Overexpressed cyclin D3 contributes to retaining the growth inhibitor p27 in the cytoplasm of thyroid tumor cells

The majority of thyroid carcinomas maintain the expression of the cell grow th suppressor p27, an inhibitor of cyclin-dependent kinase-2 (Cdk2). Howeve r, we fmd that 80% of p27-expressing tumors show an uncommon cytoplasmic lo calization of p27 protein, associated with high Cdk2 activity. To reproduce such a situation, a mutant p27 devoid of its COOH-terminal nuclear-localiz ation signal was generated (p27-NLS). p27-NLS accumulates in the cytoplasm and fails to induce growth arrest in 2 different cell lines, indicating tha t cytoplasm-residing p27 is inactive as a growth inhibitor, presumably beca use it does not interact with nuclear Cdk2. Overexpression of cyclin D3 may account in part for p27 cytoplasmic localization In thyroid tumors and cel l Lines, cyclin D3 expression was associated with cytoplasmic localization of p27. Moreover, expression of cyclin D3 in thyroid carcinoma cells induce d cytoplasmic retention of cotransfected p27 and rescued p27-imposed growth arrest. Endogenous p27 also localized prevalently to the cytoplasm in norm al thyrocytes engineered to stably overexpress cyclin D3 (PC-D3 cells). In these cells, cyclin D3 induced the formation of cytoplasmic p27-cyclin D3-C dk complexes, which titrated p27 away from intranuclear complexes that cont ain cyclins A-E and Cdk2. Our results demonstrate a novel mechanism that ma y contribute to overcoming the p27 inhibitory threshold in transformed thyr oid cells.