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D Büscher

Publications and source records attributed to D Büscher.

At least 19 recordsLinked to original sources
PubMed2001

WNT signals control FGF-dependent limb initiation and AER induction in the chick embryo.

A regulatory loop between the fibroblast growth factors FGF-8 and FGF-10 plays a key role in limb initiation and AER induction in vertebrate embryos. Here, we show that three WNT factors signaling through beta-catenin act as key regulators of the FGF-8/FGF-10 loop. The Wnt-2b gene is expressed in the intermediate mesoderm and the lateral plate mesoderm in the presumptive chick forelimb region. Cells expressing Wnt-2b are able to induce Fgf-10 and generate an extra limb when implanted into the flank. In the presumptive hindlimb region, another Wnt gene, Wnt-8c, controls Fgf-10 expression, and is also capable of inducing ectopic limb formation in the flank. Finally, we also show that the induction of Fgf-8 in the limb ectoderm by FGF-10 is mediated by the induction of Wnt-3a. Thus, three WNT signals mediated by beta-catenin control both limb initiation and AER induction in the vertebrate embryo.

Animals↗
PubMed2000

The Raf-1 kinase associates with vimentin kinases and regulates the structure of vimentin filaments.

Using immobilized GST-Raf-1 as bait, we have isolated the intermediate filament protein vimentin as a Raf-1-associated protein. Vimentin coimmunoprecipitated and colocalized with Raf-1 in fibroblasts. Vimentin was not a Raf-1 substrate, but was phosphorylated by Raf-1-associated vimentin kinases. We provide evidence for at least two Raf-1-associated vimentin kinases and identified one as casein kinase 2. They are regulated by Raf-1, since the activation status of Raf-1 correlated with the phosphorylation of vimentin. Vimentin phosphorylation by Raf-1 preparations interfered with its polymerization in vitro. A subset of tryptic vimentin phosphopeptides induced by Raf-1 in vitro matched the vimentin phosphopeptides isolated from v-raf-transfected cells labeled with orthophosphoric acid, indicating that Raf-1 also induces vimentin phosphorylation in intact cells. In NIH 3T3 fibroblasts, the selective activation of an estrogen-regulated Raf-1 mutant induced a rearrangement and depolymerization of the reticular vimentin scaffold similar to the changes elicited by serum treatment. The rearrangement of the vimentin network occurred independently of the MEK/ERK pathway. These data identify a new branch point in Raf-1 signaling, which links Raf-1 to changes in the cytoskeletal architecture.

Amino Acid Sequence↗
PubMed1999

IKK1-deficient mice exhibit abnormal development of skin and skeleton.

IkappaB kinases (IKKs) IKK1 and IKK2 are two putative IkappaBalpha kinases involved in NF-kappaB activation. To examine the in vivo functions of IKK1, we generated IKK1-deficient mice. The mutant mice are perinatally lethal and exhibit a wide range of developmental defects. Newborn mutant mice have shiny, taut, and sticky skin without whiskers. Histological analysis shows thicker epidermis, which is unable to differentiate. Limbs and tail are wrapped inside the skin and do not extend properly out of the body trunk. Skeleton staining reveals a cleft secondary palate, split sternebra 6, and deformed incisors. NF-kappaB activation mediated by TNFalpha and IL-1 is diminished in IKK1-deficient mouse embryonic fibroblast (MEF) cells. The IKK complex in the absence of IKK1 is capable of phosphorylating IkappaBalpha and IkappaBbeta in vitro. Our results support a role for IKK1 in NF-kappaB activation and uncover its involvement in skin and skeleton development. We conclude further that the two related kinases IKK1 and IKK2 have distinct functions and can not be substituted for each other's functions.

Animals↗
PubMed1999

Muscle development during vertebrate limb outgrowth.

Limb development has become one of the model systems for studying vertebrate development. One crucial aspect in limb development is the origin, differentiation and patterning of muscle. Much progress has been made in recent years towards understanding this process. One of the general observations is that the genes involved in limb muscle development appear to be very similar to those involved in muscle development in other regions of the embryo. In this review, we summarize some of the genes and mechanisms that regulate limb muscle development and discuss various avenues along which a deeper understanding can be gained of how muscle cells originate and differentiate in different tissues during vertebrate development.

