/genética , Análisis de Secuencia de ADN , Secuencia de Bases , ADN de Hongos/genética , Datos de Secuencia Molecular , Secuencias Repetitivas de Ácidos Nucleicos
9.
Proc Natl Acad Sci U S A ; 95(7): 3351-5, 1998 Mar 31.
Artículo en Inglés | MEDLINE | ID: mdl-9520368

RESUMEN

In this model, without dark matter, the flat rotation curves of galaxies and the mass-to-light ratios of clusters of galaxies are described quantitatively. The hypothesis is that the agent of gravitational force is propagated as if it were scattered with a mean free path of approximately 5 kiloparsecs. As a result, the force between moderately distant masses, separated by more than the mean free path, diminishes as the inverse first power of the distance, following diffusion equations, and describes the flat rotation curves of galaxies. The force between masses separated by <1 kiloparsec diminishes as the inverse square of distance. The excess gravitational force (ratio of 1/r:1/r2) increases with the scale of structures from galaxies to clusters of galaxies. However, there is reduced force at great distances because of the approximately 12 billion years that has been available for diffusion to occur. This model with a mean free path of approximately 5 kiloparsecs predicts a maximum excess force of a few hundredfold for objects the size of galactic clusters a few megaparsecs in size. With only a single free parameter, the predicted curve for excess gravitational force vs. size of structures fits reasonably well with observations from those for dwarf galaxies through galactic clusters. Under the diffusion model, no matter is proposed in addition to the observed baryons plus radiation and thus the proposed density of the universe is only a few percent of that required for closure.

10.
Gene ; 205(1-2): 177-82, 1997 Dec 31.
Artículo en Inglés | MEDLINE | ID: mdl-9461392

RESUMEN

Current evidence on the long-term evolutionary effect of insertion of sequence elements is reviewed. There are three criteria for inclusion of an example: (i) the element was inserted far in the past and thus the event is not a transient mutation; (ii) the element is a member of a large group of similar sequences; (iii) the element now serves a useful function. There are 21 examples from Drosophila, sea urchin, human and mouse genomes that meet these criteria. Taken together, these examples show that the insertion of sequence elements in the genome has been a significant source of regulatory variation in evolution.


Asunto(s)
Elementos Transponibles de ADN/genética , Evolución Molecular , Animales , Regulación de la Expresión Génica , Humanos , Retroelementos , Retroviridae/genética , Transcripción Genética
11.
Proc Natl Acad Sci U S A ; 93(18): 9374-7, 1996 Sep 03.
Artículo en Inglés | MEDLINE | ID: mdl-8790336

RESUMEN

Current evidence on the long-term evolutionary effect of insertion of sequence elements into gene regions is reviewed, restricted to cases where a sequence derived from a past insertion participates in the regulation of expression of a useful gene. Ten such examples in eukaryotes demonstrate that segments of repetitive DNA or mobile elements have been inserted in the past in gene regions, have been preserved, sometimes modified by selection, and now affect control of transcription of the adjacent gene. Included are only examples in which transcription control was modified by the insert. Several cases in which merely transcription initiation occurred in the insert were set aside. Two of the examples involved the long terminal repeats of mammalian endogenous retroviruses. Another two examples were control of transcription by repeated sequence inserts in sea urchin genomes. There are now six published examples in which Alu sequences were inserted long ago into human gene regions, were modified, and now are central in control/enhancement of transcription. The number of published examples of Alu sequences affecting gene control has grown threefold in the last year and is likely to continue growing. Taken together, all of these examples show that the insertion of sequence elements in the genome has been a significant source of regulatory variation in evolution.


Asunto(s)
Elementos Transponibles de ADN , Evolución Molecular , Regulación del Desarrollo de la Expresión Génica , Animales , Variación Genética , Humanos , Secuencias Repetitivas de Ácidos Nucleicos
12.
Mol Phylogenet Evol ; 5(1): 13-7, 1996 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-8673282

RESUMEN

There is no doubt that mobile elements are a major source of mutation and that in many cases insertions cause changes in the expression of genes, but a question remains open. What has the long-term effect of these processes been? The data collected here show that in many eukaryotes, segments of repetitive DNA (mobile elements) that have been inserted in the past in regions of many eukaryote genes have been preserved by selection and now affect the transcriptional control of these specific genes. At least five of the examples are considered to be solid cases demonstrating this history. There are a number of significant but less compelling sets of evidence that support the concept that the insertion of mobile elements in gene regions could be a major source of regulatory variation in evolution.


