Nuclear and Nucleolar Localization of 18-kDa Fibroblast Growth Factor-2 Is Controlled by C-terminal Signals

Zhi Sheng; John A. Lewis; William J. Chirico

doi:10.1074/jbc.M400123200

Nuclear and Nucleolar Localization of 18-kDa Fibroblast Growth Factor-2 Is Controlled by C-terminal Signals*

^‡Molecular and Cellular Biology Program, School of Graduate Studies, ^§Department of Anatomy and Cell Biology, State University of New York Downstate Medical Center, Brooklyn, New York 11203

↵¶ To whom correspondence should be addressed: Dept. of Anatomy and Cell Biology, Box 5, State University of New York Downstate Medical Center, 450 Clarkson Ave., Brooklyn, NY 11203. Tel.: 718-270-1308; Fax: 718-270-3732; E-mail: william.chirico{at}downstate.edu.

Abstract

Members of high (22-, 22.5-, 24-, and 34-kDa) and low (18-kDa) molecular mass forms of fibroblast growth factor-2 (FGF-2) regulate cell proliferation, differentiation, and migration. FGF-2s have been previously shown to accumulate in the nucleus and nucleolus. Although high molecular weight forms of FGF-2 contain at least one nuclear localization signal (NLS) in their N-terminal extension, the 18-kDa FGF-2 does not contain a standard NLS. To determine signals controlling the nuclear and subnuclear localization of the 18-kDa FGF-2, its full-length cDNA was fused to that of green fluorescent protein (GFP). The fusion protein was primarily localized to the nucleus of COS-7 and HeLa cells and accumulated in the nucleolus. The subcellular distribution was confirmed using wild type FGF-2 and FGF-2 tagged with a FLAG epitope. A 17-amino acid sequence containing two groups of basic amino acid residues separated by eight amino acid residues directed GFP and a GFP dimer into the nucleus. We systematically mutated the basic amino acid residues in this nonclassical NLS and determined the effect on nuclear and nucleolar accumulation of 18-kDa FGF-2. Lys¹¹⁹ and Arg¹²⁹ are the key amino acid residues in both nuclear and nucleolar localization, whereas Lys¹²⁸ regulates only nucleolar localization of 18-kDa FGF-2. Together, these results demonstrate that the 18-kDa FGF-2 harbors a C-terminal nonclassical bipartite NLS, a portion of which also regulates its nucleolar localization.