Regulation of body length and male tail ray pattern formation of Caenorhabditis elegans by a member of TGF-beta family.
(9/277092)
We have identified a new member of the TGF-beta superfamily, CET-1, from Caenorhabditis elegans, which is expressed in the ventral nerve cord and other neurons. cet-1 null mutants have shortened bodies and male tail abnormal phenotype resembling sma mutants, suggesting cet-1, sma-2, sma-3 and sma-4 share a common pathway. Overexpression experiments demonstrated that cet-1 function requires wild-type sma genes. Interestingly, CET-1 appears to affect body length in a dose-dependent manner. Heterozygotes for cet-1 displayed body lengths ranging between null mutant and wild type, and overexpression of CET-1 in wild-type worms elongated body length close to lon mutants. In male sensory ray patterning, lack of cet-1 function results in ray fusions. Epistasis analysis revealed that mab-21 lies downstream and is negatively regulated by the cet-1/sma pathway in the male tail. Our results show that cet-1 controls diverse biological processes during C. elegans development probably through different target genes. (+info)
Sonic hedgehog signaling by the patched-smoothened receptor complex.
(10/277092)
BACKGROUND: The Hedgehog (Hh) family of secreted proteins is involved in a number of developmental processes as well as in cancer. Genetic and biochemical data suggest that the Sonic hedgehog (Shh) receptor is composed of at least two proteins: the tumor suppressor protein Patched (Ptc) and the seven-transmembrane protein Smoothened (Smo). RESULTS: Using a biochemical assay for activation of the transcription factor Gli, a downstream component of the Hh pathway, we show here that Smo functions as the signaling component of the Shh receptor, and that this activity can be blocked by Ptc. The inhibition of Smo by Ptc can be relieved by the addition of Shh. Furthermore, oncogenic forms of Smo are insensitive to Ptc repression in this assay. Mapping of the Smo domains required for binding to Ptc and for signaling revealed that the Smo-Ptc interaction involves mainly the amino terminus of Smo, and that the third intracellular loop and the seventh transmembrane domain are required for signaling. CONCLUSIONS: These data demonstrate that Smo is the signaling component of a multicomponent Hh receptor complex and that Ptc is a ligand-regulated inhibitor of Smo. Different domains of Smo are involved in Ptc binding and activation of a Gli reporter construct. The latter requires the third intracellular loop and the seventh transmembrane domain of Smo, regions often involved in coupling to G proteins. No changes in the levels of cyclic AMP or calcium associated with such pathways could be detected following receptor activation, however. (+info)
A Drosophila TNF-receptor-associated factor (TRAF) binds the ste20 kinase Misshapen and activates Jun kinase.
(11/277092)
Two families of protein kinases that are closely related to Ste20 in their kinase domain have been identified - the p21-activated protein kinase (Pak) and SPS1 families [1-3]. In contrast to Pak family members, SPS1 family members do not bind and are not activated by GTP-bound p21Rac and Cdc42. We recently placed a member of the SPS1 family, called Misshapen (Msn), genetically upstream of the c-Jun amino-terminal (JNK) mitogen-activated protein (MAP) kinase module in Drosophila [4]. The failure to activate JNK in Drosophila leads to embryonic lethality due to the failure of these embryos to stimulate dorsal closure [5-8]. Msn probably functions as a MAP kinase kinase kinase kinase in Drosophila, activating the JNK pathway via an, as yet, undefined MAP kinase kinase kinase. We have identified a Drosophila TNF-receptor-associated factor, DTRAF1, by screening for Msn-interacting proteins using the yeast two-hybrid system. In contrast to the mammalian TRAFs that have been shown to activate JNK, DTRAF1 lacks an amino-terminal 'Ring-finger' domain, and overexpression of a truncated DTRAF1, consisting of only its TRAF domain, activates JNK. We also identified another DTRAF, DTRAF2, that contains an amino-terminal Ring-finger domain. Msn specifically binds the TRAF domain of DTRAF1 but not that of DTRAF2. In Drosophila, DTRAF1 is thus a good candidate for an upstream molecule that regulates the JNK pathway by interacting with, and activating, Msn. Consistent with this idea, expression of a dominant-negative Msn mutant protein blocks the activation of JNK by DTRAF1. Furthermore, coexpression of Msn with DTRAF1 leads to the synergistic activation of JNK. We have extended some of these observations to the mammalian homolog of Msn, Nck-interacting kinase (NIK), suggesting that TRAFs also play a critical role in regulating Ste20 kinases in mammals. (+info)
Identification of APC2, a homologue of the adenomatous polyposis coli tumour suppressor.