Asunto(s)
Elementos Transponibles de ADN , Evolución Molecular , Regulación de la Expresión Génica , Proteínas Represoras , Amilasas/genética , Animales , Secuencia de Bases , Sitios de Unión/genética , Proteínas Sanguíneas/genética , Antígenos CD8/genética , Complemento C4 , Secuencia Conservada , ADN/genética , ADN/metabolismo , Proteínas de Unión al ADN/genética , Femenino , Genes de Plantas , Humanos , Factores de Transcripción de Tipo Kruppel , Masculino , Ratones , Mutagénesis Insercional , Secuencias Repetitivas de Ácidos Nucleicos , Erizos de Mar , Factores de Transcripción/genética
13.
J Immunol ; 156(2): 593-602, 1996 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-8543810

RESUMEN

To identify some of the genes expressed in LPS-activated coelomocytes, we sequenced randomly chosen clones from a directionally constructed cDNA library to produce a set of expressed sequence tags (ESTs). Deduced amino acid sequences from 307 ESTs were compared with known protein sequences in GenBank, and significant matches to approximately 30% of the clones were identified. Eighty-nine clones matched to 55 different proteins, including several putative immune effector proteins. In this work, we show the first identification of an invertebrate homologue of a vertebrate C component. Another EST matches to several short consensus repeats that are characteristic of a variety of proteins, including CR/regulatory proteins and clotting factors. Additional putative immune effector genes include 1) a Kazal-type protease inhibitor that may function to inactivate bacterial proteases, 2) a C-type lectin similar to echinoidin, and 3) a serine protease with similarities to thrombin, elastase, haptoglobin, and plasmin. Other EST categories include 1) cell surface proteins and receptors, 2) proteins involved in signaling systems, 3) lysosomal and secreted proteins, 4) cytoskeletal and cytoskeletal modifying proteins, 5) general cell function proteins, 6) proteins with unknown function, and 7) ESTs without significant matches, 25 with open reading frames. Many of the ESTs identified in this study represent the types of genes expected to be used in lower deuterostome immune functions.


Asunto(s)
Proteínas del Sistema Complemento/genética , Equinodermos/inmunología , Regulación del Desarrollo de la Expresión Génica , Genes MHC Clase II , Genes , Fagocitos/metabolismo , Erizos de Mar/genética , Secuencia de Aminoácidos , Grupos de Población Animal/genética , Animales , Secuencia de Bases , Secuencia de Consenso , ADN Complementario/genética , Equinodermos/genética , Evolución Molecular , Hongos/genética , Expresión Génica , Lipopolisacáridos/farmacología , Datos de Secuencia Molecular , Sistemas de Lectura Abierta , Fagocitos/efectos de los fármacos , Plantas/genética , Proteínas/clasificación , Proteínas/genética , Secuencias Repetitivas de Ácidos Nucleicos , Erizos de Mar/citología , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Especificidad de la Especie
14.
Development ; 121(7): 1957-70, 1995 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-7635044

RESUMEN

The SM50 gene encodes a minor matrix protein of the sea urchin embryo spicule. We carried out a detailed functional analysis of a cis-regulatory region of this gene, extending 440 bp upstream and 120 bp downstream of the transcription start site, that had been shown earlier to confer accurate skeletogenic expression of an injected expression vector. The distal portion of this fragment contains elements controlling amplitude of expression, while the region from -200 to +105 contains spatial control elements that position expression accurately in the skeletogenic lineages of the embryo. A systematic mutagenesis analysis of this region revealed four adjacent regulatory elements, viz two copies of a positively acting sequence (element D) that are positioned just upstream of the transcription start site; an indispensable spatial control element (element C) that is positioned downstream of the start site; and further downstream, a second positively acting sequence (element A). We then constructed a series of synthetic expression constructs. These contained oligonucleotides representing normal and mutated versions of elements D, C, and A, in various combinations. We also changed the promoter of the SM50 gene from a TATA-less to a canonical TATA box form, without any effect on function. Perfect spatial regulation was also produced by a final series of constructs that consisted entirely of heterologous enhancers from the CyIIIa gene, the SV40 early promoter, and synthetic D, C, and A elements. We demonstrate that element C exercises the primary spatial control function of the region we analyzed. We term this a 'locator' element. This differs from conventional 'tissue-specific enhancers' in that while it is essential for expression, it has no transcriptional activity on its own, and it requires other, separable, positive regulatory elements for activity. In the normal configuration these ancillary positive functions are mediated by elements A and D. Only positively acting control elements were observed in the SM50 regulatory domain throughout this analysis.


