Genetic alterations disrupting the nuclear localization of the retinoblastoma-related gene RB2/p130 in human tumor cell lines and primary tumors

The prototypic tumor suppressor gene, the retinoblastoma gene (RB/p105), is mutated in a variety of human tumors. However, to date, mutational data on retinoblastoma family members p107 and RB2/p130 in tumors is lacking. We s tudied the expression of pRb2/p130 by immunocytochemistry and Western blot analysis in a panel of human osteosarcoma and lymphoid cell lines. Only the Lymphoid cell lines showed an abnormal cytoplasmic localization of pRb2/p1 30, suggesting possible alterations within the region of nuclear localizati on signaling. We screened these cell lines for genetic alterations of the R B2/p130 gene in the region of the putative bipartite nuclear localization s ignal (NLS), This region is highly homologous with that of the RB/p105 gene . In addition, we screened four primary Burkitt's lymphomas for genetic alt erations in the RB2/p130 gene. Naturally occurring mutations, which disrupt the putative bipartite NLS, were found in lymphoma cell lines and primary tumors, but not in the osteosarcoma cell lines, where normal nuclear locali zation of the protein was detectable. Site-directed mutagenesis and transfe ction assay using NLS mutants displayed markedly reduced biological activit y as measured by flow cytometric analysis. This study clearly describes RB2 /p130 as an important target for mutations and subsequent inactivation in l ymphoma pathogenesis, thus validating that RB2/p130 is a classical tumor su ppressor gene.