(12/277092)
The adenomatous polyposis coli (APC) tumour-suppressor protein controls the Wnt signalling pathway by forming a complex with glycogen synthase kinase 3beta (GSK-3beta), axin/conductin and betacatenin. Complex formation induces the rapid degradation of betacatenin. In colon carcinoma cells, loss of APC leads to the accumulation of betacatenin in the nucleus, where it binds to and activates the Tcf-4 transcription factor (reviewed in [1] [2]). Here, we report the identification and genomic structure of APC homologues. Mammalian APC2, which closely resembles APC in overall domain structure, was functionally analyzed and shown to contain two SAMP domains, both of which are required for binding to conductin. Like APC, APC2 regulates the formation of active betacatenin-Tcf complexes, as demonstrated using transient transcriptional activation assays in APC -/- colon carcinoma cells. Human APC2 maps to chromosome 19p13.3. APC and APC2 may therefore have comparable functions in development and cancer. (+info)
Intracellular signalling: PDK1--a kinase at the hub of things.
(13/277092)
Phosphoinositide-dependent kinase 1 (PDK1) is at the hub of many signalling pathways, activating PKB and PKC isoenzymes, as well as p70 S6 kinase and perhaps PKA. PDK1 action is determined by colocalization with substrate and by target site availability, features that may enable it to operate in both resting and stimulated cells. (+info)
Ultrabithorax function in butterfly wings and the evolution of insect wing patterns.
(14/277092)
BACKGROUND: . The morphological and functional evolution of appendages has played a critical role in animal evolution, but the developmental genetic mechanisms underlying appendage diversity are not understood. Given that homologous appendage development is controlled by the same Hox gene in different organisms, and that Hox genes are transcription factors, diversity may evolve from changes in the regulation of Hox target genes. Two impediments to understanding the role of Hox genes in morphological evolution have been the limited number of organisms in which Hox gene function can be studied and the paucity of known Hox-regulated target genes. We have therefore analyzed a butterfly homeotic mutant 'Hindsight', in which portions of the ventral hindwing pattern are transformed to ventral forewing identity, and we have compared the regulation of target genes by the Ultrabithorax (Ubx) gene product in Lepidopteran and Dipteran hindwings. RESULTS: . We show that Ubx gene expression is lost from patches of cells in developing Hindsight hindwings, correlating with changes in wing pigmentation, color pattern elements, and scale morphology. We use this mutant to study how regulation of target genes by Ubx protein differs between species. We find that several Ubx-regulated genes in the Drosophila haltere are not repressed by Ubx in butterfly hindwings, but that Distal-less (Dll) expression is regulated by Ubx in a unique manner in butterflies. CONCLUSIONS: . The morphological diversification of insect hindwings has involved the acquisition of different sets of target genes by Ubx in different lineages. Changes in Hox-regulated target gene sets are, in general, likely to underlie the morphological divergence of homologous structures between animals. (+info)
Molecular cloning and epitope analysis of the peanut allergen Ara h 3.
(15/277092)
Peanut allergy is a significant IgE-mediated health problem because of the increased prevalence, potential severity, and chronicity of the reaction. Following our characterization of the two peanut allergens Ara h 1 and Ara h 2, we have isolated a cDNA clone encoding a third peanut allergen, Ara h 3. The deduced amino acid sequence of Ara h 3 shows homology to 11S seed-storage proteins. The recombinant form of this protein was expressed in a bacterial system and was recognized by serum IgE from approximately 45% of our peanut-allergic patient population. Serum IgE from these patients and overlapping, synthetic peptides were used to map the linear, IgE-binding epitopes of Ara h 3. Four epitopes, between 10 and 15 amino acids in length, were found within the primary sequence, with no obvious sequence motif shared by the peptides. One epitope is recognized by all Ara h 3-allergic patients. Mutational analysis of the epitopes revealed that single amino acid changes within these peptides could lead to a reduction or loss of IgE binding. By determining which amino acids are critical for IgE binding, it might be possible to alter the Ara h 3 cDNA to encode a protein with a reduced IgE-binding capacity. These results will enable the design of improved diagnostic and therapeutic approaches for food-hypersensitivity reactions. (+info)
TIF1gamma, a novel member of the transcriptional intermediary factor 1 family.
(16/277092)
We report the cloning and characterization of a novel member of the Transcriptional Intermediary Factor 1 (TIF1) gene family, human TIF1gamma. Similar to TIF1alpha and TIF1beta, the structure of TIF1beta is characterized by multiple domains: RING finger, B boxes, Coiled coil, PHD/TTC, and bromodomain. Although structurally related to TIF1alpha and TIF1beta, TIF1gamma presents several functional differences. In contrast to TIF1alpha, but like TIF1beta, TIF1 does not interact with nuclear receptors in yeast two-hybrid or GST pull-down assays and does not interfere with retinoic acid response in transfected mammalian cells. Whereas TIF1alpha and TIF1beta were previously found to interact with the KRAB silencing domain of KOX1 and with the HP1alpha, MODI (HP1beta) and MOD2 (HP1gamma) heterochromatinic proteins, suggesting that they may participate in a complex involved in heterochromatin-induced gene repression, TIF1gamma does not interact with either the KRAB domain of KOX1 or the HP1 proteins. Nevertheless, TIF1gamma, like TIF1alpha and TIF1beta, exhibits a strong silencing activity when tethered to a promoter. Since deletion of a novel motif unique to the three TIF1 proteins, called TIF1 signature sequence (TSS), abrogates transcriptional repression by TIF1gamma, this motif likely participates in TIF1 dependent repression. (+info)