Animals↗
PubMed1999

Molecular and cellular basis of pattern formation during vertebrate limb development.

The body plan is generated by cells and tissues that become arranged precisely in the embryo. This process, termed pattern formation, involves cell interactions in which a particular group of cells produce signals that specify new cell types or patterns of differentiation in responding cells. These patterning signals emanate from very discrete centers called "organizer centers," such as the Hensen's node or Spemann organizer, the midbrain-hindbrain junction, the notochord, or in the case of the limb, the zone of polarizing activity (ZPA) or the apical ectodermal ridge (AER). The developing vertebrate limb is an ideal model system for the study of pattern formation because, in addition to surgical manipulations, molecular manipulations are now feasible. In this review we summarize early experiments that established, by means of surgical manipulations, the different organizer centers of the vertebrate limb: the ectoderm covering the limb bud, the apical ectodermal ridge, the zone of polarizing activity, and the distal mesoderm (progress zone) underlying the AER. We then describe the domains of expression of various genes present during the development of the limb and discuss some of the functional approaches (overexpression and lack of function studies) undertaken to ascertain their role in limb outgrowth. The knowledge acquired in the last few years has had an enormous impact not only on our view of how limbs develop (perhaps now one of the most approachable vertebrate model systems) but also in a more general sense of how the embryo is organized in space and time.

Animals↗
PubMed1998

Expression profile of Gli family members and Shh in normal and mutant mouse limb development.

Gli genes represent a small family, encoding zinc-finger proteins of the Krüppel-type. The family consists of Gli(1), Gli2, and Gli3, all of which are expressed in the developing mouse limb bud. To assess the role of the Gli family and Sonic hedgehog (Shh) in mouse limb development, we compared the expression domains of all three Gli genes and of Shh. Although each Gli gene has its own distinct expression pattern in limb buds, at 10.5-11.5 dpc all three genes were found not to be expressed in the posterior region, the presumptive Shh expression domain. This transient mutually exclusive expression suggested a potential interaction between Gli genes and Shh. To address this matter, we analysed the expression of Gli genes and Shh in two polydactyly mouse mutants, Extra toes (Xt) and Hemimelic-extra toes (Hx) which express Shh ectopically in the anterior region of the limb field. Since Xt mice lack Gli3 expression, the ectopic Shh expression is genetically linked to the absence of Gli3. In Hx mice we found a down-regulation of Gli3 in the anterior region of the limb bud. In both mutants Gli2 expression pattern was not altered, whereas Gli1 expression was anteriorly up-regulated adjacent to the ectopic Shh domain. These results strongly suggest a positive regulation of Gli1 by Shh and a negative interaction between Shh and Gli3.

Animals↗
PubMed1998

The role of Alx-4 in the establishment of anteroposterior polarity during vertebrate limb development.

We have determined that Strong's Luxoid (lstJ) [corrected] mice have a 16 bp deletion in the homeobox region of the Alx-4 gene. This deletion, which leads to a frame shift and a truncation of the Alx-4 protein, could cause the polydactyly phenotype observed in lstJ [corrected] mice. We have cloned the chick homologue of Alx-4 and investigated its expression during limb outgrowth. Chick Alx-4 displays an expression pattern complementary to that of shh, a mediator of polarizing activity in the limb bud. Local application of Sonic hedgehog (Shh) and Fibroblast Growth Factor (FGF), in addition to ectodermal apical ridge removal experiments suggest the existence of a negative feedback loop between Alx-4 and Shh during limb outgrowth. Analysis of polydactylous mutants indicate that the interaction between Alx-4 and Shh is independent of Gli3, a negative regulator of Shh in the limb. Our data suggest the existence of a negative feedback loop between Alx-4 and Shh during vertebrate limb outgrowth.

Amino Acid Sequence↗
PubMed1997

Evidence for genetic control of Sonic hedgehog by Gli3 in mouse limb development.

