The t(9;22)(q34;q11) is detected in 95% of CML and 20–30% of adult ALL. CML is a myeloproliferative syndrome [1
]. In contrast, Ph+-ALL is an acute disease characterized by blasts blocked at the pre-lymphatic stage of differentiation. Patients suffering from Ph+-ALL constitute a high risk group of ALL (5–10% survival rate after five years)[2
]. The factors determining the biological differences between CML and Ph+-ALL are completely unknown.
The t(9;22) is a reciprocal translocation. A portion of chromosome 9 translocates onto chromosome 22 (22+), thereby replacing a fragment which in turn translocates onto chromosome 9 (9+) [1
]. The derivative of chromosome 22 (22q+) can be revealed by cytogenetic techniques as the so-called Philadelphia chromosome (Ph).
On chromosome 22, translocation (9;22) involves the bcr
(breakpoint cluster region) locus and there are two principal regions in which the breaks occur: (major) M-bcr, which spans between exons 12 to 16, and (minor) m-bcr, in the first intron, about 50 kb 5' of M-bcr. The product of fusion between M-bcr and abl is a protein of 210 kDa, the p210((BCR-ABL))
, which is highly specific for CML. Due to the fact that the m-bcr maps within an intron, the p185((BCR-ABL))
transcript in Ph+-ALL is constant [1
]. Through fusion to BCR, the kinase activity of ABL becomes constitutively activated, leading to the constitutive activation of the "down-stream" signal transduction pathways, such as RAS, JAK-STAT and PI-3 kinase, responsible for the oncogenic potential of BCR/ABL [1
]. The suppression of constitutively active ABL kinase by specific kinase inhibitors, such as Imatinib [3
], Nilotinib [4
] and Dasatinib. [5
], reverts the oncogenic potential of BCR/ABL and these drugs are currently in clinical evaluation.
The breakpoint on chromosome 9 is located in intron 1 of the abl
gene locus. It is, in contrast to the breakpoints on chromosome 22, constant and located between exons 1 and 2. The abl/bcr
fusion genes on 9+ differ depending on the breakpoint on chromosome 22. Fusion between M-bcr and abl
results in the 'small' abl/bcr
fusion gene encoding a 'small' ABL/BCR transcript, detectable in 65% patients suffering from CML [6
], which is translated into an ABL/BCR protein with a theoretical molecular mass of about 40 kDa – p40(ABL/BCR)
(Zheng et al. in preparation). The fusion between m-bcr and abl
leads to a 'large' transcript, present in 100% of examined patients with a Ph+-ALL [7
], which encodes a fusion protein with a theoretical molecular mass of about 96 kDa – p96(ABL/BCR)
(Zheng et al. in preparation).
The ABL/BCR fusion proteins represent mutants of the protein kinase BCR. BCR is a Rho-GEF due to the presence of a dbl homology (DH) domain and a pleckstrin homology (PH) domain (Fig. ) [8
]. The GEFs activate members of the Ras superfamily by increasing the proportion of their GTP-bound form with respect to the GDP-bound form [10
]. The prototype for Rho-GEF, the diffuse B-cell lymphoma (Dbl) oncogene, has a strong transformation activity in NIH3T3 fibroblasts, and both the Rho-GEF function and the oncogenic potential of Dbl depend on its DH-domains [11
Figure 1 Modular organization of the main t(9;22) translocation products. The protein sizes are not to scale. CC – coiled coil oligomerization interface; S/T kinase – serine/threonine kinase domain; DH – dbl homology domain; PH – (more ...)
Furthermore, BCR also contains a C-terminal Rac-GAP domain. In contrast to GEFs, the GTPase activating proteins (GAPs) promote hydrolysis of GTP to GDP and increase the inactive forms of small GTPases of the Ras superfamily [10
]. In fact, BCR is a negative regulator of Rac, as demonstrated by the fact that it reduces the Rac1-dependent activation of the protein kinase Pak1, an activator of the JNK pathway, via its GAP function [12
The Rho-GEF and Rac-GAP functions strongly suggest an important role of BCR in cytoskeleton modeling by regulating the activity of Rho-like GTP-ases, such as Rac, cdc42, and Rho [11
]. Rho, cdc42 and Rac are involved in the formation and maintenance of 'stress fibers', filopodia and lamellipodia, respectively [13
]. The N-terminal 'coiled coil' dimerization interface of BCR, its serine/threonine kinase activity, and the tyrosine phosphorylation site at position 177 (Y177) are indispensable for its function [1
are mutated BCR, but nothing is known about their biology. In the ABL/BCR fusion proteins, the N-terminus of BCR is substituted by the first exon of abl
. The differences between ALL- and CML-specific ABL/BCR lie in the presence of the DH and PH domains. CML-associated p40(ABL/BCR)
lacks the DH/PH domains conserved in the ALL-specific p96(ABL/BCR)
(Fig. ). Thus the ALL-specific p96(ABL/BCR)
fusion protein is an N-terminally truncated Rho-GEF and, therefore, a putative oncogene [15
To determine the role of the ABL/BCR in Ph+ leukemia we compared the effects of BCR and the ABL/BCR proteins on the regulation of Rho-like GTPases, cytoskeleton modeling and cell motility.