Receptor-mediated transport
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We study effects of convective transport on a chemical front wave representing a signal propagation at a simple (single layer) epithelium by means of mathematical modeling. Plug flow and laminar flow regimes were considered. We observed a nonmonotonous dependence of the propagation velocity on the ligand receptor binding constant under influence of the convective transport. If the signal propagates downstream, the region of high velocities becomes much broader and spreads over several orders of magnitude of the binding constant. When the convective transport is oriented against the propagating signal, either velocity of the traveling front wave is slowed down or the traveling front wave can stop or reverse the direction of propagation. More importantly, chemical signal in epithelial systems influenced by the convective transport can propagate almost independently of the ligand-receptor binding constant in a broad range of this parameter. Furthermore, we found that the effects of the convective transport becomes more significant in systems where either the characteristic dimension of the extracellular space is larger/comparable with the spatial extent of the ligand diffusion trafficking or the ligand-receptor binding/ligand diffusion rate ratio is high.
The fibroblast growth factor receptors (FGFRs) are important oncogenes promoting tumor progression in many types of cancer, such as breast, bladder, and lung cancer as well as multiple myeloma and rhabdomyosarcoma. However, little is known about how these receptors are internalized and down-regulated in cells. We have here applied proximity biotin labeling to identify proteins involved in FGFR4 signaling and trafficking. For this purpose we fused a mutated biotin ligase, BirA*, to the C-terminal tail of FGFR4 (FGFR4-BirA*) and the fusion protein was stably expressed in U2OS cells. Upon addition of biotin to these cells, proteins in proximity to the FGFR4-BirA* fusion protein became biotinylated and could be isolated and identified by quantitative mass spectrometry. We identified in total 291 proteins, including 80 proteins that were enriched in samples where the receptor was activated by the ligand (FGF1), among them several proteins previously found to be involved in FGFR signaling (e.g., FRS2, PLCγ, RSK2 and NCK2). Interestingly, many of the identified proteins were implicated in endosomal transport, and by precise annotation we were able to trace the intracellular pathways of activated FGFR4. Validating the data by confocal and three-dimensional structured illumination microscopy analysis, we concluded that FGFR4 uses clathrin-mediated endocytosis for internalization and is further sorted from early endosomes to the recycling compartment and the trans-Golgi network. Depletion of cells for clathrin heavy chain led to accumulation of FGFR4 at the cell surface and increased levels of active FGFR4 and PLCγ, while AKT and ERK signaling was diminished, demonstrating that functional clathrin-mediated endocytosis is required for proper FGFR4 signaling. Thus, this study reveals proteins and pathways involved in FGFR4 transport and signaling that provide possible targets and opportunities for therapeutic intervention in FGFR4 aberrant cancer.
- MeSH
- barvení a značení MeSH
- biotinylace MeSH
- endocytóza MeSH
- endozomy metabolismus MeSH
- klathrin metabolismus MeSH
- lidé MeSH
- mikroskopie metody MeSH
- nádorové buněčné linie MeSH
- receptor fibroblastových růstových faktorů, typ 4 metabolismus MeSH
- signální transdukce MeSH
- trans-Golgiho síť metabolismus MeSH
- transport proteinů MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Membrane-tethered signalling proteins such as TNFα and many EGF receptor ligands undergo shedding by the metalloproteinase ADAM17 to get released. The pseudoproteases iRhom1 and iRhom2 are important for the transport, maturation and activity of ADAM17. Yet, the structural and functional requirements to promote the transport of the iRhom-ADAM17 complex have not yet been thoroughly investigated. Utilising in silico and in vitro methods, we here map the conserved iRhom homology domain (IRHD) and provide first insights into its structure and function. By focusing on iRhom2, we identified different structural and functional factors within the IRHD. We found that the structural integrity of the IRHD is a key factor for ADAM17 binding. In addition, we identified a highly conserved motif within an unstructured region of the IRHD, that, when mutated, restricts the transport of the iRhom-ADAM17 complex through the secretory pathway in in vitro, ex vivo and in vivo systems and also increases the half-life of iRhom2 and ADAM17. Furthermore, the disruption of this IRHD motif was also reflected by changes in the yet undescribed interaction profile of iRhom2 with proteins involved in intracellular vesicle transport. Overall, we provide the first insights into the forward trafficking of iRhoms which is critical for TNFα and EGF receptor signalling.