Sonic hedgehog (Shh) expression in the developing limb is associated with the zone of polarising activity (ZPA), and both are restricted to the posterior part of the limb bud. We show that the expression patterns of Shh and Gli3, a member of the Gli-family believed to function in transcriptional control, appear to be mutually exclusive in limb buds of mouse embryos. In the polydactyly mouse mutant extra toes (Xt), possessing a null mutation of Gli3, Shh is additionally expressed in the anterior region of the limb bud. The transcript of Ptc, the putative receptor for Shh protein, can be detected anteriorly as well. Other genes known to be involved in limb outgrowth and patterning, like Fibroblast growth factor (Fgf), Bone morphogenetic protein (Bmp), and Hoxd are misexpressed in relation to the ectopic Shh expression domain in Xt limb buds. This data suggest that Gli3 is a regulator of Shh expression in mouse limb development.

Animals↗
PubMed1996

Cloning and sequence analysis of the murine Gli3 cDNA.

The zinc finger gene Gli3 plays a role in limb and brain development. To facilitate the molecular analysis of different mouse mutations of this gene, the murine cDNA was isolated and sequenced. This 5113 bp cDNA encodes a putative protein of 1596 amino acids. Comparison of the murine and human GLI3 cDNA revealed an overall homology of 85% between the deduced amino acid sequences. More importantly, several regions of the protein, including the zinc fingers, are more highly conserved ( > 95%), suggesting that these represent functional domains in the Gli3 protein.

Amino Acid Sequence↗
PubMed1996

Involvement of the protein tyrosine phosphatase SHP-1 in Ras-mediated activation of the mitogen-activated protein kinase pathway.

Ubiquitously expressed SH2-containing tyrosine phosphatases interact physically with tyrosine kinase receptors or their substrates and relay positive mitogenic signals via the activation of the Ras-mitogen-activated protein kinase (MAPK) pathway. Conversely, the structurally related phosphatase SHP-1 is predominantly expressed in hemopoietic cells and becomes tyrosine phosphorylated upon colony-stimulating factor 1 treatment of macrophages without associating with the colony-stimulating factor 1 receptor tyrosine kinase. Mice lacking functional SHP-1 (me/me and me(v)/me(v)) develop systemic autoimmune disease with accumulation of macrophages, suggesting that SHP-1 may be a negative regulator of hemopoietic cell growth. By using macrophages expressing dominant negative Ras and the me(v)/me(v) mouse mutant, we show that SHP-1 is activated in the course of mitogenic signal transduction in a Ras-dependent manner and that its activity is necessary for the Ras-dependent activation of the MAPK pathway but not of the Raf-1 kinase. Consistent with a role for SHP-1 as an intermediate between Ras and the MEK-MAPK pathway, Ras-independent activation of the latter kinases by bacterial lipopolysaccharide occurred normally in me(v)/me(v) cells. Our results sharply accentuate the diversity of signal transduction in mammalian cells, in which the same signaling intermediates can be rearranged to form different pathways.

Animals↗
PubMed1995

Suppression of growth factor-mediated MAP kinase activation by v-raf in macrophages: a putative role for the MKP-1 phosphatase.

Many tyrosine kinase growth factor receptors activate the MAP Kinase (MAPK) pathway by stimulating the activity of the RAF kinase. In some, but not all cell types, the expression of activated RAF is sufficient to induce constitutive MAPK activation. In BAC-1.2F5 macrophages the expression of virally activated RAF does not correlate with constitutive MAPK activation; on the contrary, growth factor-mediated stimulation of MAPK activity is suppressed in these cells. Suppression correlates with v-RAF expression, as MAPK activation is normal in a revertant cell line that stopped expressing v-RAF. Inhibition of MAPK activation is associated with lack of ERK-2 tyrosine phosphorylation, and is not due to the suppression of CSF-1-mediated MEK activation. Pretreatment with vanadate restores growth factor-stimulated activation and tyrosine phosphorylation of MAPK in v-RAF-expressing macrophages, indicating the involvement of a tyrosine phosphatase. Interestingly, v-RAF-expressing macrophages contain low constitutive levels of MKP-1 mRNA, an immediate early gene that encodes a MAPK-specific phosphatase and is induced in the parental cell line by CSF-1 treatment. The restoration of MAPK activation by vanadate pretreatment and the presence of MKP-1 mRNA in v-RAF-expressing macrophages raise the intriguing possibility that in macrophages RAF may be feeding back on the MAPK pathway by participating in the control of MKP-1 expression.

