MBI PhD Qualifying Exam
Time: 2pm
Date: Wednesday, 14 March 2018
Venue: MBI, level 5 meeting rooms
Supervisors: Prof Virgile Viasnoff (Main supervisor), Prof Michael Sheetz (Co-supervisor)
Non-canonical role of RTKs (EGFR) in E-cadherin junction formation
Similar to cell-matrix adhesion formation and as a complex cellular mechanical function, E-cad mediated cell-cell adhesion is critical for cells to sense their neighboring cells. It has been noticed that classical E-cad homophilic ligation will result in EGF-independent activation of EGFR in epithelial cells. While the EGF-independent activity of EGFR was found to be necessary for forming contractile units to sense rigidity in focal adhesion, the role of ligand-free EGFR activity in regulating E-cad adhesions was remained to be clarified. Thus, it is of interest to investigate a non-canonical mechanism of EGFR, in which ligand-free EGFR activity is hypothesized to have a positive contribution to E-cad junction formation. In this project, we first started to investigate its role in regulating E-cad local contraction in a second-scale at a single cell level. In this regard, Cos-7 cell was used to spread on an E-cad pillar system, where we could analyze the local forces to indicate the E-cad contraction activity. Preliminary observation showed a decrease E-cad contraction pair density in EGFR inhibited cells on the rigid surface, supporting our hypothesis that ligand-free EGFR activity regulated E-cad local contraction. Future investigations are aimed to confirm the aforementioned result and evaluate whether the regulation of EGFR is mediated by Src and through myosin IIB contraction. Second, we used cell doublets system to illustrate the role of ligand-free EGFR activity in regulating dynamics of E-cad junction formation in a minute-scale at a cell doublet level. Cell contact size (ρ) and E-cad junction formation time (τ) were measured in S180 cells to demonstrate the dynamic process of E-cad junction formation. Changes of contact size were found to be positively correlated with changes in cortical tension. However, inhibiting ligand-free EGFR activity decreased contact size of doublets without affecting the cortical tension of single cells. Furthermore, junction formation time was found to be determined by myosin contractility and actin dynamics of the cortex, cooperatively. Consistent with these results, the decreases of actin dynamics in the free cortex of doublets were found to speed up further junction formation. In addition, we found that inhibiting ligand-free EGFR activity prevented the reorganization of actin dynamics in the free cortex of doublets. Taken together, we hypothesized that EGFR affected contact size through regulating myosin contractility in the contact plain and reorganized actin dynamics in the free cortex of doublets, which determined further junction formation time, and planned a few future experiments to confirm this hypothesis. We further plan to focus on the role of Rho family of GTPases (Rac1 and Cdc42) in mediating the EGFR regulation of dynamics of E-cad junction formation. Finally, we plan to perform collective cell migration with both MDCK and MCF-10A cells in geometrical confinements on a microcontact printed pattern. Based on this system, we will evaluate the role of ligand-free EGFR activity in regulating the speed of collective cell migration in an hour-scale at a tissue level.
**Please note the examination following the seminar is closed-door**
