Subcellular distribution of p210BCR-ABL in CML cell lines and primary CD34+ CML cells.

We analysed the subcellular distribution of p210BCR-ABL protein using a junction-specific anti-BCR-ABL monoclonal antibody and confocal laser scanning microscopy (CLSM). Our studies have shown that p210BCR-ABL is arranged in discrete foci in the cytoplasm of cell lines and primary CD34+ cells but not mononuclear cells suggesting the foci may be a feature of immature chronic myeloid leukaemia cells. We have devised a strategy to score the foci and found the mean number of foci varies between the cell types. The number of foci per cell is directly related to the level of p210BCR-ABL expression. CLSM was also used to analyse the distribution and colocalization of CT10 regulator-like (CRKL) p210BCR-ABL. CRKL-p210BCR-ABL foci were completely or partially associated, touching or separate in different regions of the same cell. We also analysed the distribution of phosphorylated CRKL (pCRKL) with p210BCR-ABL and unexpectedly found only a small proportion of pCRKL in complex with p210BCR-ABL. The foci distribution and high levels of uncomplexed p210BCR-ABL, pCRKL and CRKL protein suggested the possibility of a dynamic equilibrium. Imatinib promoted nuclear transport of p210BCR-ABL-positive foci. It also disrupted complex formation between p210BCR-ABL and casitas B-cell lymphoma and CRKL but not between p210BCR-ABL and GRB2. Our observations of the CRKL and p210BCR-ABL complex may be important for understanding the function of CRKL.Leukemia (2008) 22, 559–571; doi:10.1038/sj.leu.2405057; published online 6 December 2007 [ABSTRACT FROM AUTHOR]