- MeSH
- aminokyselinové motivy MeSH
- buněčné linie MeSH
- epidermální růstové faktory metabolismus MeSH
- lidé MeSH
- malá interferující RNA metabolismus MeSH
- membránové proteiny genetika metabolismus MeSH
- mutageneze MeSH
- myši inbrední C57BL MeSH
- myši MeSH
- poločas MeSH
- protein ADAM17 chemie metabolismus MeSH
- proteinové domény MeSH
- RNA interference MeSH
- signální transdukce MeSH
- TNF-alfa metabolismus MeSH
- transport proteinů MeSH
- transportní proteiny antagonisté a inhibitory genetika metabolismus MeSH
- vazba proteinů MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
To examine reciprocal or unilateral implications between two cell destruction processes, autophagy and apoptosis, in 5-Fluorouracil (5-FU)-treated tumor cells, a combination of chemical inhibitors, RNAi and genetic approaches were used. In contrast to cancer cells harboring obstructed apoptosis, either at the DISC or the mitochondrial level, p53-deficiency generated signs of autophagy deregulation upon chemotherapy. On the other, hand disruption of lysosomal function by chloroquine, caused a profound decrease in apoptotic markers appearing in response to 5-FU. DR5, which is essential for 5-FU-induced apoptosis, accumulated in lysosomes and autophagosomes upon chloroquine treatment. Since neither 3-MA, RNAi of critical autophagy regulators or inhibition of cathepsins reversed apoptosis in a similar manner, it is likely that not autophagy per se but rather correct receptor transport is an important factor for 5-FU cytotoxicity. We found that apoptosis generated by TRAIL, the cognate ligand for DR5, remained unchanged upon chloroquine lysosomal interference, indicating that 5-FU activates the receptor by a discrete mechanism. In support, depletion of membrane cholesterol or hampering cholesterol transport drastically reduced 5-FU cytotoxicity. We conclude that targeting of lysosomes by chloroquine deregulates DR5 trafficking and abrogates 5-FU- but not TRAIL-stimulated cell elimination, hence suggesting a novel mechanism for receptor activation.
- MeSH
- apoptóza MeSH
- autofagie * MeSH
- buněčná membrána metabolismus MeSH
- chlorochin chemie MeSH
- cholesterol chemie MeSH
- fagozomy MeSH
- fluoruracil chemie MeSH
- HCT116 buňky MeSH
- lidé MeSH
- ligandy MeSH
- lyzozomy metabolismus MeSH
- makrolidy chemie MeSH
- mitochondrie metabolismus MeSH
- nádorový supresorový protein p53 metabolismus MeSH
- protein TRAIL farmakologie MeSH
- protinádorové antimetabolity farmakologie MeSH
- RNA interference MeSH
- signální transdukce účinky léků MeSH
- TRAIL receptory metabolismus MeSH
- transport proteinů MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Auxin and cytokinins have been identified as key regulators of plant development. Recently, these phytohormones have been shown to interact during important developmental processes, including positioning, identity acquisition and maintenance of meristem organizing centres, regulation of balance between cell division and differentiation, and postembryonic de novo organogenesis. Here, we discuss recent advances in our understanding of the underlying molecular mechanisms at the levels of regulating metabolism, signalling, gene expression and protein stability.
