Molecular analysis of 12 patients with the t(8;2l) translocation and M2 acute myelogenous leukemia

Abstract

The t(8;21)(q22;q22) is a nonrandom cytogenetic abnormality associated with acute myelogenous leukemia of the M2 subtype (FAB classification). The 8q- and 21q+ derivative chromosomes have previously been isolated in somatic cell hybrids and used to map the anonymous sequences D21S65 and D21S17, which were proximal and distal, respectively, to the breakpoint on chromosome 21. DNA from a series of 12 t(8;21) patients and 7 controls was analyzed by pulsed field gel electrophoresis. Physical linkage of probes D21S65 and D21S17 on a 2100 kb NruI fragment was established by partial digestion experiments. In all the patients, the translocation generated a rearranged D21S65 NruI fragment of 650 to 750 kb, suggesting heterogeneity in the breakpoints. This heterogeneity was confirmed by using BssHII, SacII, and EagI enzymes. Our results are consistent with the presence of a 100 Kb breakpoint cluster region on chromosome 21 encompassing the AML1 gene. Interestingly, in half of the patients, demethylation of an NruI site located 7 kb proximal to the last exon of the AML1 gene occurred on the nontranslocated chromosome 21.

Identification of AML-1 and the (8;21) translocation protein (AML-1/ETO) as sequence-specific DNA-binding proteins: the runt homology domain is required for DNA binding and protein-protein interactions.

“…Addition ofAMLl-programmed lysates to extracts from the leukemic bone marrow aspirate enhanced the formation of the endogenous A complex, indicating that the reconstituted complex comigrates with the A complex (Fig. 7C) (16,45,48), consistently conserves AML-1 sequences to the end of a domain homologous to the Drosophila runt protein (the runt homology domain, rhd) (5,28,40), suggesting that this region of the protein is critical for the transformation process. Random oligonucleotide selection was used to identify AML-1 as a DNAbinding protein with the consensus recognition sequence TGT/cGGT.…”