RESUMEN
Zebrafish are a valuable model for mammalian lipid metabolism; larvae process lipids similarly through the intestine and hepatobiliary system and respond to drugs that block cholesterol synthesis in humans. After ingestion of fluorescently quenched phospholipids, endogenous lipase activity and rapid transport of cleavage products results in intense gall bladder fluorescence. Genetic screening identifies zebrafish mutants, such as fat free, that show normal digestive organ morphology but severely reduced phospholipid and cholesterol processing. Thus, fluorescent lipids provide a sensitive readout of lipid metabolism and are a powerful tool for identifying genes that mediate vertebrate digestive physiology.
Asunto(s)
Fenómenos Fisiológicos del Sistema Digestivo , Sistema Digestivo/metabolismo , Colorantes Fluorescentes/metabolismo , Fosfolípidos/metabolismo , Pez Cebra/metabolismo , Animales , Anticolesterolemiantes/farmacología , Atorvastatina , Ácidos y Sales Biliares/farmacología , Compuestos de Boro/metabolismo , Colesterol/metabolismo , Sistema Digestivo/efectos de los fármacos , Sistema Digestivo/patología , Fenómenos Fisiológicos del Sistema Digestivo/efectos de los fármacos , Vesícula Biliar/efectos de los fármacos , Vesícula Biliar/metabolismo , Ácidos Heptanoicos/farmacología , Larva/efectos de los fármacos , Larva/metabolismo , Lipasa/metabolismo , Ratones , Microscopía Fluorescente , Microscopía por Video , Mutación/genética , Pirroles/farmacología , Transducción de Señal/efectos de los fármacos , Pez Cebra/embriología , Pez Cebra/genética , Pez Cebra/fisiologíaRESUMEN
The amphibian Spemann organizer is subdivided in trunk and head organizer and it is unclear how this division is regulated. The Xenopus trunk organizer expresses anti-dorsalizing morphogenetic protein (ADMP), a potent organizer antagonist. We show that ADMP represses head formation during gastrulation and that its expression is activated by BMP antagonists. A specifically acting dominant-negative ADMP anteriorizes embryos and its coexpression with BMP antagonists induces secondary embryonic axes with heads as well as expression of head inducers. Unlike other BMPs, ADMP is not inhibited by a dominant-negative BMP type I receptor, Noggin, Cerberus and Chordin but by Follistatin, suggesting that it utilizes a distinct TGF-beta receptor pathway and displays differential sensitivity to BMP antagonists. The results indicate that ADMP functions in the trunk organizer to antagonize head formation, thereby regulating organizer patterning.
Asunto(s)
Tipificación del Cuerpo/fisiología , Proteínas Morfogenéticas Óseas/metabolismo , Péptidos y Proteínas de Señalización Intercelular , Receptores de Factores de Crecimiento , Animales , Receptores de Proteínas Morfogenéticas Óseas , Proteínas Morfogenéticas Óseas/genética , Proteínas Morfogenéticas Óseas/fisiología , Proteínas Portadoras , Folistatina , Glicoproteínas/metabolismo , Cabeza/embriología , Péptidos y Proteínas de Señalización Intracelular , Morfogénesis , Proteínas/genética , Proteínas/metabolismo , Receptores de Superficie Celular/metabolismo , Transducción de Señal , Xenopus , Proteínas de Xenopus , Proteínas de Pez Cebra , CigotoRESUMEN
Members of the transforming growth factor-beta (TGF-beta) superfamily, including TGF-beta, bone morphogenetic proteins (BMPs), activins and nodals, are vital for regulating growth and differentiation. These growth factors transduce their signals through pairs of transmembrane type I and type II receptor kinases. Here, we have cloned a transmembrane protein, BAMBI, which is related to TGF-beta-family type I receptors but lacks an intracellular kinase domain. We show that BAMBI is co-expressed with the ventralizing morphogen BMP4 (refs 5, 6) during Xenopus embryogenesis and that it requires BMP signalling for its expression. The protein stably associates with TGF-beta-family receptors and inhibits BMP and activin as well as TGF-beta signalling. Finally, we provide evidence that BAMBI's inhibitory effects are mediated by its intracellular domain, which resembles the homodimerization interface of a type I receptor and prevents the formation of receptor complexes. The results indicate that BAMBI negatively regulates TGF-beta-family signalling by a regulatory mechanism involving the interaction of signalling receptors with a pseudoreceptor.
