Membrane-anchored forms of EGF stimulate focus formation and intercellular communication

Epidermal growth factor (EGF) is the prototype for a small family of soluble proteins that bind to and activate the EGF receptor. These proteins are derived from larger propeptides that are anchored to the plasma membrane. Although the signalling properties of soluble EGF are well-characterized, the signalling potential of the membrane-anchored form had not been determined. We therefore investigated whether membrane-anchored EGF can stimulate the EGF receptor. An EGF mini-gene expression system that we had previously constructed for expression of soluble EGF was modified to encode anchored forms of EGF. In the encoded proteins EGF was fused to the spacer in the EGF propeptide that separates EGF from the transmembrane domain. The spacer was followed by vesicular stomatitis virus G protein transmembrane and cytoplasmic domain sequences. Three forms of EGF/G fusion protein were expressed in rat fibroblasts. The plasmids for expression of anchored EGF induced focus formation in rat fibroblasts, indicating that anchored EGF can cause autocrine transformation. When mixed with indicator HeLa cells, cell lines expressing EGF/G fusion proteins activated the HeLa EGF receptor. This activation was mediated by cell-associated, rather than soluble EGF. The finding that membrane-anchored EGF is capable of activating the EGF receptor on neighboring cells has broad implications for the functions of EGF in the organism.