Figure 1. ErbB3 mutations block tyrosine phosphorylation, PI3K binding and show a defect in in vitro chemotaxis and invasion to HRGβ1.
A. Flow cytometry analysis of surface ErbB3 levels in pLXSN (solid line), ErbB3WT (dotted line), and ErbB3-Mutant (dashed line) cells. A mouse anti-ErbB3 primary followed by an APC-labeled secondary was used. The curve with solid shading represents ErbB3WT cells incubated with just the APC-conjugated secondary antibody.
B. HRGβ1-induced ErbB3 tyrosine phosphorylation and association to the p85 subunit of PI3K. Serum starved MTLn3 ErbB3WT or ErbB3-Mutant cells were stimulated with buffer or HRGβ1 (0 or 0.4 HRG) for 5 minutes. ErbB3 immunoprecipitates (IP) and whole cell lysates (WCL) were resolved by SDS/PAGE, and probed for (PTyr) or p85α. Membranes were stripped and reprobed to control for ErbB3 levels.
C Chemotaxis to HRGβ1 of pLXSN (light gray bars), ErbB3WT (dark gray bars) and ErbB3-Mutant (patterned bars). Data are mean and SEM of 6-27 wells in 3-6 independent experiments.
D. In vitro invasion responses of ErbB3WT (black bars) and ErbB3-Mutant cells (patterned bar) into Matrigel-coated transwells stimulated by buffer or 12.5 nM HRGβ1. Data are presented as % Area invaded, and are mean and SEM of 3 independent experiments. *:p< 0.02