RESUMEN
Genetic isolation by distance (IBD) has rarely been described in marine species with high potential for dispersal at both the larval and adult life-history stages. Here, we report significant relationships between inferred levels of gene flow and geographic distance in the Atlantic cod, Gadus morhua, at 10 nuclear restriction-fragment-length-polymorphism (RFLP) loci at small regional scales in the western north Atlantic region (< 1,600 km) that mirror those previously detected over its entire geographic range (up to 7,300 km). Highly significant allele frequency differences were observed among eight northwestern Atlantic populations, although the mean FST for all 10 loci was only 0.014. Despite this weak population structuring, the distance separating populations explained between 54% and 62% of the variation in gene flow depending on whether nine or 10 loci were used to estimate Nm. Across the species' entire geographic range, highly significant differences were observed among six regional populations at nine of the 10 loci (mean FST = 0.068) and seven loci exhibited significant negative relationships between gene flow and distance. At this large geographic scale, natural selection acting in the vicinity of one RFLP locus (GM798) had a significant effect on the correlation between gene flow and distance, and eliminating it from the analysis caused the coefficient of determination to increase from 17% to 62%. The role of vicariance was assessed by sequentially removing populations from the analysis and was found to play a minor role in contributing to the relationship between gene flow and distance at either geographic scale. The correlation between gene flow and distance detected in G. morhua at small and large spatial scales suggests that dispersal distances and effective population sizes are much smaller than predicted for the species and that the recent age of populations, rather than extensive gene flow, may be responsible for its weak population structure. Our results suggest that interpreting limited genetic differences among populations as reflecting high levels of ongoing gene flow should be made with caution.
Asunto(s)
Peces/genética , Genética de Población , Polimorfismo de Longitud del Fragmento de Restricción , Animales , Femenino , Geografía , Larva/crecimiento & desarrollo , Masculino , Dinámica PoblacionalRESUMEN
Specimens of Nautilus pompilius were trapped at depths of 225-300 m off the sunken barrier reef southeast of Port Moresby, Papua New Guinea. Animals transported to the Motupore Island laboratory were acclimated to normal habitat temperatures of 18 degrees C and then cannulated for arterial and venous blood sampling. When animals were forced to undergo a period of progressive hypoxia eventually to encounter ambient partial pressure of oxygen (PO2) levels of approximately 10 mmHg (and corresponding arterial PO2's of approximately 5 mmHg), they responded by lowering their aerobic metabolic rates to 5-10% of those seen in resting normoxic animals. Coincident with this profound metabolic suppression was an overall decrease in activity, with brief periods of jet propulsion punctuating long periods of rest. Below ambient PO2 levels of 30-40 mmHg, ventilatory movements became highly periodic and at the lowest PO2 levels encountered, ventilation occasionally ceased altogether. Cardiac output estimated by the Fick equation decreased during progressive hypoxia by as much as 75 80%, and in the deepest hypometabolic states heart rates slowed to one to two cycles of very low amplitude per minute. By the end of 500 min exposure to ambient PO2 levels of 10 mmHg or less, the anaerobic end products octopine and succinate had increased significantly in adductor muscle and heart, respectively. Increased concentrations of octopine in adductor muscle apparently contributed to a small intracellular acidosis and to the development of a combined respiratory and metabolic acidosis in the extracellular compartment. On the other hand, increases in succinate in heart muscle occurred in the absence of any change in cardiac pHi. Taken together, we estimate that these anaerobic end products would make up less than 2% of the energy deficit arising from the decrease in aerobic metabolism. Thus, metabolic suppression is combined with a massive downregulation of systemic O2 delivery to match metabolic supply to demand.
