CRKL (CRK-like) is an adapter protein predominantly phosphorylated in cells that express the tyrosine kinase p210(BCR-ABL), the fusion product of a (9;22) chromosomal translocation causative for chronic myeloid leukemia. It has been unclear, however, whether CRKL plays a functional role in p210(BCR-ABL) transformation. Here, we show that CRKL is required for p210(BCR-ABL) to support interleukin-3-independent growth of myeloid progenitor cells and long-term outgrowth of B-lymphoid cells from fetal liver-derived hematopoietic progenitor cells. Furthermore, a synthetic phosphotyrosyl peptide that binds to the CRKL SH2 domain with high affinity blocks association of endogenous CRKL with the p210(BCR-ABL) complex and reduces c-MYC levels in K562 human leukemic cells as well as in mouse hematopoietic cells transformed by p210(BCR-ABL) or the imatinib-resistant mutant T315I. These results indicate that the function of CRKL as an adapter protein is essential for p210(BCR-ABL)-induced transformation.