Asunto(s)
Proteínas del Citoesqueleto/genética , Proteínas de la Matriz Extracelular , Glicoproteínas/genética , Mesodermo/fisiología , Secuencias Reguladoras de Ácidos Nucleicos , Erizos de Mar/embriología , Animales , Secuencia de Bases , Diferenciación Celular/genética , Cloranfenicol O-Acetiltransferasa/metabolismo , ADN , Expresión Génica , Marcadores Genéticos , Hibridación in Situ , Mesodermo/enzimología , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Erizos de Mar/enzimología , Erizos de Mar/genética
15.
Dev Biol ; 170(1): 75-82, 1995 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-7601316

RESUMEN

SpP3A1 and SpP3A2 are DNA-binding proteins that interact specifically with the same target sites in the regulatory domains of the Strongylocentrotus purpuratus CyIIIa gene and also of several other known genes. In this work we used antibodies raised against recombinant P3A1 and P3A2 to quantitate these transcription factors in eggs and in the nuclear compartments of embryos of various stages. Both proteins are present in unfertilized eggs, and both enter the embryonic nuclei early in development, but only P3A2 remains present in nuclei at functional concentrations beyond the early gastrula stage. Combined with earlier measurements of P3A site binding at cleavage stages, these measurements show that P3A1 would be replaced by P3A2 at target sites in genes regulated by these factors.


Asunto(s)
Proteínas de Unión al ADN/fisiología , ADN/metabolismo , Regulación del Desarrollo de la Expresión Génica , Erizos de Mar/genética , Factores de Transcripción/fisiología , Animales , Secuencia de Bases , Sitios de Unión , Núcleo Celular/química , Citoplasma/química , Proteínas de Unión al ADN/inmunología , Embrión no Mamífero/metabolismo , Femenino , Datos de Secuencia Molecular , Oocitos/metabolismo , Conejos , Erizos de Mar/embriología , Factores de Transcripción/inmunología , Dedos de Zinc
16.
Mol Mar Biol Biotechnol ; 4(2): 148-53, 1995 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-7773332

RESUMEN

SpZ12-1 is a zinc-finger transcription factor. Previous work has indicated that this factor functions late in embryogenesis as a spatial transcriptional repressor. We show here that this factor is present in significant quantities even in unfertilized egg cytoplasm, and in similar quantities in mesenchyme blastula-stage embryo cytoplasm. Taken together with earlier measurements of Calzone and associates, our observations indicate that SpZ12-1 enters the embryonic nuclei between late cleavage and mesenchyme blastula stages. A low-prevalence mRNA encoding SpZ12-1 is also present throughout development. Translation of this mRNA could, however, easily account for the complete complement of SpZ12-1 protein in the embryo, as estimated from its DNA binding activity. SpZ12-1 probably functions at several developmental stages and is evidently of both maternal and embryonic provenance.


Asunto(s)
Proteínas de Unión al ADN/análisis , ARN Mensajero/análisis , Proteínas Represoras , Erizos de Mar/embriología , Factores de Transcripción/análisis , Dedos de Zinc , Animales , Blastocisto/química , Compartimento Celular , Núcleo Celular/química , Citoplasma/química , Proteínas de Unión al ADN/genética , Femenino , Óvulo/química , Sondas ARN , ARN sin Sentido , Fracciones Subcelulares/química , Factores de Transcripción/genética
17.
Dev Biol ; 169(2): 713-27, 1995 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-7781910

RESUMEN

Several Strongylocentrotus purpuratus gene cis-regulatory regions contain asymmetric C4 sequences which are core elements of target sites for a specific DNA-protein interaction. Blastula stage nuclear extract contains five proteins which specifically bind to these target sites, resulting in a characteristic pattern of complexes in gel mobility shift assays. We used automated affinity chromatography to purify a protein which binds to these sites and have isolated the corresponding cDNA. This protein, SpGCF1, is a novel sea urchin DNA-binding protein with no overall homology to proteins reported in the databases currently available. The DNA-binding domain of this protein was identified by a deletion analysis. As demonstrated both for protein translated in vitro and for bacterial protein expressed from a cDNA clone, a single SpGCF1 mRNA serves as a template for the synthesis of five DNA-binding polypeptides. We show that these five polypeptides are most likely produced by differential usage of a nested set of AUG start codons in the SpGCF1 cDNA and thus contain variable amounts of a proline-rich N-terminal domain. Since proline-rich regions often serve as transcriptional activation domains, the five SpGCF1 proteins apparently possess different "activation potentials."


