Molecular analysis of the chromosomal breakpoint and fusion transcripts in the acute lymphoblastic SEM cell line with chromosomal translocation t(4;ll)

Abstract

The chromosomal breakpoint and fusion transcripts of the pre-B-leukaemia-derived SEM cell line carrying a reciprocal t(4;11)(q21;q23) translocation were analysed. The breakpoint from derivative chromosome der4 was cloned and sequenced. The crossover site was localized in intron 7 of the ALL-1 gene on chromosome 11q23 and in a large intron of the AF-4 (FEL) gene. RNA transcripts from both wild-type genes and both hybrid genes were detected by reverse transcriptase polymerase chain reaction (RT-PCR) assays. In addition, alternatively spliced mRNA species derived from the der4 chromosome were found. They were generated by using the exon 5' of the breakpoint on der4 as a common splice donor site and the 5' boundaries of exons 8 or 9 of the ALL-1 gene as alternative splice acceptor sites. The hypothesis is proposed that selective pressure operators to maintain the presence of both derivative chromosomes as important elements in the leukaemogenic process.

A DNA damage repair mechanism is involved in the origin of chromosomal translocations t(4;11) in primary leukemic cells

“…A third explanantion proposes that only translocations occuring in these breakpoint cluster regions produce chimeric proteins that confer a selectable advantage. Translocations may also accur elsewhere with equal frequencies, but these would not lead to the outgrowth of leukemic clones (Marschalek et al, 1995.…”

“…Finally, the analysis of chromosomal breakpoints in t(4;11) cells was greatly facilitated by the improved knowledge of the breakpoint cluster regions of the MLL and AF-4 genes (Gu et al, 1994;Marschalek et al, 1995;Nilson et al, 1997;Reichel et al, 1998). Together with the new techniques described above (long range PCR,¯uorescence detection of restriction polymorphic fragments; (Repp et al, 1995;Leis et al, 1998) this combination of improvements enabled us to amplify and analyse chromosomal breakpoints in a reasonably short time.…”

“…The same is true for the SIL-TAL deletion involved in a frequent form of T-cell leukemias, where nonamer and heptamer sequences were identi®ed at the chromosomal breakpoints (Breit et al, 1993). None of these characteristic signs were observed for t(4;11) translocations in leukemia-derived cell lines and a limited number of primary blasts (Marschalek et al, 1995Reichel et al, 1998). From the limited evidence to date it was concluded, that the illegitimate recombination leading to these translocations was probably initiated by a severe DNA damage and subsequent defects in the DNA repair mechanism (Reichel et al, 1998).…”

“…The analysis was greatly facilitated by the recent improvement of our knowledge of the breakpoint cluster regions of the MLL and AF-4 genes (Gu et al, 1994;Marschalek et al, 1995;Nilson et al, 1997;Reichel et al, 1998). Based on this extended knowledge, it was possible to design a novel set of several dozen speci®c oligonucleotides and thus to amplify and sequence both reciprocal breakpoints for each patient.…”

Isolation and characterization of a Pufferfish MLL (Mixed lineage leukemia)-like gene (fMll) reveals evolutionary conservation in vertebrate genes related to Drosophila trithorax

“…The fMll gene has 34 exons while the human MLL gene has 36 exons (37 exons with the alternative splice form containing exon 1a, see Nilson 77 90 80 12 114 114 287 423 PHD 3 70 84 54 13 185 185 151 1095 PHD 3 64 89 75 14 174 174 251 1015 60 74 65 15 111 111 109 293 77 92 78 16 74 74 1766 932 70 76 72 17 215 194 88 368 66 80 72 18 95 107 75 1552 63 72 59 19 138 138 91 295 ZNF 74 96 85 20 159 159 153 745 70 89 77 21 112 118 86 413 71 92 82 22 79 79 199 1035 ATA1 73 88 85 23 161 161 111 519 64 70 61 24 147 186 128 539 46 47 31 25a 2596 The number of bases for each exon and each intron are given in the table for the Fugu and human MLL gene. Data for the human gene is based upon our own data and those published (Gu et al, 1994;Marschalek et al, 1995;Rasio et al, 1996). a The ®gures in parentheses are the coding region of the ®rst and last exons, the precise exon sizes inc...…”