Expression in mammalian cell cultures reveals interdependent, but distinct, functions for Star and Rhomboid proteins in the processing of the Drosophila transforming-growth-factor-α homologue Spitz

2002 ◽  
Vol 363 (2) ◽  
pp. 347-352 ◽  
Author(s):  
John C. PASCALL ◽  
Jane E. LUCK ◽  
Kenneth D. BROWN
Keyword(s):  
Growth Factor ◽  
Cell Surface ◽  
Mammalian Cells ◽  
Egf Receptor ◽  
Transforming Growth Factor ◽  
Surface Protein ◽  
Immunocytochemical Staining ◽  
Transforming Growth Factor Α ◽  
Mammalian Cell Cultures ◽  
Factor Α

We report here distinct interdependent functions for two proteins, Star and Rhomboid, that are key determinants of the epidermal-growth-factor (EGF)-receptor signalling pathway in Drosophila. When we expressed the Drosophila EGF-receptor ligand Spitz in mammalian cells, the protein failed to traffic to the plasma membrane, as assessed by either cell-surface protein biotinylation or immunocytochemical staining. However, when we co-expressed Star with Spitz, trafficking of Spitz to the cell surface could be demonstrated. Only when we co-expressed Spitz, Star and Rhomboid could the release of soluble Spitz protein into the medium be shown. Taken together, our results indicate that Star is required for the intracellular trafficking of Spitz, and that Rhomboid is essential for the release of soluble Spitz protein from cells.

EGF-Receptor binding peptides from transforming growth factor α (TGFα): Evidence for a multi-domain interaction

Peptides ◽  
1992 ◽  
pp. 410-412
Author(s):  
Donna E. Davies ◽  
Audrey Richter ◽  
J. Wayne Conlan ◽  
Clement Higginbotham ◽  
Michael E. Ward ◽  
...  
Keyword(s):  
Growth Factor ◽  
Receptor Binding ◽  
Egf Receptor ◽  
Transforming Growth Factor ◽  
Domain Interaction ◽  
Transforming Growth Factor Α ◽  
Binding Peptides ◽  
Factor Α

scholarly journals EGF and amphiregulin differentially regulate Cbl recruitment to endosomes and EGF receptor fate

2008 ◽  
Vol 410 (3) ◽  
pp. 585-594 ◽  
Author(s):  
Kathryn A. Stern ◽  
Trenton L. Place ◽  
Nancy L. Lill
Keyword(s):  
Growth Factor ◽  
Egf Receptor ◽  
Transforming Growth Factor ◽  
Partial Agonist ◽  
Ectopic Expression ◽  
Growth Factor Receptor ◽  
Receptor Activation ◽  
Transforming Growth Factor Α ◽  
Molecular Events ◽  
Factor Α

EGF-R [EGF (epidermal growth factor) receptor] ligands can promote or inhibit cell growth. The biological outcome of receptor activation is dictated, at least in part, by ligand-specified patterns of endocytic trafficking. EGF-R trafficking downstream of the ligands EGF and TGF-α (transforming growth factor-α) has been investigated extensively. However, less is known about EGF-R fates induced by the ligands BTC (betacellulin) and AR (amphiregulin). We undertook comparative analyses to identify ligand-specific molecular events that regulate EGF-R trafficking and degradation. EGF (17 nM) and BTC (8.5 nM) induced significant EGF-R degradation, with or without ectopic expression of the ubiquitin ligase Cbl. Human recombinant AR (17 nM) failed to affect receptor degradation in either case. Notably, levels of ligand-induced EGF-R ubiquitination did not correlate strictly with receptor degradation. Dose–response experiments revealed that AR at a saturating concentration was a partial agonist at the EGF-R, with approx. 40% efficacy (relative to EGF) at inducing receptor tyrosine phosphorylation, ubiquitination and association with Cbl. EGF-R down-regulation and degradation also were compromised upon cell stimulation with AR (136 nM). These outcomes correlated with decreased degradation of the Cbl substrate and internalization inhibitor hSprouty2. Downstream of the hSprouty2 checkpoint in AR-stimulated cells, Cbl-free EGF-R was incorporated into endosomes from which Cbl–EGF-R complexes were excluded. Our results suggest that the AR-specific EGF-R fate results from decreased hSprouty2 degradation and reduced Cbl recruitment to underphosphorylated EGF-R, two effects that impair EGF-R trafficking to lysosomes.

Transforming growth factor-α: receptor binding and action on DNA synthesis in the sheep mammary gland

1995 ◽  
Vol 144 (1) ◽  
pp. 165-171 ◽  
Author(s):  
C D Moorby ◽  
J A Taylor ◽  
I A Forsyth
Keyword(s):  
Growth Factor ◽  
Dna Synthesis ◽  
Binding Sites ◽  
Egf Receptor ◽  
Transforming Growth Factor ◽  
Mammary Development ◽  
Transforming Growth Factor Α ◽  
Pregnant Sheep ◽  
Igf I ◽  
Factor Α

Abstract Microsome fractions prepared from the mammary glands of non-pregnant, pregnant and lactating sheep have been used to study binding of 125I-labelled transforming growth factor-α (TGF-α). Binding was dependent on microsomal protein concentration, time and temperature. It showed the characteristics of an epidermal growth factor (EGF) receptor, being displaced by TGF-α and EGF, but not by insulin or IGF-I. The non-linear curve fitting program LIGAND was used to determine affinity and number of binding sites. A single class of high-affinity binding sites was found. The apparent dissociation constant (Kd) was similar in all physiological states (2·43±0·27 mol/l × 10−10, n=23). Numbers of binding sites were lower in late-pregnant (20 weeks) and lactating sheep (14·07± 2·45 fmol/mg protein, n=10) than in non-pregnant, 10-or 15-week pregnant sheep (43·04±5·93 fmol/mg protein, n=13). DNA synthesis by mammary alveolar epithelial cells cultured on collagen gels was increased twofold by TGF-a (maximum response at 10 μg/l; 1·8 nmol/l) but not by EGF. Cells derived from 15- to 20-week pregnant sheep responded significantly to TGF-α on day 3 of culture, but the response was delayed to day 4–5 of culture in cells from other physiological states. Dose–response was not significantly affected. TGF-α and IGF-I produced an additive effect on DNA synthesis. Oestradiol (10−12 to 10−9 m), a potential stimulator of the TGF-α gene, did not stimulate DNA synthesis alone, or in combination with IGF-I. It is concluded that growth factors acting via the EGF receptor play a role in ruminant mammary development, but whether they mediate oestradiol effects remains unresolved. Journal of Endocrinology (1995) 144, 165–171

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