- MeSH
- Arabidopsis genetika růst a vývoj metabolismus MeSH
- biologický transport genetika MeSH
- cytokininy metabolismus MeSH
- kyseliny indoloctové metabolismus MeSH
- meristém růst a vývoj MeSH
- regulace genové exprese u rostlin MeSH
- regulátory růstu rostlin metabolismus MeSH
- rostliny genetika metabolismus MeSH
- signální transdukce MeSH
- vývoj rostlin MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
- MeSH
- biologický transport MeSH
- glukosa fyziologie MeSH
- homeostáza MeSH
- inzulin fyziologie MeSH
- inzulinová rezistence MeSH
- krysa rodu rattus MeSH
- modely nemocí na zvířatech enzymologie genetika patofyziologie MeSH
- signální transdukce MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
- Publikační typ
- přehledy MeSH
Nonsense-mediated mRNA decay (NMD) is a conserved eukaryotic RNA surveillance mechanism that degrades aberrant mRNAs. NMD impairment in Arabidopsis is linked to constitutive immune response activation and enhanced antibacterial resistance, but the underlying mechanisms are unknown. Here we show that NMD contributes to innate immunity in Arabidopsis by controlling the turnover of numerous TIR domain-containing, nucleotide-binding, leucine-rich repeat (TNL) immune receptor-encoding mRNAs. Autoimmunity resulting from NMD impairment depends on TNL signaling pathway components and can be triggered through deregulation of a single TNL gene, RPS6. Bacterial infection of plants causes host-programmed inhibition of NMD, leading to stabilization of NMD-regulated TNL transcripts. Conversely, constitutive NMD activity prevents TNL stabilization and impairs plant defense, demonstrating that host-regulated NMD contributes to disease resistance. Thus, NMD shapes plant innate immunity by controlling the threshold for activation of TNL resistance pathways.
- MeSH
- Arabidopsis genetika imunologie mikrobiologie MeSH
- interakce hostitele a patogenu MeSH
- messenger RNA genetika imunologie MeSH
- nemoci rostlin genetika imunologie mikrobiologie MeSH
- nesmyslný kodon MeSH
- nonsense mediated mRNA decay * MeSH
- proteiny huseníčku genetika imunologie MeSH
- Pseudomonas syringae genetika fyziologie MeSH
- receptory imunologické genetika imunologie MeSH
- RNA-helikasy genetika imunologie MeSH
- transportní proteiny genetika imunologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Polar auxin transport plays a pivotal role in plant growth and development. PIN-FORMED (PIN) auxin efflux carriers regulate directional auxin movement by establishing local auxin maxima, minima, and gradients that drive multiple developmental processes and responses to environmental signals. Auxin has been proposed to modulate its own transport by regulating subcellular PIN trafficking via processes such as clathrin-mediated PIN endocytosis and constitutive recycling. Here, we further investigated the mechanisms by which auxin affects PIN trafficking by screening auxin analogs and identified pinstatic acid (PISA) as a positive modulator of polar auxin transport in Arabidopsis (Arabidopsis thaliana). PISA had an auxin-like effect on hypocotyl elongation and adventitious root formation via positive regulation of auxin transport. PISA did not activate SCFTIR1/AFB signaling and yet induced PIN accumulation at the cell surface by inhibiting PIN internalization from the plasma membrane. This work demonstrates PISA to be a promising chemical tool to dissect the regulatory mechanisms behind subcellular PIN trafficking and auxin transport.
- MeSH
- Arabidopsis účinky léků metabolismus MeSH
- biologický transport účinky léků MeSH
- buněčná membrána účinky léků metabolismus MeSH
- endocytóza * účinky léků MeSH
- fenotyp MeSH
- fenylacetáty farmakologie MeSH
- gravitropismus účinky léků MeSH
- hypokotyl účinky léků růst a vývoj MeSH
- kořeny rostlin účinky léků růst a vývoj MeSH
- kyseliny indoloctové metabolismus MeSH
- proteiny huseníčku metabolismus MeSH
- signální transdukce MeSH
- výhonky rostlin metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
