Transducing the Dpp morphogen gradient in the wing of Drosophila: regulation of Dpp targets by brinker.
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Dpp, a TGFbeta, organizes pattern in the Drosophila wing by acting as a graded morphogen, activating different targets above distinct threshold concentrations. Like other TGFbetas, Dpp appears to induce transcription directly via activation of a SMAD, Mad. However, here we demonstrate that Dpp can also control gene expression indirectly by downregulating the expression of the brinker gene, which encodes a putative transcription factor that functions to repress Dpp targets. The medial-to-lateral Dpp gradient along the anterior-posterior axis is complemented by a lateral-to-medial gradient of Brinker, and the presence of these two opposing gradients may function to allow cells to detect small differences in Dpp concentration and respond by activating different target genes. (+info)
The Drosophila gene brinker reveals a novel mechanism of Dpp target gene regulation.
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decapentaplegic (dpp), a Drosophila member of the TGFbeta family of secreted molecules, functions as a long-range morphogen in patterning of the embryo and the adult appendages. Dpp signals via the SMAD proteins Mad and Medea. Here we show that in the absence of brinker (brk), Mad is not required for the activation of Dpp target genes that depend on low levels of Dpp. brk encodes a novel protein with features of a transcriptional repressor. brk itself is negatively regulated by Dpp. Dpp signaling might relieve brk's repression of low-level target genes either by transcriptional repression of brk or by antagonizing a repressor function of brk at the target gene promoters. (+info)
Restricted expression of the receptor serine/threonine kinase BMPR-IB in zebrafish.
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Bone morphogenetic proteins (BMPs) comprise a rapidly expanding subclass of the transforming growth factor-beta superfamily. They are known to regulate a diverse range of developmental phenomena including cell differentiation, morphogenesis and apoptosis. In this study, we have isolated a zebrafish homolog of BMP type IB receptor (BMPR-IB) and examined the localization of the transcripts during embryogenesis. Whole-mount in situ hybridization analysis revealed that unlike other type I and type II receptors that mediate BMP signal, it is expressed in developing somite and in mid-hind brain region in a restricted manner. (+info)
Bmp signaling regulates proximal-distal differentiation of endoderm in mouse lung development.
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In the mature mouse lung, the proximal-distal (P-D) axis is delineated by two distinct epithelial subpopulations: the proximal bronchiolar epithelium and the distal respiratory epithelium. Little is known about the signaling molecules that pattern the lung along the P-D axis. One candidate is Bone Morphogenetic Protein 4 (Bmp4), which is expressed in a dynamic pattern in the epithelial cells in the tips of growing lung buds. Previous studies in which Bmp4 was overexpressed in the lung endoderm (Bellusci, S., Henderson, R., Winnier, G., Oikawa, T. and Hogan, B. L. M. (1996) Development 122, 1693-1702) suggested that this factor plays an important role in lung morphogenesis. To further investigate this question, two complementary approaches were utilized to inhibit Bmp signaling in vivo. The Bmp antagonist Xnoggin and, independently, a dominant negative Bmp receptor (dnAlk6), were overexpressed using the surfactant protein C (Sp-C) promoter/enhancer. Inhibiting Bmp signaling results in a severe reduction in distal epithelial cell types and a concurrent increase in proximal cell types, as indicated by morphology and expression of marker genes, including the proximally expressed hepatocyte nuclear factor/forkhead homologue 4 (Hfh4) and Clara cell marker CC10, and the distal marker Sp-C. In addition, electron microscopy demonstrates the presence of ciliated cells, a proximal cell type, in the most peripheral regions of the transgenic lungs. We propose a model in which Bmp4 is a component of an apical signaling center controlling P-D patterning. Endodermal cells at the periphery of the lung, which are exposed to high levels of Bmp4, maintain or adopt a distal character, while cells receiving little or no Bmp4 signal initiate a proximal differentiation program. (+info)
Characterization of bone morphogenetic protein-6 signaling pathways in osteoblast differentiation.
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Bone morphogenetic protein (BMP)-6 is a member of the transforming growth factor (TGF)-(&bgr;) superfamily, and is most similar to BMP-5, osteogenic protein (OP)-1/BMP-7, and OP-2/BMP-8. In the present study, we characterized the endogenous BMP-6 signaling pathway during osteoblast differentiation. BMP-6 strongly induced alkaline phosphatase (ALP) activity in cells of osteoblast lineage, including C2C12 cells, MC3T3-E1 cells, and ROB-C26 cells. The profile of binding of BMP-6 to type I and type II receptors was similar to that of OP-1/BMP-7 in C2C12 cells and MC3T3-E1 cells; BMP-6 strongly bound to activin receptor-like kinase (ALK)-2 (also termed ActR-I), together with type II receptors, i.e. BMP type II receptor (BMPR-II) and activin type II receptor (ActR-II). In addition, BMP-6 weakly bound to BMPR-IA (ALK-3), to which BMP-2 also bound. In contrast, binding of BMP-6 to BMPR-IB (ALK-6), and less efficiently to ALK-2 and BMPR-IA, together with BMPR-II was detected in ROB-C26 cells. Intracellular signalling was further studied using C2C12 and MC3T3-E1 cells. Among the receptor-regulated Smads activated by BMP receptors, BMP-6 strongly induced phosphorylation and nuclear accumulation of Smad5, and less efficiently those of Smad1. However, Smad8 was constitutively phosphorylated, and no further phosphorylation or nuclear accumulation of Smad8 by BMP-6 was observed. These findings indicate that in the process of differentiation to osteoblasts, BMP-6 binds to ALK-2 as well as other type I receptors, and transduces signals mainly through Smad5 and possibly through Smad1. (+info)
Roles of bone morphogenetic protein type I receptors and Smad proteins in osteoblast and chondroblast differentiation.