Amino Acid Sequence↗
PubMed1995

Ras-dependent and -independent pathways target the mitogen-activated protein kinase network in macrophages.

Mitogen-activated protein kinases (MAPKs) are activated upon a variety of extracellular stimuli in different cells. In macrophages, colony-stimulating factor 1 (CSF-1) stimulates proliferation, while bacterial lipopolysaccharide (LPS) inhibits cell growth and causes differentiation and activation. Both CSF-1 and LPS rapidly activate the MAPK network and induce the phosphorylation of two distinct ternary complex factors (TCFs), TCF/Elk and TCF/SAP. CSF-1, but not LPS, stimulated the formation of p21ras. GTP complexes. Expression of a dominant negative ras mutant reduced, but did not abolish, CSF-1-mediated stimulation of MEK and MAPK. In contrast, activation of the MEK kinase Raf-1 was Ras independent. Treatment with the phosphatidylcholine-specific phospholipase C inhibitor D609 suppressed LPS-mediated, but not CSF-1-mediated, activation of Raf-1, MEK, and MAPK. Similarly, down-regulation or inhibition of protein kinase C blocked MEK and MAPK induction by LPS but not that by CSF-1. Phorbol 12-myristate 13-acetate pretreatment led to the sustained activation of the Raf-1 kinase but not that of MEK and MAPK. Thus, activated Raf-1 alone does not support MEK/MAPK activation in macrophages. Phosphorylation of TCF/Elk but not that of TCF/SAP was blocked by all treatments that interfered with MAPK activation, implying that TCF/SAP was targeted by a MAPK-independent pathway. Therefore, CSF-1 and LPS target the MAPK network by two alternative pathways, both of which induce Raf-1 activation. The mitogenic pathway depends on Ras activity, while the differentiation signal relies on protein kinase C and phosphatidylcholine-specific phospholipase C activation.

Amino Acid Sequence↗
PubMed1994

Lipopolysaccharide induces activation of the Raf-1/MAP kinase pathway. A putative role for Raf-1 in the induction of the IL-1 beta and the TNF-alpha genes.

Bacterial LPS is a potent macrophage activator. The early steps in LPS signal transduction involve the tyrosine phosphorylation and activation of a number of kinases of the src family, and inhibition of this pathway causes a severe impairment in the production of the cytokines TNF-alpha and IL-1 beta. We find that LPS-induced macrophages activation also involves the Raf-1 kinase, a key component in mitogenic signal transduction. Treatment of BAC-1.2F5 macrophages with LPS causes phosphorylation and activation of Raf-1. This is paralleled by the stimulation of MEK-1 and MAP-kinase activity and by the phosphorylation of the transcription factor Elk-1, a nuclear target of MAP-kinase. Activation of the Raf/MAP-kinase pathway was inhibited upon pretreatment of the cells with genistein, a tyrosine kinase inhibitor. Raf-1 must thus lie downstream of tyrosine kinase in LPS signal transduction. However, Raf-1 is not a direct substrate of a LPS-induced tyrosine kinase, because Raf-1 immunoisolated from LPS-induced cells contains only phosphoserine. This resembles the situation after CSF-1-stimulation of macrophages, in which Raf-1 clearly transduces a signal generated by the CSF-1 receptor kinase, but is phosphorylated exclusively in serine. Phosphopeptide maps of Raf-1 immunoprecipitated from LPS- or CSF-1-treated cells are indistinguishable, suggesting that these agents activate Raf-1 by similar mechanisms. Finally, v-raf-infected BAC-1.2F5 macrophages were found to constitutively express low levels of IL-1 beta and TNF-alpha. These data argue that Raf-1 functions downstream of tyrosine kinases in LPS-mediated macrophage activation and cytokine production.

Amino Acid Sequence↗
PubMed1994

Ras/MAP kinase-dependent and -independent signaling pathways target distinct ternary complex factors.

