Down-regulation of Drosophila Egf-r mRNA levels following hyperactivated receptor signaling

Development ◽  
1994 ◽  
Vol 120 (9) ◽  
pp. 2593-2600 ◽  
Author(s):  
M.A. Sturtevant ◽  
J.W. O'Neill ◽  
E. Bier
Keyword(s):  
Mrna Stability ◽  
Egf Receptor ◽  
Receptor Internalization ◽  
Ras Signaling ◽  
Mrna Levels ◽  
Down Regulation ◽  
Feedback Mechanisms ◽  
Receptor Complexes ◽  
Surface Receptor ◽  
Receptor Pool

Internalization of ligand-receptor complexes is a well-documented mechanism for limiting the duration and magnitude of a signaling event. In the case of the EGF-Receptor (EGF-R), exposure to EGF or TGF-alpha results in internalization of up to 95% of the surface receptor pool within 5 minutes of exposure to ligand. In this report, we show that levels of Drosophila Egf-r mRNA are strongly down-regulated in epidermal cells likely to have recently undergone high levels of EGF-R signaling. The cells in which Egf-r mRNA levels are down-regulated express the rhomboid gene, which is thought to locally amplify EGF-R signaling. Widespread Egf-r mRNA down-regulation can be induced by ubiquitous expression of rhomboid or by eliminating the Gap1 gene. These results suggest that cells engaged in intense EGF-R/RAS signaling limit the duration of the signal through a combination of short-acting negative feedback mechanisms such as receptor internalization followed by a longer lasting reduction in receptor transcript levels. Control of Egf-r mRNA levels by altering transcription or mRNA stability is a new tier of regulation to be considered in analysis of EGF-R signaling during development.

Both proteasomes and lysosomes degrade the activated erythropoietin receptor

Blood ◽  
2005 ◽  
Vol 105 (2) ◽  
pp. 600-608 ◽  
Author(s):  
Pierre Walrafen ◽  
Frédérique Verdier ◽  
Zahra Kadri ◽  
Stany Chrétien ◽  
Catherine Lacombe ◽  
...  
Keyword(s):  
Cell Surface ◽  
Intracellular Signaling ◽  
Erythropoietin Receptor ◽  
Janus Kinase ◽  
Receptor Internalization ◽  
Down Regulation ◽  
Epo Receptor ◽  
Receptor Complexes ◽  
Regulation Mechanisms ◽  
Rapid Activation

AbstractActivation of the erythropoietin receptor (EpoR) after Epo binding is very transient because of the rapid activation of strong down-regulation mechanisms that quickly decrease Epo sensitivity of the cells. Among these down-regulation mechanisms, receptor internalization and degradation are probably the most efficient. Here, we show that the Epo receptor was rapidly ubiquitinated after ligand stimulation and that the C-terminal part of the Epo receptor was degraded by the proteasomes. Both ubiquitination and receptor degradation by the proteasomes occurred at the cell surface and required Janus kinase 2 (Jak2) activation. Moreover, Epo-EpoR complexes were rapidly internalized and targeted to the lysosomes for degradation. Neither Jak2 nor proteasome activities were required for internalization. In contrast, Jak2 activation was necessary for lysosome targeting of the Epo-EpoR complexes. Blocking Jak2 with the tyrphostin AG490 led to some recycling of internalized Epo-Epo receptor complexes to the cell surface. Thus, activated Epo receptors appear to be quickly degraded after ubiquitination by 2 proteolytic systems that proceed successively: the proteasomes remove part of the intracellular domain at the cell surface, and the lysosomes degrade the remaining part of the receptor-hormone complex. The efficiency of these processes probably explains the short duration of intracellular signaling activated by Epo.

scholarly journals Ligand-mediated internalization, recycling, and downregulation of the epidermal growth factor receptor in vivo.

1989 ◽  
Vol 109 (6) ◽  
pp. 2741-2749 ◽  
Author(s):  
W H Lai ◽  
P H Cameron ◽  
I Wada ◽  
J J Doherty ◽  
D G Kay ◽  
...  
Keyword(s):  
Body Weight ◽  
Western Blotting ◽  
Egf Receptor ◽  
Growth Factor Receptor ◽  
Receptor Occupancy ◽  
Liver Parenchyma ◽  
Receptor Internalization ◽  
Scatchard Analysis ◽  
Surface Receptor

