Abstract 4003: The role of ubiquitin-mediated binding in p210 BCR/ABL induced leukemogenesis

Abstract

Abstract Chronic myelogenous leukemia (CML) is a malignant hematopoietic stem cell disorder that is invariably associated with a balanced reciprocal translocation between chromosomes 22 and 9. The major product of the rearrangement is a 210 kD in-frame fusion protein (p210 BCR/ABL) that contains amino-terminal sequences from BCR, and carboxy-terminal sequences from the non-receptor tyrosine kinase ABL. This fusion event leads to the constitutive activation of the ABL-encoded kinase activity, which is the principal driving force behind CML. Thus, inhibitors that target the kinase activity produce durable clinical remissions. Although the ABL-encoded kinase activity is essential for disease progression, several studies have shown that BCR encoded sequences are also necessary for BCR/ABL-mediated leukemogenesis. In this current study we have identified ubiquitin as a binding partner for BCR, and demonstrated that binding is retained in p210 BCR/ABL. We have localized the docking site for ubiquitin to residues 180-191 which is present in all fusion variants of BCR/ABL (p190, p210 and p230). This putative ubiquitin binding domain (UBD) does not conform to any known consensus sequence, and is unique to BCR. Deletion of the UBD does not impair the auto-or transkinase activity of p210 BCR/ABL, nor does it impair the interaction between p210 BCR/ABL and GRB2, or the ability of p210 BCR/ABL to activate ERK1/2. A mutation at residue tyr-177 of p210 BCR/ABL also does not impair the interaction with ubiquitin suggesting that the GRB2 and ubiquitin binding sites are adjacent, but separable. β-catenin has been identified as a binding partner for BCR and BCR/ABL, and the docking site has been mapped to a region of BCR/ABL that contains the UBD. Over-expression of p210 BCR/ABL, but not the ubiquitin binding mutant, leads to an accumulation of β-catenin that is phosphorylated on the serine residues that normally trigger ubiquitin-mediated turnover. This difference cannot be attributed to a difference in the activation status of GSK-3β or Pin-1. These results suggest that either β-catenin and ubiquitin share a docking site on p210 BCR/ABL, or that the interaction between β-catenin and p210 BCR/ABL is ubiquitination dependent. The latter possibility is consistent with a previous study which showed that glycine residue in β-catenin is required for Bcr interaction. We propose a model in which p210 BCR/ABL may influence the Wnt signaling pathway by binding ubiquitinated β-catenin, and stabilizing it against degradation. In a CML animal model, mice received the ubiquitin binding mutant have significantly increased the survival time compared with p210 BCR-ABL positive transplant mice. The role of ubiquitin mediated binding in p210 BCR/ABL-mediated leukemogenesis is currently being explored. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4003.