Transcriptional activation of the immediate early genes c-fos and egr-1 by extracellular signals appears to be mediated by ternary complex factors (TCFs). In BAC-1 macrophages, growth factor stimulation leads to the retardation of protein-DNA complexes containing distinct TCFs. One TCF is recognized by Elk-1 antisera, whereas the other is immunologically related to SAP-1. The appearance and decay of hyperphosphorylated TCF/Elk-1-containing complexes after stimulation coincide with the activation of mitogen-activated protein kinase (MAPK) and the induction and repression of c-fos and egr-1, whereas modified TCF/SAP-1-containing complexes decay more slowly. Suppression of MAPK activation in macrophages and fibroblasts correlates with the failure to induce TCF/Elk-1 hyperphosphorylation without blocking TCF/SAP-1 modification. Accordingly the modified Elk-1 complex is generated in vitro by activated MAPK, whereas that of SAP-1 is not. Expression of a dominant-negative Ras mutant (RasAsn17) in BAC-1 cells does not affect CSF-1-induced TCF/SAP-1 modification while suppressing TCF/Elk-1 phosphorylation. Neither PKC down-regulation by TPA nor inhibition of Gi proteins by pertussis toxin pretreatment influences CSF-1-induced signaling to TCFs. These data indicate the existence of two separate signaling pathways for the modification of distinct TCFs: one dependent on Ras and MAPK and converging on TCF/Elk-1, and the other targeting TCF/SAP-1 independently of Ras and MAPK.

3T3 Cells↗
PubMed1993

v-raf confers CSF-1 independent growth to a macrophage cell line and leads to immediate early gene expression without MAP-kinase activation.

The BAC-1.2F5 macrophage cell line depends on CSF-1 for proliferation and survival. Phosphorylation and activation of the RAF-1 kinase are among the early events in CSF-1 signal transduction. To characterize the role of RAF-1 in CSF-1-induced proliferation, we overexpressed oncogenically activated RAF-1, cellular RAF-1 and RAF-1 kinase-defective mutant proteins in BAC-1.2F5 cells. We were unable to establish stable cell lines expressing either kinase-negative or full length RAF-1 proteins, implying that expression of these molecules is not tolerated in BAC-1.2F5 cells. Oncogenically activated RAF-1 induces CSF-1-independent growth in the absence of autocrine growth factor production. Autonomous growth is not associated with dedifferentiation, since v-raf-expressing macrophages perform the same immunological functions as control cells. Intriguingly, autonomous growth correlates with the suppression of CSF-1-mediated MAP-Kinase activation and with the low constitutive expression of a number of CSF-1-inducible genes, including fos, jun, ets2, and myc, but also the genes for the inflammatory cytokines TNF alpha and IL-1 beta. Many of these genes have AP-1 binding sites in their promoters, and the v-raf-expressing cells contain constitutive AP-1 binding activity. These data indicate that RAF-1, but not MAP-Kinase, is a key component in CSF-1 mitogenic signal transduction, and are consistent with a working hypothesis in which RAF-1 mediates transcriptional activation of genes via AP-1.

Amino Acid Sequence↗
PubMed1979

[Reconstructive measures to prevent lower extremity amputation for atherosclerotic occlusive disease in old age and the results obtained between 1973 to 1977 (author's transl)].

Within a 5-year-period 102 patients over the age of 70 were admitted with atherosclerotic occlusive disease. All patients had rest pain or necrosis. In 12,8% only a primary amputation could be performed. The procedures and results of reconstructive surgery on 89 patients were demonstrated: mortality rate of 11,2%, a secondary amputation rate of 13,5% and improvement of 75,3%. The problems after amputation in the senium are discussed.

Age Factors↗
PubMed1979

[Effects of bacterial peritonitis on the low-pressure system in man].

The effective compliance (C) of the "low-pressure system" has been measured in ten patients with bacterial peritonitis by means of plotting pressure and volume on the first postoperative day (deltaV = 500 ml/10 min). The value of C is decreased in one group of patients, while it increases to as much as five times that of "normal patients" in the other. The cause might be a summation effect of hypovolemia, blood volume shift from intra- to extrathoracic space, and endotoxin reaction on the tone of vascular smooth muscle. The augmentation of the intravascular volume results in an increase of portal venous pressure, which is less than central venous pressure. Splanchnic pooling through increased vascular resistance does not appear.

Bacterial Infections↗