Asunto(s)
Proteínas de la Membrana/metabolismo , Transducción de Señal , Factor de Crecimiento Transformador beta/metabolismo , Proteínas de Xenopus , Activinas , Secuencia de Aminoácidos , Animales , Proteína Morfogenética Ósea 4 , Receptores de Proteínas Morfogenéticas Óseas de Tipo 1 , Proteínas Morfogenéticas Óseas/antagonistas & inhibidores , Proteínas Morfogenéticas Óseas/metabolismo , Células COS , Técnicas de Cultivo , Embrión no Mamífero/metabolismo , Expresión Génica , Humanos , Inhibinas/antagonistas & inhibidores , Inhibinas/metabolismo , Ligandos , Proteínas de la Membrana/química , Proteínas de la Membrana/genética , Ratones , Datos de Secuencia Molecular , Proteínas Serina-Treonina Quinasas/química , Receptores de Factores de Crecimiento/química , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Homología de Secuencia , Células Tumorales Cultivadas , XenopusRESUMEN
The marginal zone is a ring of tissue that gives rise to a characteristic dorsoventral pattern of mesoderm in amphibian embryos. Bmp-4 is thought to play an important role in specifying ventral mesodermal fate. Here we show (1) that different doses of Bmp-4 are sufficient to pattern four distinct mesodermal cell types and to pattern gene expression in the early gastrula marginal zone into three domains, (2) that there is a graded requirement for a Bmp signal in mesodermal patterning, and (3) that Bmp-4 has long-range activity which can become graded in the marginal zone by the antagonizing action of noggin. The results argue that Bmp-4 acts as a morphogen in dorsoventral patterning of mesoderm.
Asunto(s)
Proteínas Morfogenéticas Óseas/fisiología , Proteínas de Unión al ADN , Regulación del Desarrollo de la Expresión Génica , Mesodermo/fisiología , Transactivadores , Factores de Transcripción , Proteínas de Xenopus , Xenopus/embriología , Animales , Biomarcadores , Blastocisto , Proteína Morfogenética Ósea 4 , Proteínas Portadoras , Diferenciación Celular/genética , Embrión no Mamífero/fisiología , Inducción Embrionaria/genética , Gástrula/fisiología , Dosificación de Gen , Proteínas de Homeodominio/genética , Proteínas Musculares/fisiología , Factor 5 Regulador Miogénico , Proteínas/fisiologíaRESUMEN
We describe a novel Xenopus homeobox gene, Xvent-2, which together with the previously identified homeobox gene Xvent-1, defines a novel class of homeobox genes. vent genes are related by sequence homology, expression pattern and gain-of-function phenotype. Evidence is presented for a role of Xvent-2 in the BMP-4 pathway involved in dorsoventral patterning of mesoderm. (1) Xvent-2 is expressed in regions that also express BMP-4. (2) Xvent-2 and BMP-4 interact in a positive feedback loop. (3) Xvent-2 ventralizes dorsal mesoderm in a dose-dependent manner resulting in phenoytpes ranging from microcephaly to Bauchstück pieces, as does BMP-4. (4) Like BMP-4 and gsc, Xvent-2 and gsc are able to interact in a crossregulatory loop to suppress each other. (5) Microinjection of Xvent-2 mRNA can rescue dorsalization by a dominant-negative BMP-4 receptor. The results suggest that Xvent-2 functions in the BMP-4 signalling pathway that antagonizes the Spemann organizer.
Asunto(s)
Proteínas Morfogenéticas Óseas/metabolismo , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Receptores de Factores de Crecimiento , Transducción de Señal/fisiología , Factores de Transcripción , Proteínas de Xenopus , Xenopus/embriología , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Receptores de Proteínas Morfogenéticas Óseas , Diferenciación Celular , Clonación Molecular , ADN Complementario , Expresión Génica , Mesodermo , Microinyecciones , Datos de Secuencia Molecular , Notocorda/citología , Receptores de Superficie CelularRESUMEN
The Myc protein binds to and transactivates the expression of genes via E-box elements containing a central CAC(G/A)TG sequence. The transcriptional activation function of Myc is required for its ability to induce cell cycle progression, cellular transformation and apoptosis. Here we show that transactivation by Myc is under negative control by the transcription factor AP-2. AP-2 inhibits transactivation by Myc via two distinct mechanisms. First, high affinity binding sites for AP-2 overlap Myc-response elements in two bona fide target genes of Myc, prothymosin-alpha and ornithine decarboxylase. On these sites, AP-2 competes for binding of either Myc/Max heterodimers or Max/Max homodimers. The second mechanism involves a specific interaction between C-terminal domains of AP-2 and the BR/HLH/LZ domain of Myc, but not Max or Mad. Binding of AP-2 to Myc does not preclude association of Myc with Max, but impairs DNA binding of the Myc/Max complex and inhibits transactivation by Myc even in the absence of an overlapping AP-2 binding site. Taken together, our data suggest that AP-2 acts as a negative regulator of transactivation by Myc.