Regulation of retinoblastoma protein activity by the c terminal domain

Purpose. The retinoblastoma protein (Rb) is a iumor suppressor that regulates cell proliferation by controlling cellular transition from Gl to S phase of the cell cycle. The A-B pocket of Rb forms a transcriptional represser motif which is regulated through phosphorylation of the C-terminal of Rb. We examined the mechanism of C-terminal regulation of the A-B pocket. Methods. Transient iransfections of plasmids expressing GAL4-R5 or LexA-Rb fusion proteins in C33A (Rb-t cells, and detection of transcriptional activity using a reporter plasmid containing the chloramphenico! acetyttransferase (CAT) gene, SV40 promoter, and GAL4 or LexA binding sites. Results. Overexpression of the A-B pocket as a GAL4 or LexA fusion protein resulted in repression of CAT activity. Coexpression of the C-terminal domain on a separate protein blocked this transcriptional repression. Conclusions. Since the C-terminal can block transcriptional repression by the A-B pocket when expressed on a separate protein, the mechanism of C-terminal regulation may depend on interdomain interactions, rather than intramolecular conformational changes.