Nuclear import mechanism of EGFR in breast cancer cells

Abstract

Receptor tyrosine kinases (RTKs), such as epidermal growth factor receptor (EGFR) are internalised from the plasma membrane by endocytosis and may be transported to the nucleus. EGFR, a receptor for EGF and other RTKs, HER-2 and HER-4 has an important role in signalling; it contains transactivational activity and can function as a transcription co-factor to activate gene promoters. High nuclear accumulation of imported full length EGFR is associated with an increased tumour proliferation and a reduced survival in cancer patients. However, little is known about the mechanism by which membrane-bound proteins, such as EGFR, translocate from the cell surface into the cell nucleus; how nuclear membrane proteins cross through the NPC to reach the INM. The mechanism of translocation for soluble proteins is also presently unclear. EGFR nuclear import is mediated by importin α/β. And it is exported from the nucleus by the exportin CRM1. Sec61β which may reside in the inner nuclear membrane (INM) is required for the release of EGFR from the INM into the nucleus. Nuclear transport involves binding of nuclear localisation sequences (NLSs) within the cargo to a transport receptor (karyopherins or importin). Karyopherins interact with certain nuclear pore complex (NPC) proteins (nucleoporins). Membrane proteins can access the INM through the NPC membrane: by diffusion, using classical nuclear transport factors (the importin/Ran system); or by an ATP dependent mechanism. EGFR may use the former mechanism. This work concentrates to show by electron microscopy and by Immuno-Fluorescence that upon EGF treatment, the biotinylated cell surface EGFR is trafficked to the INM through the NPC, yet remaining a membrane-bound protein. We also confirm that importin regulates EGFR nuclear transport to the INM and in addition, Sec61β is required for EGFR release to the nucleoplasm.
Altogether, this study of the mechanism of EGFR nuclear-cytoplasmic import in breast cancer cells, further confirms previous reports and provides an understanding of the nature and regulation of the nuclear EGFR pathway and the mechanism by which cell-surface EGFR is shuttled in the cell cytoplasm and channelled through the Golgi and Endoplasmic Reticulum (ER) compartments and into the nucleus through the NPC.