A role of cyclin G in the process of apoptosis. (33/1354)

Cyclin G was previously identified as a target gene of the p53 tumor suppresser protein, and levels of cyclin G are increased after induction of p53 by DNA damage. However, the function of cyclin G has not been established. To determine the effect of increased expression of cyclin G, retroviruses encoding cyclin G were constructed and used to infect three different murine cell lines. Cyclin G protein levels induced by the retroviruses were within the range seen after DNA damage induction of p53. In each case we observed that such over-expression of cyclin G augments the apoptotic process. TNF-alpha induction of apoptosis is increased by expression of cyclin G in NIH3T3 fibroblasts which express p53, as well as in 10.1 fibroblasts which contain no p53 allele. Additionally, we observed that while cyclin G expression is markedly reduced upon aggregate formation in embryonic carcinoma P19 cells, retrovirus-mediated over-expression of cyclin G enhances apoptotic cell death in aggregated P19 cells, and increases the extent of apoptosis caused by retinoic acid or serum starvation of these cells. These data demonstrate that cyclin G plays a facilitating role in modulating apoptosis induced by different stimuli. Moreover, we have discovered that cyclin G expression is rapidly induced in P19 cells after exposure to Bone Morphogenic Protein-4 (BMP-4), suggesting that cyclin G may mediate apoptotic signals generated by BMP-4.  (+info)

Isolation and characterization of bone morphogenetic protein-binding proteins from the early Xenopus embryo. (34/1354)

Using a surface plasmon resonance biosensor as a sensitive and specific monitor, we have isolated two distinct bone morphogenetic protein (BMP)-binding proteins, and identified them as lipovitellin 1 and Ep45, respectively. Lipovitellin 1 is an egg yolk protein that is processed from vitellogenin. Both vitellogenin and Ep45 are synthesized under estrogen control in the liver, secreted, and taken up by developing oocytes. In this paper, we have shown that of the TGF-beta family members tested, Ep45 can bind only to BMP-4, whereas lipovitellin 1 can bind to both BMP-4 and activin A. Because of this difference in specificity, we have focused on and further studied Ep45. Kinetic parameters were determined by surface plasmon resonance studies and showed that Ep45 associated rapidly with BMP-4 (k(a) = 1.06 x 10(4) M(-1)s(-1)) and dissociated slowly (k(d) = 1.6 x 10(-4) s(-1)). In Xenopus embryos microinjected with Ep45 mRNA, Ep45 blocked the ability of follistatin to inhibit BMP activity and to induce a secondary body axis in a dose-dependent manner, whereas it had no effect on other BMP antagonists, chordin and noggin. These results support the possibility that Ep45 interacts with BMP to modulate its activities in vivo.  (+info)

Developmental effects of ectopic expression of the glucocorticoid receptor DNA binding domain are alleviated by an amino acid substitution that interferes with homeodomain binding. (35/1354)

Steroid hormone receptors are distinguished from other members of the nuclear hormone receptor family through their association with heat shock proteins and immunophilins in the absence of ligands. Heat shock protein association represses steroid receptor DNA binding and protein-protein interactions with other transcription factors and facilitates hormone binding. In this study, we investigated the hormone-dependent interaction between the DNA binding domain (DBD) of the glucocorticoid receptor (GR) and the POU domains of octamer transcription factors 1 and 2 (Oct-1 and Oct-2, respectively). Our results indicate that the GR DBD binds directly, not only to the homeodomains of Oct-1 and Oct-2 but also to the homeodomains of several other homeodomain proteins. As these results suggest that the determinants for binding to the GR DBD are conserved within the homeodomain, we examined whether the ectopic expression of GR DBD peptides affected early embryonic development. The expression of GR DBD peptides in one-cell-stage zebra fish embryos severely affected their development, beginning with a delay in the epibolic movement during the blastula stage and followed by defects in convergence-extension movements during gastrulation, as revealed by the abnormal patterns of expression of several dorsal gene markers. In contrast, embryos injected with mRNA encoding a GR peptide with a point mutation that disrupted homeodomain binding or with mRNA encoding the DBD of the closely related mineralocorticoid receptor, which does not bind octamer factors, developed normally. Moreover, coinjection of mRNA encoding the homeodomain of Oct-2 completely rescued embryos from the effects of the GR DBD. These results highlight the potential of DNA-independent effects of GR in a whole-animal model and suggest that at least some of these effects may result from direct interactions with homeodomain proteins.  (+info)

Three different noggin genes antagonize the activity of bone morphogenetic proteins in the zebrafish embryo. (36/1354)

The dorsoventral polarity of the vertebrate embryo is established through interactions between ventrally expressed bone morphogenetic proteins and their organizer-borne antagonists Noggin, Chordin, and Follistatin. While the opposing interactions between Short Gastrulation/Chordin and Decapentaplegic/BMP4 have been evolutionarily conserved in arthropods and vertebrates, there has been up to now no functional evidence of an implication of Noggin in the early patterning of organisms other than Xenopus. We have studied the contribution of Noggin to the embryonic development of the zebrafish. While single-copy noggin genes have been characterized in several vertebrate species, we report that the zebrafish genome harbors three noggin homologues. Overexpression experiments show that Noggin1, Noggin2, and Noggin3 can antagonize ventralizing BMPs. While all three factors have similar biological activities, their embryonic expression is different. The combined expression of the three genes recapitulates the different aspects of the expression of the single-copy noggin genes of other organisms. This suggests that the three zebrafish noggin genes and the single noggin genes of other vertebrates have evolved from a common ancestor and that subsequent differential loss of tissue-specific elements in the promoters of the different zebrafish genes accounts for their more restricted spatiotemporal expression. Finally we show that noggin1 is expressed in the fish organizer and able to dorsalize the embryo, suggesting its implication in the dorsoventral patterning of the zebrafish.  (+info)