Asunto(s)
Adaptación Fisiológica/fisiología , Arginina/análogos & derivados , Metabolismo Basal/fisiología , Hipoxia/metabolismo , Moluscos/metabolismo , Equilibrio Ácido-Base/fisiología , Animales , Arginina/metabolismo , Dióxido de Carbono/metabolismo , Frecuencia Cardíaca , Concentración de Iones de Hidrógeno , Músculos/metabolismo , Miocardio/metabolismo , Oxígeno/metabolismo , Respiración , Ácido Succínico/metabolismoRESUMEN
Sp1-like transcription factors are characterized by three highly homologous C-terminal zinc finger motifs that bind GC-rich sequences. These proteins behave as either activators or repressors and have begun to be classified into different subfamilies based upon the presence of conserved motifs outside the zinc finger domain. This classification predicts that different Sp1-like subfamilies share certain functional properties. TIEG1 and TIEG2 constitute a new subfamily of transforming growth factor-beta-inducible Sp1-like proteins whose zinc finger motifs also bind GC-rich sequences. However, regions outside of the DNA-binding domain that differ in structure from other Sp1-like family members remain poorly characterized. Here, we have used extensive mutagenesis and GAL4-based transcriptional assays to identify three repression domains within TIEG1 and TIEG2 that we call R1, R2, and R3. R1 is 10 amino acids, R2 is 12 amino acids, and R3 is approximately 80 amino acids long. None of these domains share homology with previously described transcriptional regulatory motifs, but they share strong sequence homology and are functionally conserved between TIEG1 and TIEG2. Together, these data demonstrate that TIEG proteins are capable of repressing transcription, define domains critical for this function, and further support the idea that different subfamilies of Sp1-like proteins have evolved to mediate distinct transcriptional functions.
Asunto(s)
Proteínas de Unión al ADN/química , Proteínas Represoras/genética , Proteínas de Saccharomyces cerevisiae , Factor de Transcripción Sp1/química , Factores de Transcripción/química , Dedos de Zinc/genética , Secuencia de Aminoácidos , Animales , Células CHO , Mapeo Cromosómico , Secuencia Conservada , Cricetinae , Proteínas de Unión al ADN/genética , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Regulación de la Expresión Génica , Silenciador del Gen , Genes Reporteros , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Homología de Secuencia de Aminoácido , Factor de Transcripción Sp1/genética , Factores de Transcripción/genética , TransfecciónRESUMEN
Sp1-like zinc finger transcription factors are involved in the regulation of cell growth and differentiation. Recent evidence demonstrating that mammalian cells express novel, yet uncharacterized, Sp1-like proteins has stimulated a search for new members of this family. We and others have recently reported that the transforming growth factor (TGF)-beta-regulated gene TIEG encodes a new Sp1-like protein that inhibits cell growth in cultured cells. Here we report the identification, nuclear localization, DNA binding activity, transcriptional repression activity, and growth inhibitory effects of TIEG2, a novel TGF-beta-inducible gene related to TIEG. TIEG2 is ubiquitously expressed in human tissues, with an enrichment in pancreas and muscle. TIEG2 shares 91% homology with TIEG1 within the zinc finger region and 44% homology within the N terminus. Biochemical characterization reveals that TIEG2 is a nuclear protein, which, as predicted from the primary structure, specifically binds to an Sp1-like DNA sequence in vitro and can repress a promoter containing Sp1-like binding sites in transfected Chinese hamster ovary epithelial cells. Furthermore, functional studies using [3H]thymidine uptake and MTS (3-(4, 3-dimethyltiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-su lfophenyl)-2 H-tetrazolium) assays demonstrate that the overexpression of TIEG2 in Chinese hamster ovary cells inhibits cell proliferation. Thus, TIEG2, together with TIEG1, defines a new subfamily of TGF-beta-inducible Sp1-like proteins involved in the regulation of cell growth.
Asunto(s)
Proteínas de Ciclo Celular/química , División Celular/fisiología , Proteínas de Unión al ADN/química , Proteínas Represoras/química , Factores de Transcripción/química , Factor de Crecimiento Transformador beta/fisiología , Dedos de Zinc/fisiología , Secuencia de Aminoácidos , Animales , Apoptosis/genética , Proteínas Reguladoras de la Apoptosis , Células CHO/citología , Clonación Molecular , Cricetinae , Factores de Transcripción de la Respuesta de Crecimiento Precoz , Regulación de la Expresión Génica/genética , Humanos , Factores de Transcripción de Tipo Kruppel , Microscopía Fluorescente , Datos de Secuencia Molecular , Proteínas Nucleares/química , ARN Mensajero/metabolismo , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Factor de Transcripción Sp1/químicaRESUMEN
The authors conducted a survey of dentists reporting tooth avulsions from basketball net accidents. Although the number of people injured was small, the dental injuries were serious. In many cases, multiple teeth were avulsed as a result of the maxillary anterior teeth becoming entangled in the basketball net while the patients were attempting to slamdunk a basketball either on a lowered backboard or from a raised take-off area. The authors present recommendations for preventing tooth avulsions resulting from basketball net entanglement.