EGF receptor internalization, recycling,a nd downregulation were evaluated in liver parenchyma as a function of increasing doses of injected EGF. The effect of ligand occupancy in vivo on the kinetics and extent of internalization was studied with changes in the receptor content of isolated plasmalemma and endosome fractions evaluated by direct binding, Scatchard analysis, and Western blotting. For all doses of injected EGF, receptor was lost from the plasmalemma and accumulated in endosomes in a time- and dose-dependent fashion. However, at doses of injected EGF equivalent to less than or equal to 50% surface receptor occupancy (i.e., less than or equal to 1 microgram/100 g body weight), receptor levels returned by 120 min to initial values. This return was resistant to cycloheximide and therefore did not represent newly synthesized receptor. Neither was the return due to replenishment by an intracellular pool of low-affinity receptors as such a pool could not be detected by Scatchard analysis or Western blotting. Therefore, receptor return was due to the recycling of previously internalized receptor. At doses of injected EGF greater than 50% receptor occupancy, net receptor loss-i.e., downregulation-was observed by evaluating the receptor content of total particulate fractions of liver homogenates. At the higher saturating doses of injected EGF (5 and 10 micrograms/100 g body weight), the majority of surface receptor content was lost by 15 min and remained low for at least an additional 105 min. As the kinetics of ligand clearance from the circulation and liver parenchyma were similar for all doses of EGF injected, then the ligand-mediated regulation of surface receptor content and downregulation were not a result of a prolonged temporal interaction of ligand with receptor. Rather, the phenomena must be a consequence of the absolute concentrations of EGF interacting with receptor at the cell surface and/or in endosomes.

scholarly journals Down-Regulation of the Proteoglycan Decorin Fills in the Tumor-Promoting Phenotype of Ionizing Radiation-Induced Senescent Human Breast Stromal Fibroblasts

Cancers ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1987
Author(s):  
Eleni Mavrogonatou ◽  
Adamantia Papadopoulou ◽  
Asimina Fotopoulou ◽  
Stathis Tsimelis ◽  
Heba Bassiony ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Ionizing Radiation ◽  
Cancer Progression ◽  
Breast Cancer Cell Lines ◽  
Phenotypic Trait ◽  
Mrna Levels ◽  
Human Breast ◽  
Down Regulation ◽  
Stromal Fibroblasts ◽  
Poor Prognostic Factor

Down-regulation of the small leucine-rich proteoglycan decorin in the stroma is considered a poor prognostic factor for breast cancer progression. Ionizing radiation, an established treatment for breast cancer, provokes the premature senescence of the adjacent to the tumor stromal fibroblasts. Here, we showed that senescent human breast stromal fibroblasts are characterized by the down-regulation of decorin at the mRNA and protein level, as well as by its decreased deposition in the pericellular extracellular matrix in vitro. Senescence-associated decorin down-regulation is a long-lasting process rather than an immediate response to γ-irradiation. Growth factors were demonstrated to participate in an autocrine manner in decorin down-regulation, with bFGF and VEGF being the critical mediators of the phenomenon. Autophagy inhibition by chloroquine reduced decorin mRNA levels, while autophagy activation using the mTOR inhibitor rapamycin enhanced decorin transcription. Interestingly, the secretome from a series of both untreated and irradiated human breast cancer cell lines with different molecular profiles inhibited decorin expression in young and senescent stromal fibroblasts, which was annulled by SU5402, a bFGF and VEGF inhibitor. The novel phenotypic trait of senescent human breast stromal fibroblasts revealed here is added to their already described cancer-promoting role via the formation of a tumor-permissive environment.

Nitric oxide increases IL-8 gene transcription and mRNA stability to enhance IL-8 gene expression in lung epithelial cells

2004 ◽  
Vol 287 (4) ◽  
pp. L764-L773 ◽  
Author(s):  
Loretta Sparkman ◽  
Vijayakumar Boggaram
Keyword(s):  
Gene Expression ◽  
Nitric Oxide ◽  
Protein Kinase ◽  
Epithelial Cells ◽  
Gene Transcription ◽  
Mrna Stability ◽  
Inflammatory Responses ◽  
Lung Epithelial Cells ◽  
Mrna Levels ◽  
Lung Epithelial