Patterning of the mesoderm involves several threshold responses to BMP-4 and Xwnt-8. (37/1354)

Two secreted signaling molecules, Xwnt-8 and BMP-4, play an essential role in the dorso-ventral patterning of the mesoderm in Xenopus. Here we investigate how the Wnt-8 and the BMP-4 pathways are connected and how they regulate target genes in the lateral and ventral marginal zone. BMP-4 regulates the transcription of Xwnt-8 in a threshold dependent manner. High levels of BMP-4 induce the expression of the Wnt antagonist sizzled in the ventral marginal zone, independent of Xwnt-8 signaling. Xwnt-8 induces the early muscle marker myf-5 in the lateral marginal zone in a BMP independent manner. The expression of the homeobox gene Xvent-1 can be modulated through both the BMP-4 and the Xwnt-8 pathways. The spatial distribution and the level of BMP-4 activity in the lateral and ventral marginal zone is reflected in the dynamic expression pattern of Xwnt-8. The data support the view that Xwnt-8 is involved in the specification of lateral (somitogenic) mesoderm and BMP-4 in the specification of ventral mesoderm.  (+info)

Dual role of brain factor-1 in regulating growth and patterning of the cerebral hemispheres. (38/1354)

Brain factor-1 (BF-1) is a winged-helix (WH) transcription factor with a restricted pattern of expression in the neural tube. In the embryo, BF-1 is localized to the progenitor cells of the most rostral neural tube, the telencephalic neuroepithelium. Expression of BF-1 persists in the adult brain in the structures derived from the telencephalon, including the cerebral cortex, the hippocampus, the olfactory bulbs and the basal ganglia. Targeted disruption of the BF-1 gene in mice results in hypoplasia of the cerebral hemispheres. Proliferation of the telencephalic neuroepithelium is decreased and neuronal differentiation occurs prematurely. The forebrain of the BF-1 (-/-) mutant also displays dorsal-ventral patterning defects. Development of the ventral (basal) region of the telencephalon is more severely affected than the dorsal region. These anomalies are associated with the ectopic expression of BMP4 in the dorsal telencephalic neuroepithelium and the loss of shh in the ventral telencephalon. These results raise the possibility that BF-1 may modulate both progenitor cell proliferation and regional patterning by regulating the expression or activity of inductive signals which act on the telencephalic neuroepithelium.  (+info)

The dominant hemimelia mutation uncouples epithelial-mesenchymal interactions and disrupts anterior mesenchyme formation in mouse hindlimbs. (39/1354)

Epithelial-mesenchymal interactions are essential for both limb outgrowth and pattern formation in the limb. Molecules capable of communication between these two tissues are known and include the signaling molecules SHH and FGF4, FGF8 and FGF10. Evidence suggests that the pattern and maintenance of expression of these genes are dependent on a number of factors including regulatory loops between genes expressed in the AER and those in the underlying mesenchyme. We show here that the mouse mutation dominant hemimelia (Dh) alters the pattern of gene expression in the AER such that Fgf4, which is normally expressed in a posterior domain, and Fgf8, which is expressed throughout are expressed in anterior patterns. We show that maintenance of Shh expression in the posterior mesenchyme is not dependent on either expression of Fgf4 or normal levels of Fgf8 in the overlying AER. Conversely, AER expression of Fgf4 is not directly dependent on Shh expression. Also the reciprocal regulatory loop proposed for Fgf8 in the AER and Fgf10 in the underlying mesenchyme is also uncoupled by this mutation. Early during the process of limb initiation, Dh is involved in regulating the width of the limb bud, the mutation resulting in selective loss of anterior mesenchyme. The Dh gene functions in the initial stages of limb development and we suggest that these initial roles are linked to mechanisms that pattern gene expression in the AER.  (+info)

Regulation of the onset of neural crest migration by coordinated activity of BMP4 and Noggin in the dorsal neural tube. (40/1354)

For neural crest cells to engage in migration, it is necessary that epithelial premigratory crest cells convert into mesenchyme. The mechanisms that trigger cell delamination from the dorsal neural tube remain poorly understood. We find that, in 15- to 40-somite-stage avian embryos, BMP4 mRNA is homogeneously distributed along the longitudinal extent of the dorsal neural tube, whereas its specific inhibitor noggin exists in a gradient of expression that decreases caudorostrally. This rostralward reduction in signal intensity coincides with the onset of emigration of neural crest cells. Hence, we hypothesized that an interplay between Noggin and BMP4 in the dorsal tube generates graded concentrations of the latter that in turn triggers the delamination of neural crest progenitors. Consistent with this suggestion, disruption of the gradient by grafting Noggin-producing cells dorsal to the neural tube at levels opposite the segmental plate or newly formed somites, inhibited emigration of HNK-1-positive crest cells, which instead accumulated within the dorsal tube. Similar results were obtained with explanted neural tubes from the same somitic levels exposed to Noggin. Exposure to Follistatin, however, had no effect. The Noggin-dependent inhibition was overcome by concomitant treatment with BMP4, which when added alone, also accelerated cell emigration compared to untreated controls. Furthermore, the observed inhibition of neural crest emigration in vivo was preceded by a partial or total reduction in the expression of cadherin-6B and rhoB but not in the expression of slug mRNA or protein. Altogether, these results suggest that a coordinated activity of Noggin and BMP4 in the dorsal neural tube triggers delamination of specified, slug-expressing neural crest cells. Thus, BMPs play multiple and discernible roles at sequential stages of neural crest ontogeny, from specification through delamination and later differentiation of specific neural crest derivatives.  (+info)