Flotillin 2 is distinct from epidermal surface antigen (ESA) and is associated with filopodia formation

ECS-1, a monoclonal antibody (MoAb) raised to cultured human keratinocytes, stains the intercellular glycocalyx with a pemphigus-like pattern and reco gnizes a 35-kDa epidermal surface antigen (ESA) on Western blotting of kera tinocyte extracts. When ECS-I MoAb was used to screen a keratinocyte expres sion library, a unique cDNA was identified that predicted a 42-kDa globular protein of un known function. Th is putative ESA was conserved between mic e and humans and was encoded by a gene on chromosome 17q11-12 in linkage wi th neurofibromin. Homology between the cDNA sequence has been reported with flotillin 1, a caveolae associated protein, as well as Reggie 1 and 2, neu ronal proteins expressed during axonal regeneration present in activated CP I-anchored cell adhesion molecules in non-caveolar-associated micropatches. In order to determine whether the cDNA predicted protein and ECS-1 antigen were identical, we compared ECS-1 with the immunoreactivity of a new antib ody raised to the cDNA fusion protein in epidermis and cultured cells. The cDNA fusion protein was expressed in bacteria and in cos cells with his, FL AG, and EGFP reporter tags and by stable transfection as an EGFP fusion pro tein. The fusion protein and native protein of 42 kDa were detected by the new antibody, but not by the original ECS-1. Thus, the ECS-1 antigen, ESA ( 35 kDa), is clearly distinct from the protein predicted by the cDNA (rename d flotillin 2). Stable transfection of ESA/flotillin 2 fusion protein in co s cells induced filopodia formation and changed epithelial cells to a neuro nal appearance. Thus, the function of flotillin 2 may resemble that of the goldfish optic nerve neuronal regeneration proteins, Reggie 1 and 2. (C) 19 99 Wiley-Liss, Inc.