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The biological effects of type I serine/threonine kinase receptors and Smad proteins were examined using an adenovirus-based vector system. Constitutively active forms of bone morphogenetic protein (BMP) type I receptors (BMPR-IA and BMPR-IB; BMPR-I group) and those of activin receptor-like kinase (ALK)-1 and ALK-2 (ALK-1 group) induced alkaline phosphatase activity in C2C12 cells. Receptor-regulated Smads (R-Smads) that act in the BMP pathways, such as Smad1 and Smad5, also induced the alkaline phosphatase activity in C2C12 cells. BMP-6 dramatically enhanced alkaline phosphatase activity induced by Smad1 or Smad5, probably because of the nuclear translocation of R-Smads triggered by the ligand. Inhibitory Smads, i.e., Smad6 and Smad7, repressed the alkaline phosphatase activity induced by BMP-6 or the type I receptors. Chondrogenic differentiation of ATDC5 cells was induced by the receptors of the BMPR-I group but not by those of the ALK-1 group. However, kinase-inactive forms of the receptors of the ALK-1 and BMPR-I groups blocked chondrogenic differentiation. Although R-Smads failed to induce cartilage nodule formation, inhibitory Smads blocked it. Osteoblast differentiation induced by BMPs is thus mediated mainly via the Smad-signaling pathway, whereas chondrogenic differentiation may be transmitted by Smad-dependent and independent pathways. (+info)
Combinatorial signaling through BMP receptor IB and GDF5: shaping of the distal mouse limb and the genetics of distal limb diversity.
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In this study, we use a mouse insertional mutant to delineate gene activities that shape the distal limb skeleton. A recessive mutation that results in brachydactyly was found in a lineage of transgenic mice. Sequences flanking the transgene insertion site were cloned, mapped to chromosome 3, and used to identify the brachydactyly gene as the type IB bone morphogenetic protein receptor, BmprIB (ALK6). Expression analyses in wild-type mice revealed two major classes of BmprIB transcripts. Rather than representing unique coding RNAs generated by alternative splicing of a single pro-mRNA transcribed from one promoter, the distinct isoforms reflect evolution of two BmprIB promoters: one located distally, driving expression in the developing limb skeleton, and one situated proximally, initiating transcription in neural epithelium. The distal promoter is deleted in the insertional mutant, resulting in a regulatory allele (BmprIB(Tg)) lacking cis-sequences necessary for limb BmprIB expression. Mutants fail to generate digit cartilage, indicating that BMPRIB is the physiologic transducer for the formation of digit cartilage from the skeletal blastema. Expansion of BmprIB expression into the limb through acquisition of these distal cis-regulatory sequences appears, therefore, to be an important genetic component driving morphological diversity in distal extremities. GDF5 is a BMP-related signal, which is also required for proper digit formation. Analyses incorporating both Gdf5 and BmprIB(Tg) alleles revealed that BMPRIB regulates chondrogenesis and segmentation through both GDF5-dependent and -independent processes, and that, reciprocally, GDF5 acts through both IB and other type I receptors. Together, these findings provide in vivo support for the concept of combinatorial BMP signaling, in which distinct outcomes result both from a single receptor being triggered by different ligands and from a single ligand binding to different receptors. (+info)
The type I BMP receptor BMPRIB is required for chondrogenesis in the mouse limb.
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Mice carrying a targeted disruption of BmprIB were generated by homologous recombination in embryonic stem cells. BmprIB(-/-) mice are viable and, in spite of the widespread expression of BMPRIB throughout the developing skeleton, exhibit defects that are largely restricted to the appendicular skeleton. Using molecular markers, we show that the initial formation of the digital rays occurs normally in null mutants, but proliferation of prechondrogenic cells and chondrocyte differentiation in the phalangeal region are markedly reduced. Our results suggest that BMPRIB-mediated signaling is required for cell proliferation after commitment to the chondrogenic lineage. Analyses of BmprIB and Gdf5 single mutants, as well as BmprIB; Gdf5 double mutants suggests that GDF5 is a ligand for BMPRIB in vivo. BmprIB; Bmp7 double mutants were constructed in order to examine whether BMPRIB has overlapping functions with other type I BMP receptors. BmprIB; Bmp7 double mutants exhibit severe appendicular skeletal defects, suggesting that BMPRIB and BMP7 act in distinct, but overlapping pathways. These results also demonstrate that in the absence of BMPRIB, BMP7 plays an essential role in appendicular skeletal development. Therefore, rather than having a unique role, BMPRIB has broadly overlapping functions with other BMP receptors during skeletal development. (+info)