Spatial-temporal regulation of EGFR phosporylation by protein tyrosine phosphatases
| dc.contributor.advisor | Bastiaens, Philippe | |
| dc.contributor.author | Fengler, Sven | |
| dc.contributor.referee | Grecco, Hernán E. | |
| dc.date.accepted | 2014-08-29 | |
| dc.date.accessioned | 2017-06-23T13:30:21Z | |
| dc.date.available | 2017-06-23T13:30:21Z | |
| dc.date.issued | 2017 | |
| dc.description.abstract | The activity of the epidermal growth factor receptor (EGFR) and its interactions with protein tyrosine phosphatases (PTPs) is what determines growth factor signaling. Due to its intrinsic autocatalytic properties, EGFR can undergo autonomous ligand independent activation. EGFR in this case, cycles between the plasma membrane (PM) and recycling endosomes where PTPs dephosphorylate the receptor to avoid spurious receptor signals. EGF-induced receptor dimerization results in a robust trans-phosphorylation of tyrosine residues that allows stable binding of signaling effectors. The majority of EGFR at the PM is internalized and undergoes degradation in lysosomal compartment. After internalization, the EGFR encounters PTPs at different cellular locations, which thereby regulate the signal duration of the receptor. Moreover, specific dephosphorylation of phosphotyrosines that are required for ubiquitin ligase (Cbl) binding reduces the receptor ubiquitylation and thereby its degradation rate. Due to their dephosphorylation activity, PTPs were thought as negative regulators of RTK signalling, but it has been shown that PTPs also promote receptor phosphorylation by activating cytosolic kinases that in turn phosphorylate RTKs (Julien et al., 2011). To understand the spatial-temporal regulation of EGFR phosphorylation by PTPs, we used CA-FLIM (Grecco et al., 2010). This method allowed us to quantify the phosphorylated fraction of EGFR in cells, upon perturbation of PTP expression by siRNAs or cDNAs. Upon opposing perturbations, we identified several PTPs that have indicated either a negative or positive effect on EGFR phosphorylation. Classification of the temporal phosphorylation profiles of EGFR discovered 5 functional groups of PTPs acting at early and/or late time points after EGF stimulation. PTPs within each group showed differences in their regulatory influence highlighting individual impact in EGF signaling. Predominantly cytosolic PTPs regulated early EGFR phosphorylation, whereas receptor-like PTPs (RPTPs) induced a transient response profile. Analysis of the spatial-temporal phosphorylation profile of EGFR upon PTPRA, PTPN1 or PTPN2 expression showed an almost abolished axial phosphorylation of EGFR that might promote receptor recycling. In contrast, we identified a positive regulatory function of MTM1, DUSP7 and PTPN21 that was further validated using multi-parametric single cell information. These PTPs induced an early amplification of receptor phosphorylation near the PM. Our results strongly suggest that MTM1 and PTPN21 inhibit the degradation pathway and thereby enhancing the phosphorylated fraction of internalized EGFR. In summary, the presented work provides novel insights about when and where PTPs regulate EGFR phosphorylation and how this could affect cellular responses. | en |
| dc.identifier.uri | http://hdl.handle.net/2003/36007 | |
| dc.identifier.uri | http://dx.doi.org/10.17877/DE290R-18025 | |
| dc.language.iso | en | de |
| dc.subject | Cell-array | en |
| dc.subject | EGFR | en |
| dc.subject | PTP | en |
| dc.subject | CA-FLIM | en |
| dc.subject.ddc | 570 | |
| dc.title | Spatial-temporal regulation of EGFR phosporylation by protein tyrosine phosphatases | en |
| dc.type | Text | de |
| dc.type.publicationtype | doctoralThesis | en |
| dcterms.accessRights | open access |