Differential activation of Epidermal Growth Factor (EGF) receptor downstream signaling pathways by betacellulin and EGF

Department of Medicine and Molecular Science, Gunma University Graduate School of Medicine, Gunma 371-8511, Japan. .
Endocrinology (Impact Factor: 4.64). 10/2004; 145(9):4232-43. DOI: 10.1210/en.2004-0401
Source: PubMed

ABSTRACT To determine the downstream signaling pathways regulated by betacellulin (BTC) in comparison with epidermal growth factor (EGF), we used Chinese hamster ovary cells overexpressing the human EGF receptor (ErbB1/EGFR). The overall time-dependent activation of EGFR autophosphorylation was identical in cells treated with 1 nm BTC or 1.5 nm EGF. Analysis of site-specific EGFR phosphorylation demonstrated that the BTC and EGF tyrosine phosphorylation of Y1086 was not significantly different. In contrast, the autophosphorylation of Y1173 was markedly reduced in BTC-stimulated cells, compared with EGF stimulation that directly correlated with a reduced BTC stimulation of Shc tyrosine phosphorylation, Ras, and Raf-1 activation. On the other hand, Y1068 phosphorylation was significantly increased after BTC stimulation, compared with EGF in parallel with a greater extent of Erk phosphorylation. Expression of a dominant interfering MEK kinase 1 (MEKK1) and Y1068F EGFR more efficiently blocked the enhanced Erk activation by BTC, compared with EGF. Interestingly BTC had a greater inhibitory effect on apoptosis, compared with EGF, and expression of Y1068F EGFR abolished this enhanced inhibitory effect. Together, these data indicated that although BTC and EGF share overlapping signaling properties, the ability of BTC to enhance Erk activation occurs independent of Ras. The increased BTC activation results from a greater extent of Y1068 EGFR tyrosine phosphorylation and subsequent increased recruitment of the Grb2-MEKK1 complex to the plasma membrane, compared with EGF stimulation. The increased Erk activation by BTC associated with antiapoptotic function.

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    • "BTC has anti-apoptotic effects on intestinal epithelial cells Western blots carried out employing protein extracts from the small intestine epithelium revealed markedly lower levels of activated caspase-3 in samples from BTC-tg mice, as compared to control mice (Fig. 3D). This result confirms previous observations indicating that BTC has stronger anti-apoptotic actions as compared to EGF [26]. Furthermore, this finding suggests that BTC overexpression creates an anti-apoptotic environment in the gut epithelium, supporting the survival of newly formed adenomas. "
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    ABSTRACT: We employed transgenic mice overexpressing betacellulin (BTC) to study its effects in the gut. BTC stimulated crypt cell proliferation and markedly increased intestinal size, while the crypt-villus architecture was preserved. Introduction of a dominant negative epidermal growth factor receptor (EGFR) completely abolished the intestinal hyperplasia. BTC increased polyp multiplicity but did not change the mean size or the histological quality of intestinal polyps in Apc(+/Min) mice. Analysis of intact and cleaved caspase-3 levels indicated that BTC has anti-apoptotic effects in the intestinal epithelium. We conclude that increased BTC levels support the survival of nascent adenomas in Apc(+/Min) mice, resulting in a larger total polyp number at later stages.
    FEBS Letters 08/2008; 582(19):2911-5. DOI:10.1016/j.febslet.2008.07.026 · 3.34 Impact Factor
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    • "The α v integrin subunit induced the phosphorylation of 845, 1068, 1086, and 1173 (Moro et al., 2002). In addition, ionizing radiation induced phosphorylation of 992 and 1173 while betacellulin induced 1068 (Sturla et al., 2005; Saito et al., 2004). While these are major sites of phosphorylation in response to EGF, it is interesting that differential phosphorylation occurs in response to specific biologic activation. "
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    ABSTRACT: The early events that occur rapidly after injury trigger signal cascades that are essential for proper wound closure of corneal epithelial cells. We hypothesize that injury releases ATP, which stimulates purinergic receptors and elicits the phosphorylation of epidermal growth factor receptor (EGFR) tyrosine residues and subsequent cell migration by a MMP and HB-EGF dependent pathway. We demonstrated that the inhibition of purinergic receptors with the antagonist, Reactive Blue 2, abrogated the phosphorylation of EGFR and ERK. Pre-incubation of cells with the EGFR kinase inhibitor, AG1478, and subsequent stimulation by injury or ATP resulted in a decrease in phosphorylation of EGFR and migration. Furthermore, downregulation of EGFR by siRNA, inhibited the EGF-induced intracellular Ca(2+) wave. However, the response to injury and ATP was retained indicating the presence of two signaling pathways. Inhibition with either CRM197 or TIMP-3 decreased injury and nucleotide-induced phosphorylation of both EGFR and ERK. Incubation in the presence of a functional blocking antibody to HB-EGF also resulted in a decrease in the phosphorylation of EGFR. In addition, cell migration was inhibited by CRM197 and rescued when cells were incubated with HB-EGF. We showed that injury-induced phosphorylation of specific tyrosine residues and found that a similar pattern of phosphorylation was induced by trinucleotides. These studies indicate that injury-induced purinergic receptor activation leads to phosphorylation of EGFR, ERK and migration.
    Experimental Eye Research 08/2007; 85(1):130-41. DOI:10.1016/j.exer.2007.03.009 · 3.02 Impact Factor
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    • "BTC acts by binding to members of the tyrosine kinase erbB receptor family. This triggers receptor phosphorylation and signaling to the cell nucleus through a cascade of mediators, resulting in modulation of gene transcription [30] [31]. The modulated genes may include key b-cell transcription factors such as Pdx1, NeuroD, Nkx2.2, and Nkx6.1, whose expression was observed to be induced by BTC in PHID cells. "
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    ABSTRACT: Beta-cell replacement represents the ultimate cure for type 1 diabetes, however it is limited by availability of organ donors. Adult human islets are difficult to propagate in culture, and efforts to expand them result in dedifferentiation. Here we describe conditions for expansion of adult human islet cells, as well as a way for their redifferentiation. Most cells in islets isolated from human pancreata were induced to replicate within the first week of culture in expansion medium. Cells were propagated for 16 population doublings, without a change in replication rate or noticeable cell mortality, representing an expansion of over 65,000-fold. Replication was accompanied by a decrease in expression of key beta-cell genes. Shift of the cells to differentiation medium containing betacellulin resulted in redifferentiation, as manifested by restoration of beta-cell gene expression and insulin content. These methods may allow transplantation of functional islet cells from single donors into multiple recipients.
    Biochemical and Biophysical Research Communications 04/2006; 341(2):291-8. DOI:10.1016/j.bbrc.2005.12.187 · 2.28 Impact Factor
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