Asunto(s)
Baloncesto/lesiones , Avulsión de Diente/etiología , Accidentes Domésticos/prevención & control , Adolescente , Adulto , Traumatismos en Atletas/economía , Traumatismos en Atletas/prevención & control , Niño , Restauración Dental Permanente/economía , Femenino , Humanos , Incidencia , Incisivo/lesiones , Masculino , Maxilar , Protectores Bucales , Factores de Riesgo , Tratamiento del Conducto Radicular/economía , Avulsión de Diente/economía , Avulsión de Diente/prevención & control , Avulsión de Diente/cirugía , Pérdida de Diente/etiología , Reimplante Dental/economíaRESUMEN
Members of the dynamin superfamily are GTPases which have been shown to support receptor-mediated endocytosis in vivo and bind to growth factor receptor-associated proteins in vitro. In acinar cells of the pancreas, receptor-mediated endocytosis is very important for the recycling of membranes after secretory granule release. Therefore, characterization of the molecular machinery responsible for this process is critical for a better understanding of this phenomenon. In this study we sought to determine the expression pattern of the endocytic GTPase dynamin II during pancreatic acinar cell differentiation in developing rat embryos and in dexamethasone-treated AR42J cells using Western blot, Northern blot, and immunocytochemical analyses. During pancreatic development, dynamin immunoreactivity is almost undetectable until day E17 but undergoes significant upregulation in acinar cells starting at E18. In addition, the levels of dynamin mRNA and protein in AR42J cells increase approximately threefold during dexamethasone-induced acinar differentiation. The increase in dynamin levels that occurs in both embryonic pancreatic cells and dexamethasone-treated AR42J cells correlates with the establishment of a more differentiated acinar phenotype. Therefore, these results suggest a potential role for dynamin in supporting receptor-mediated endocytosis in mature pancreatic acinar cells.
Asunto(s)
GTP Fosfohidrolasas/genética , Páncreas/inmunología , Regulación hacia Arriba , Animales , Diferenciación Celular/efectos de los fármacos , Línea Celular , Dexametasona/farmacología , Dinaminas , GTP Fosfohidrolasas/metabolismo , Páncreas/citología , Fenotipo , ARN Mensajero/genética , RatasRESUMEN
Dynamin proteins are members of a recently described family of GTPases involved in receptor-mediated processes. To date, three different dynamin-encoding genes have been identified in mammalian tissues. Dynamin I is expressed only in neurons, whereas dynamin II is ubiquitously expressed. A third isoform, dynamin III, was originally isolated from a rat testis cDNA library and shown to be testis-specific. However, here we report the cloning and characterization of dynamin III from brain and lung, demonstrating a more extended pattern of expression for this isoform. In addition, we have investigated the temporal pattern of expression of these three genes during brain development. We find that both dynamin I and dynamin III mRNA levels are up-regulated during embryogenesis, whereas dynamin II mRNA levels remain unchanged. From these results, we conclude that dynamin III is not a testis-specific isoform and, furthermore, that rat brain expresses three different dynamin-encoding genes that are differentially regulated during development. Therefore, this large isoform diversity of dynamin proteins in brain predicts a significant complexity in the understanding of dynamin-based processes in this tissue.
Asunto(s)
Encéfalo/embriología , GTP Fosfohidrolasas/genética , Empalme Alternativo/fisiología , Secuencia de Aminoácidos , Animales , Encéfalo/fisiología , Dinamina I , Dinamina III , Dinaminas , Endocitosis/genética , GTP Fosfohidrolasas/química , Regulación del Desarrollo de la Expresión Génica/fisiología , Biblioteca de Genes , Pruebas Genéticas , Isoenzimas/química , Isoenzimas/genética , Pulmón/fisiología , Masculino , Microtúbulos/química , Microtúbulos/genética , Datos de Secuencia Molecular , Células PC12/fisiología , ARN Mensajero/análisis , Ratas , Ratas Sprague-Dawley , Testículo/fisiología , Factores de TiempoRESUMEN
Genes encoding for C2H2 zinc finger proteins are known to regulate normal cell proliferation and differentiation and have often been found to be mutated in different forms of cancer. We are interested in understanding the role of these genes as regulators of cell proliferation and differentiation in the exocrine pancreas. Therefore, we have generated expressed sequence tags (ESTs) encoding pancreas-enriched zinc finger peptides using the polymerase chain reaction and hybridization techniques [Adams, M.D. et al. (1991) Science, 252, 1651-1656]. Here we report the primary structure and expression pattern of 18 different zinc finger-encoding cDNAs (DZF-1-18) from the azaserine-derived tumoral cell line AR4IP which displays a poorly differentiated phenotype. Sequence analysis shows that all of these clones encode peptides which share the consensus DNA-binding motif with the Drosophila zinc finger transcription factor krüppel. High stringency Northern blot analysis shows that eight different zinc finger transcripts are expressed at high levels in normal adult rat pancreas and therefore constitute good candidates to play a role as transcription factors in exocrine pancreatic cells.
