Loss of the xeroderma pigmentosum group B protein binding site impairs p210 BCR/ABL1 leukemogenic activity.

Previous studies have demonstrated that p210 BCR/ABL1 interacts directly with the xeroderma pigmentosum group B (XPB) protein, and that XPB is phosphorylated on tyrosine in cells that express p210 BCR/ABL1. In the current study, we have constructed a p210 BCR/ABL1 mutant that can no longer bind to XPB. The mutant has normal kinase activity and interacts with GRB2, but can no longer phosphorylate XPB. Loss of XPB binding is associated with reduced expression of c-MYC and reduced transforming potential in ex-vivo clonogenicity assays, but does not affect nucleotide excision repair in lymphoid or myeloid cells. When examined in a bone marrow transplantation (BMT) model for chronic myelogenous leukemia, mice that express the mutant exhibit attenuated myeloproliferation and lymphoproliferation when compared with mice that express unmodified p210 BCR/ABL1. Thus, the mutant-transplanted mice show predominantly neutrophilic expansion and altered progenitor expansion, and have significantly extended lifespans. This was confirmed in a BMT model for B-cell acute lymphoblastic leukemia, wherein the majority of the mutant-transplanted mice remain disease free. These results suggest that the interaction between p210 BCR/ABL1 and XPB can contribute to disease progression by influencing the lineage commitment of lymphoid and myeloid progenitors.

The RhoGEF domain of p210 Bcr-Abl activates RhoA and is required for transformation.

The BCR-ABL oncogene encodes an in-frame fusion protein containing N-terminal sequences derived from Bcr and C-terminal sequences derived from Abl. Bcr contains a centrally located Rho-specific guanine nucleotide exchange factor (RhoGEF) domain that is retained within p210 Bcr-Abl. Although this domain is subject to autoinhibition in the context of Bcr, here we show that it is constitutively activated in p210 Bcr-Abl. p210 Bcr-Abl can stimulate RhoA activation independently of its tyrosine kinase activity, and mutations within the RhoGEF domain that are predicted to eliminate RhoGEF activity inhibit RhoA activation. The RhoGEF mutant of p210 Bcr-Abl does not affect the tyrosine kinase activity of the molecule, nor the ability of p210 Bcr-Abl to interact with XPB through the RhoGEF domain. Despite retaining normal levels of tyrosine kinase activity, the RhoGEF mutant of p210 Bcr-Abl is impaired in transforming activity as measured by anchorage-independent growth. However, the mutant is still able to confer the phenotype of growth factor independence in myeloid cells, suggesting that some, but not all parameters of p210 Bcr-Abl transformation, are dependent upon a catalytically active RhoGEF domain. Collectively, these observations identify a gain-of-function activity attributable to the RhoGEF domain of p210 Bcr-Abl that is required to support the transformed phenotype.