Intrinsic regulation of the interactions between the SH3 domain of p85 subunit of phosphatidylinositol-3 kinase and the protein network of BCR/ABL oncogenic tyrosine kinase

Objective. BCR/ABL fusion tyrosine kinase is responsible for the initiation and maintenance of the Philadelphia chromosome-positive chronic myelogenous leukemia (CML) and a cohort of acute lymphocytic leukemias. We show that a signaling protein, phosphatidylinositol-3 kinase (PI-3k), is essential for growth of CML cells, but not of normal hematopoietic cells, and that p85α subunit of PI-3k co-immunoprecipitates with BCR/ABL. Therefore, we made an attempt to better characterize p85α-BCR/ABL interactions. Materials and Methods. The mutants of p85α-SH3 domain were generated by in vitro site-directed mutagenesis system. Protein lysates were obtained from p210BCR/ABL-transformed murine 32Dcl3 myeloid cells, and in vitro transcription/translation was used to produce BCR/ABL protein. Pull-down and Western analyses were performed to detect the interaction between BCR/ABL and p85α-SH3. BCR/ABL-transformed 32Dcl3 cells were infected with internal ribosome entry site-green fluorescent protein retroviral construct encoding p85α-SH3 mutants to assess their biological effects. Results. We show here that the SH3 domain of p85α (p85α-SH3) pulls down the p210BCR/ABL kinase from hematopoietic cell lysates. The interaction between p85α-SH3 and BCR/ABL may be intermediated by proteins such as c-Cbl, Shc, Grb2, and/or Gab2. Mutations in the p85α-SH3 region responsible for proline-rich motif binding either abrogate or enhance these interactions. These mutants exert a modest inhibitory effect on growth factor-independent proliferation of BCR/ABL-positive 32Dcl3 cells. Conclusions. Based on this information we speculate on the capability of p85α-SH3 to interact with the protein network of BCR/ABL oncoprotein.