Retinoblastoma protein interacts with and modulates the transcriptional activity of the tal-l/E2A/Lmo2 complex in differentiating erythroid precursors

Recent observations suggest that pRb plays a lineage- and stage-specific functional role in normal human adult hematopolesis. Thus, pRb expression in the erythrold lineage peaks at the CFU-E/proerythroblast stage, when proliferation Is exponential and the erythroid differentiation program becomes fully expressed; functional studies by antlsense ollgomers Indicate that suppression of Rb mRNA causes selective Inhibition of erythroid but not granulopoletic colony formation (Condorelli et al., Proc. Natl. Acad. Sei. USA, 1995). We have recently described the presence of a complex containing tal-l/E2A proteins in differentiated erythroid cells (Condorelli et al., Blood, 1995). In this report gel shift and immunopreclpltation/Western blot experiments Indicate that erythroblasts contain a multlprotein transcriptional complex comprising tal-l/E2A/Lmo2 and pRb. Furthermore, pRb Increases the E box binding activity of the tall/E2A/Lmo2 protein complex. In fact, addition of synthetic fulllength pRb causes a dose-dependent increase up to 10 fold in tal-l/E2A/Lmo2 complex binding to its recognition sequence. Finally, to explore the functional significance of the Interaction between pRb and the tal-l/E2A/Lmo2 complex, we have evaluated the effect of pRb expression on an artificial reporter plasmid (ElbLuc-E6) containing six binding sites for tai-l/E2A heterodimers m transiently transfected pRb negative SAOS-2 cell line. Results showed that pRb markedly enhances the transcriptional activity of the tal-1 /E2A/Lmo2 complex. Altogether, these findings suggest that the positive functional role of pRb In erythroid differentiation is mediated via modulation of the activity of the erythroid-specific tal-1/E2A/Lmo2 transcriptional complex.