AMINO-ACID-SEQUENCE RERMS REPRESENTS THE ACTIVE DOMAIN OF AMYLOID-BETA A4-PROTEIN PRECURSOR THAT PROMOTES FIBROBLAST GROWTH/

The growth of A-1 fibroblasts depends on exogenous amyloid beta/A4 pro tein precursor (APP), providing a simple bioassay to study the functio n of APP. Our preliminary study, testing the activity of a series of f ragments derived from the secreted form of APP-695 (sAPP-695) on this bioassay, has shown that at least one of the active sites of sAPP-695 was localized within a 40-mer sequence (APP296-335, Kang sequence; Roc h, J.-M., I. P. Shapiro, M. P. Sundsmo, D. A. C. Otero, L. M. Refolo, N. K. Robakis, and T. Saitoh. 1992. J. Biol. Chem. 267:2214-2221). In the present study, to further characterize the growth-promoting activi ty of sAPP-695 on fibroblasts, we applied a battery of synthetic pepti des on this bioassay and found that: (a) the sequence of five amino ac ids, RERMS (APP328-332), was uniquely required for the growth-promotin g activity of sAPP-695; (b) the activity was sequence-specific because the reverse-sequence peptide of the active domain had no activity; an d (c) the four-amino-acid peptide RMSQ (APP330-333), which partially o verlaps the COOH-terminal side of the active sequence RERMS, could ant agonize the activity of sAPP-695. Furthermore, a recombinant protein w hich lacks this active domain (APP20-591 without 306-335) did not prom ote fibroblast cell growth, suggesting that this domain is the only si te of sAPP-695 involved in the growth stimulation. The availability of these biologically active, short peptides and their antagonists shoul d prove to be an essential step for the elucidation of APP involvement in regulation of cellular homeostasis.