Asunto(s)
ADN Complementario/aislamiento & purificación , Regulación de la Expresión Génica/genética , Páncreas/química , Lugares Marcados de Secuencia , Dedos de Zinc/genética , Animales , Secuencia de Bases , Northern Blotting , Células Cultivadas , Datos de Secuencia Molecular , Péptidos/química , ARN/química , ARN/aislamiento & purificación , Ratas , Factores de Transcripción/análisisRESUMEN
Zinc finger transcription factors are DNA-binding proteins that are known to determine the identity of cells by regulating cell-specific gene expression. In addition, mutations in some members of this family have been found to be associated with several neoplasias, including Wilms' tumor and leukemias. Because the mechanisms that regulate normal, as well as neoplastic, pancreatic cell differentiation are poorly understood, we are searching for pancreas-enriched transcription factors that may determine the identity of pancreatic cells. Towards this end, we have used the polymerase chain reaction and degenerate primers against the highly conserved (Cys2-His2) zinc finger domain to amplify novel transcription factor encoding cDNAs from the well-characterized pancreatic acinar cell line AR42J. Using this approach, we have identified 17 different zinc finger encoding cDNAs (AZF-1 to -17). Sequence analysis shows that all of these clones encode for different zinc finger peptides which share the consensus DNA binding motif with the Drosophila transcription factor krüppel. As a first step in the characterization of these genes, the purified PCR products were used to determine their spatial pattern of expression by northern blot analysis. Using these techniques, we find that numerous zinc finger encoding genes are expressed in AR42J acinar cells as well as in normal adult rat pancreas and suggest that they may play a role as transcription factors in exocrine pancreatic cells.
Asunto(s)
Neoplasias Pancreáticas/genética , Factores de Edad , Secuencia de Aminoácidos , Animales , Diferenciación Celular , ADN Complementario/genética , ADN de Neoplasias/genética , Datos de Secuencia Molecular , ARN Neoplásico/genética , Ratas , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Distribución Tisular , Células Tumorales Cultivadas , Dedos de ZincRESUMEN
High levels of gene flow have been implicated in producing uniform patterns of allozyme variation among populations of many marine fish species. We have examined whether gene flow is responsible for the limited population structure in the Atlantic cod, Gadus morhua L., by comparing the previously published patterns of variation at 10 allozyme loci to 17 nuclear restriction fragment length polymorphism (RFLP) loci scored by 11 anonymous cDNA clones. Unlike the allozyme loci, highly significant differences were observed among all populations at the DNA markers in a pattern consistent with an isolation-by-distance model of population structure. The magnitude of allele frequency variation at the nuclear RFLP loci significantly exceeded that observed at the protein loci (chi 2 = 24.6, d.f. = 5, P < 0.001). Estimates of gene flow from the private alleles method were similar for the allozymes and nuclear RFLPs. From the infinite island model, however, estimates of gene flow from the DNA markers were fivefold lower than indicated by the proteins. The discrepancy between gene flow estimates, combined with the observation of a large excess of rare RFLP alleles, suggests that the Atlantic cod has undergone a recent expansion in population size and that populations are significantly displaced from equilibrium. Because gene flow is a process that affects all loci equally, the heterogeneity observed among populations at the DNA level eliminates gene flow as the explanation for the homogeneous allozyme patterns. Our results suggest that a recent origin of cod populations has acted to constrain the extent of population differentiation observed at weakly polymorphic loci and implicate a role for selection in affecting the distribution of protein variation among natural populations in this species.