Dual Retinoblastoma-binding Proteins with Properties Related to a Negative Regulator of Ras in Yeast

Yue-Wei Qian,Eva Y.-H.P Lee

doi:10.1074/jbc.270.43.25507

Abstract

The retinoblastoma protein (Rb) interacts with multiple cellular proteins that mediate its cellular function. We have identified nine polypeptides that bind to the T-binding domains of Rb using an Rb affinity resin. RbAp48 and RbAp46 are quantitatively the major Rb-associated proteins purified by this approach. RbAp48 was characterized previously and was found to be related to MSI1, a negative regulator of Ras in the yeast Saccharomyces cerevisiae. Here we report the cloning and characterization of RbAp46. RbAp46 shares 89.4% amino acid identity with RbAp48. The internal WD repeats, which are found in a growing number of eukaryotic proteins, are conserved between RbAp46 and RbAp48. Like RbAp48, RbAp46 forms a complex with Rb both in vitro and in vivo and suppresses the heat-shock sensitivity of the yeast RAS2Val-19 strains. We have also isolated the murine cDNA homologs of RbAp48 and RbAp46. Although both mRNA can be detected in all mouse tissues, their mRNA levels vary dramatically between different tissues. No significant differences were observed in the expression patterns of these genes in most tissues except thymus, testis, and ovary/uterus, in which 2-fold differences were observed. Interestingly, the mouse and human RbAp48 amino acid sequences are completely identical, and the mouse and human RbAp46 differ only by one conserved amino acid substitution. These results suggest that RbAp48 and RbAp46 may have shared as well as unique functions in the regulation of cell proliferation and differentiation.

Highlights

Studies of familial cancer syndromes, such as retinoblastoma, have lead to the identification of tumor suppressor genes
The nucleotide sequence(s) reported in this paper has been submitted to the GenBankTM/EMBL Data Bank with accession number(s) U35141, U35142, and U35143 (RbAp46)
retinoblastoma protein (Rb) protein plays a major role in the inhibition of G1 to S phase transition [12, 13], but is important for cell differentiation

Summary

Introduction

Studies of familial cancer syndromes, such as retinoblastoma, have lead to the identification of tumor suppressor genes. Lens epithelial cells fail to terminally differentiate and undergo programmed cell death [14, 15]. These pleiotropic cellular functions of Rb are likely to be mediated by association with multiple cellular proteins [16]. One of the best studied Rb-associated protein is the transcriptional factor E2F, which regulates expression of several genes essential for S phase entry [17]. UBF, a ribosomal transcription factor, is another well studied Rb-associated protein (19 –21). The UBF-Rb interaction results in suppression of the synthesis of ribosomal RNA by RNA polymerase I, thereby globally inhibiting cell proliferation

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Conclusion