INTRANUCLEAR LOCALIZATION OF INSULIN-LIKE-GROWTH-FACTOR BINDING PROTEIN-3 (IGFBP-3) DURING CELL-DIVISION IN HUMAN KERATINOCYTES

Insulin-like growth factor-I (IGF-I) stimulation of basal keratinocyte s is an essential component of normal epidermal homeostasis, In additi on to the IGF receptor, basal keratinocytes synthesize insulin-like gr owth factor binding protein-3 (IGFBP-3), The HaCaT keratinocyte cell l ine, which has many characteristics of basal keratinocytes, synthesize s IGFBP-3 that in vitro reduces its IGF-I responsiveness. IGFBP-3 has attracted interest as a potential growth arrest protein, both via its ability to modulate IGF-I responsiveness, and more controversially via IGF-I-independent mechanisms. Intracellular modes of action have been proposed, and a nuclear localization consensus sequence has previousl y been identified within IGFBP-3, Using immunocytochemistry with a bio tinylated antibody specific for IGFBP-3, we investigated the intracell ular localization of IGFBP-3 in subconfluent monolayer cultures of HaC aT cells. Diffuse cellular staining was visible, potentially correspon ding to cell surface and nascent cytoplasmic IGFBP-3, Of particular in terest however, was the localization of staining over the nuclei of a large proportion of cells that were undergoing cell division. Antibody staining was specific for IGFBP-3 because addition of recombinant hum an IGFBP-3 to the antibody prior to incubation with the cells inhibite d these staining patterns. Optical sections obtained using a confocal laser scanning microscope showed that in keratinocytes undergoing cell division, IGFBP-3 was localized inside the nucleus. These results sho w that intracellular IGFBP-3 localization is altered during the cell c ycle and suggest a possible nuclear role for IGFBP-3 during cell divis ion.