NCK adaptor proteins regulate clathrin-coated pit dynamics and EGF-stimulated PI3K-Akt signaling

The epidermal growth factor (EGF) receptor (EGFR) promotes cell growth, proliferation, and survival. EGF binding to EGFR activates receptor kinase activity, leading to membrane recruitment of class IA phosphoinositide-3-kinase (PI3K), thus activating Akt signaling. Clathrin-coated pits (CCPs) are plasma membrane endocytic structures that are enriched in signaling intermediates and regulate in EGF-stimulated Akt activation. The mechanisms and impact of recruitment of certain signaling molecules within the PI3K-Akt pathway to CCPs remains poorly understood. Using total internal reflection fluorescence microscopy (TIRFM), we observed that the adaptor proteins Nck1 and Nck2 are enriched within CCPs and regulate early CCP formation and maturation. Nck1 and Nck2 each support EGF-stimulated Akt phosphorylation. Notably, EGF stimulation triggers Nck-dependent enrichment of PI3K within CCPs, and perturbation of Nck adaptors suppressed cell proliferation and survival. This study identifies novel functions for Nck adaptor proteins in EGFR-mediated recruitment of PI3K-Akt signals within CCPs, Akt activation, and cell physiology.