Asunto(s)
Proteínas de Unión al ADN/genética , ARN Mensajero/genética , Erizos de Mar/genética , Factores de Transcripción/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Sitios de Unión , Clonación Molecular , ADN/metabolismo , ADN Complementario , Regulación del Desarrollo de la Expresión Génica , Datos de Secuencia Molecular , Erizos de Mar/embriología
18.
Dev Comp Immunol ; 19(3): 217-24, 1995.
Artículo en Inglés | MEDLINE | ID: mdl-8595820

RESUMEN

Profilin is a small, actin-binding protein that functions at the intersection of signal transduction and cytoskeletal modifications. Increases in the number of profilin messages per cell correlate with sea urchin coelomocytes activation in response to injury. Here we show that coelomocytes respond to immune challenge from lipopolysaccharide with significant elevations in profilin transcripts per coelomyocyte.


Asunto(s)
Proteínas Contráctiles , Lipopolisacáridos/farmacología , Proteínas de Microfilamentos/genética , ARN Mensajero/análisis , Erizos de Mar/inmunología , Animales , Inmunidad Innata/efectos de los fármacos , Profilinas
19.
Development ; 121(4): 1111-22, 1995 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-7743924

RESUMEN

The CyIIIa cytoskeletal actin gene of the sea urchin Strongylocentrotus purpuratus is activated in late cleavage and expressed exclusively in the aboral ectoderm territory of the embryo. Previous gene transfer studies defined a 2.3 kb cis-regulatory region that is necessary and sufficient for correct temporal and spatial expression of a CyIIIa.CAT fusion gene. In this paper, a negative regulatory element within this region was identified that is required for repression of the CyIIIa gene in skeletogenic mesenchyme cells. The repression mediated by this regulatory element takes place after initial territorial specification. A cDNA clone encoding a DNA-binding protein with twelve Zn fingers (SpZ12-1) was isolated by probing an expression library with this cis-element. Deletion analysis of the SpZ12-1 protein confirmed that a DNA-binding domain is located within the Zn finger region. SpZ12-1 is the only DNA-binding protein in embryo nuclear extract that interacts with the specific cis-target sites required for repression of CyIIIa.CAT in skeletogenic mesenchyme and is likely to be the trans factor that mediates this repression.


Asunto(s)
Actinas/genética , Proteínas de Unión al ADN/genética , Regulación del Desarrollo de la Expresión Génica , Genes Reguladores , Mesodermo/fisiología , Proteínas Represoras , Erizos de Mar/genética , Factores de Transcripción/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Técnicas Genéticas , Hibridación in Situ , Datos de Secuencia Molecular , Sondas de Oligonucleótidos , Erizos de Mar/embriología , Dedos de Zinc
20.
Proc Natl Acad Sci U S A ; 92(7): 2989-93, 1995 Mar 28.
Artículo en Inglés | MEDLINE | ID: mdl-7708761

RESUMEN

SpGCF1 is a recently cloned sea urchin transcription factor that recognizes target sites in several different sea urchin genes. We find that in gel-shift experiments this factor is able to multimerize. A quantitative simulation of the gel-shift results suggests that SpGCF1 molecules that are bound to DNA target sites may also bind to one another, thus associating several DNA probe molecules. SpGCF1 might therefore be able to loop DNA molecules bearing its target sites at distant locations. We demonstrate this prediction by electron microscopy, and using the well-characterized cis-regulatory domain of the CyIIIa cytoskeletal actin gene, we show that the loop conformations predicted from the known SpGCF1 target site locations are actually formed in vitro. We speculate that the multimerization of this factor in vivo may function to bring distant regions of extended regulatory domains into immediate proximity so that they can interact with one another.


Asunto(s)
Proteínas de Unión al ADN/metabolismo , ADN/ultraestructura , Embrión no Mamífero/metabolismo , Erizos de Mar/embriología , Factores de Transcripción/metabolismo , Actinas/biosíntesis , Animales , Secuencia de Bases , Sitios de Unión , ADN/química , ADN/metabolismo , Sondas de ADN , Proteínas de Unión al ADN/biosíntesis , Cinética , Sustancias Macromoleculares , Microscopía Electrónica , Secuencias Reguladoras de Ácidos Nucleicos , Factores de Transcripción/biosíntesis
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