Interleukin (IL)-8, a C-X-C chemokine, is a potent chemoattractant and an activator for neutrophils, T cells, and other immune cells. The airway and respiratory epithelia play important roles in the initiation and modulation of inflammatory responses via production of cytokines and surfactant. The association between elevated levels of nitric oxide (NO) and IL-8 in acute lung injury associated with sepsis, acute respiratory distress syndrome, respiratory syncytial virus infection in infants, and other inflammatory diseases suggested that NO may play important roles in the control of IL-8 gene expression in the lung. We investigated the role of NO in the control of IL-8 gene expression in H441 lung epithelial cells. We found that a variety of NO donors significantly induced IL-8 mRNA levels, and the increase in IL-8 mRNA was associated with an increase in IL-8 protein. NO induction of IL-8 mRNA was due to increases in IL-8 gene transcription and mRNA stability. NO induction of IL-8 mRNA levels was not inhibited by 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one and KT-5823, inhibitors of soluble guanylate cyclase and protein kinase G, respectively, and 8-bromo-cGMP did not increase IL-8 mRNA levels. This indicated that NO induces IL-8 mRNA levels independently of changes in the intracellular cGMP levels. NO induction of IL-8 mRNA was significantly reduced by inhibitors of extracellular regulated kinase and protein kinase C. IL-8 induction by NO was also reduced by hydroxyl radical scavengers such as dimethyl sulfoxide and dimethylthiourea, indicating the involvement of hydroxyl radicals in the induction process. NO induction of IL-8 gene expression could be a significant contributing factor in the initiation and induction of inflammatory response in the respiratory epithelium.

scholarly journals E3-13.7 Integral Membrane Proteins Encoded by Human Adenoviruses Alter Epidermal Growth Factor Receptor Trafficking by Interacting Directly with Receptors in Early Endosomes

2000 ◽  
Vol 11 (10) ◽  
pp. 3559-3572 ◽  
Author(s):  
Denise Crooks ◽  
Song Jae Kil ◽  
J. Michael McCaffery ◽  
Cathleen Carlin
Keyword(s):  
Plasma Membrane ◽  
Membrane Protein ◽  
Viral Protein ◽  
Egf Receptor ◽  
Receptor Trafficking ◽  
Model Systems ◽  
Egf Receptors ◽  
Down Regulation ◽  
Human Adenoviruses ◽  
Early Endosomes

Animal cell viruses provide valuable model systems for studying many normal cellular processes, including membrane protein sorting. The focus of this study is an integral membrane protein encoded by the E3 transcription region of human adenoviruses called E3-13.7, which diverts recycling EGF receptors to lysosomes without increasing the rate of receptor internalization or intrinsic receptor tyrosine kinase activity. Although E3-13.7 can be found on the plasma membrane when it is overexpressed, its effect on EGF receptor trafficking suggests that the plasma membrane is not its primary site of action. Using cell fractionation and immunocytochemical experimental approaches, we now report that the viral protein is located predominantly in early endosomes and limiting membranes of endosome-to-lysosome transport intermediates called multivesicular endosomes. We also demonstrate that E3-13.7 physically associates with EGF receptors undergoing E3-13.7–mediated down-regulation in early endosomes. Receptor–viral protein complexes then dissociate, and EGF receptors proceed to lysosomes, where they are degraded, while E3-13.7 is retained in endosomes. We conclude that E3-13.7 is a resident early endocytic protein independent of EGF receptor expression, because it has identical intracellular localization in mouse cells lacking endogenous receptors and cells expressing a human cytomegalovirus-driven receptor cDNA. Finally, we demonstrate that EGF receptor residues 675–697 are required for E3-13.7–mediated down-regulation. Interestingly, this sequence includes a known EGF receptor leucine-based lysosomal sorting signal used during ligand-induced trafficking, which is also conserved in the viral protein. E3-13.7, therefore, provides a novel model system for determining the molecular basis of selective membrane protein transport in the endocytic pathway. Our studies also suggest new paradigms for understanding EGF receptor sorting in endosomes and adenovirus pathogenesis.

scholarly journals Parathyroid hormone induces bone formation in phosphorylation-deficient PTHR1 knockin mice

2012 ◽  
Vol 302 (10) ◽  
pp. E1183-E1188 ◽  
Author(s):  
Nabanita S. Datta ◽  
Tareq A. Samra ◽  
Abdul B. Abou-Samra
Keyword(s):  
Parathyroid Hormone ◽  
Bone Formation ◽  
Physiological Role ◽  
Receptor Internalization ◽  
Mineral Density ◽  
Trabecular Thickness ◽  
Receptor Complexes ◽  
Diaphyseal Bone ◽  
Anabolic Action

Activation of G protein-coupled receptors by agonists leads to receptor phosphorylation, internalization of ligand receptor complexes, and desensitization of hormonal response. The role of parathyroid hormone (PTH) receptor 1, PTHR1, is well characterized and known to regulate cellular responsiveness in vitro. However, the role of PTHR1 phosphorylation in bone formation is yet to be investigated. We have previously demonstrated that impaired internalization and sustained cAMP stimulation of phosphorylation-deficient (PD) PTHR1 leads to exaggerated cAMP response to subcutaneous PTH infusion in a PD knockin mouse model. To understand the physiological role of receptor internalization on PTH bone anabolic action, we examined bone parameters of wild-type (WT) and PD knockin female and male mice following PTH treatment. We found a decrease in total and diaphyseal bone mineral density in female but not in male PD mice compared with WT controls at 3–6 mo of age. This effect was attenuated at older age groups. PTH administration displayed increased bone volume and trabecular thickness in the vertebrae and distal femora of both WT and PD animals. These results suggest that PTHR1 phosphorylation does not play a major role in the anabolic action of PTH.

A new function for a phosphotyrosine phosphatase: linking GRB2-Sos to a receptor tyrosine kinase

1994 ◽  
Vol 14 (1) ◽  
pp. 509-517
Author(s):  
W Li ◽  
R Nishimura ◽  
A Kashishian ◽  
A G Batzer ◽  
W J Kim ◽  
...  
Keyword(s):  
Growth Factor ◽  
Intracellular Signaling ◽  
Egf Receptor ◽  
Tyrosine Phosphatase ◽  
Growth Factor Receptors ◽  
Ras Signaling ◽  
Pdgf Receptor ◽  
Phosphotyrosine Phosphatase ◽  
Src Homology

Autophosphorylated growth factor receptors provide binding sites for the src homology 2 domains of intracellular signaling molecules. In response to epidermal growth factor (EGF), the activated EGF receptor binds to a complex containing the signaling protein GRB2 and the Ras guanine nucleotide-releasing factor Sos, leading to activation of the Ras signaling pathway. We have investigated whether the platelet-derived growth factor (PDGF) receptor binds GRB2-Sos. In contrast with the EGF receptor, the GRB2 does not bind to the PDGF receptor directly. Instead, PDGF stimulation induces the formation of a complex containing GRB2; 70-, 80-, and 110-kDa tyrosine-phosphorylated proteins; and the PDGF receptor. Moreover, GRB2 binds directly to the 70-kDa protein but not to the PDGF receptor. Using a panel of PDGF beta-receptor mutants with altered tyrosine phosphorylation sites, we identified Tyr-1009 in the PDGF receptor as required for GRB2 binding. Binding is inhibited by a phosphopeptide containing a YXNX motif. The protein tyrosine phosphatase Syp/PTP1D/SHPTP2/PTP2C is approximately 70 kDa, binds to the PDGF receptor via Tyr-1009, and contains several YXNX sequences. We found that the 70-kDa protein that binds to the PDGF receptor and to GRB2 comigrates with Syp and is recognized by anti-Syp antibodies. Furthermore, both GRB2 and Sos coimmunoprecipitate with Syp from lysates of PDGF-stimulated cells, and GRB2 binds directly to tyrosine-phosphorylated Syp in vitro. These results indicate that GRB2 interacts with different growth factor receptors by different mechanisms and the cytoplasmic phosphotyrosine phosphatase Syp acts as an adapter between the PDGF receptor and the GRB2-Sos complex.

Emergence of salivary gland cell lineage diversity suggests a role for androgen-independent epidermal growth factor receptor signaling

1995 ◽  
Vol 108 (6) ◽  
pp. 2205-2212
Author(s):  
E.M. Durban ◽  
P.G. Nagpala ◽  
P.D. Barreto ◽  
E. Durban
Keyword(s):  
Salivary Gland ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Egf Receptor ◽  
Cell Lineage ◽  
Mrna Levels ◽  
Receptor Signaling ◽  
Receptor Mrna ◽  
Epidermal Growth ◽  
Lineage Diversity

Diversity of cell lineages within glandular organs is generated postnatally by differentiation of committed progenitor cells. Fundamental regulatory aspects of this process are not understood. The mouse submandibular salivary gland (SSG) served as model to assess the role of epidermal growth factor (EGF) receptor signaling during emergence of cell lineage diversity. Temporal fluctuations in EGF receptor mRNA levels coincident with crucial differentiative cell lineage transitions were revealed by RNase protection analyses. Between days 2 and 5, when proacinar cells are maturing and striated duct cells emerge, EGF receptor mRNA levels were highest and all differentiating cells exhibited EGF receptor immunoreactivity. EGF receptor mRNA levels then declined sharply and immunoreactivity became confined to ductal cells. At day 11 in male mice, and days 11 and 16 in females, a second increase in EGF receptor mRNA was detected coincident with emergence of granular convoluted tubule (GCT) cells. With completion of androgen-dependent GCT cell differentiation at the onset of puberty, EGF receptor mRNA levels and intensity of immunoreactivity decreased. Androgen effects on EGF receptor mRNA or immunoreactivity could not be detected. These temporally distinct patterns of EGF receptor expression suggest that this signaling pathway is a mechanism of potential importance in emergence of cell lineage diversity in a glandular organ.

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