The recurrent t(1;22)(p13;q13) translocation is exclusively associated with infant acute megakaryoblastic leukemia. We have identified the two genes involved in this translocation. Both genes possess related sequences in the Drosophila genome. The chromosome 22 gene (megakaryocytic acute leukemia, MAL) product is predicted to be involved in chromatin organization, and the chromosome 1 gene (one twenty-two, OTT) product is related to the Drosophila split-end (spen) family of proteins. Drosophila genetic experiments identified spen as involved in connecting the Raf and Hox pathways. Because almost all of the sequences and all of the identified domains of both OTT and MAL proteins are included in the predicted fusion protein, the OTT-MAL fusion could aberrantly modulate chromatin organization, Hox differentiation pathways, or extracellular signaling.

The most frequent oncogenic activation events characterized in childhood T acute lymphoblastic leukemia (T-ALL) result in the transcriptional activation of genes coding for transcription factors. The main genes are TAL1/SCL, a member of the basic region helix-loop-helix gene family, and HOX11L2, a member of the homeobox-containing protein family. To gain insight into the pathogenesis of this type of hematologic malignancy, we analyzed 28 T-ALL samples. SIL-TAL1/SCL fusion was detected in 6 patients; expression of HOX11L2 was observed in 6 patients and of HOX11 in 3 patients. With one exception, these activations did not occur simultaneously in the same patients, and they allowed the subclassification of 50% of the patients. SIL-TAL1 fusion was detected in association with HOX11 expression in one patient and with a t(8;14) (q24;q11) in another. High expression of LYL1, LMO2, or TAL1 was observed mainly in samples negative for HOX11L2 expression. HOX11L1 and HOX11 expression were observed in one instance each, in the absence of detectable chromosomal abnormality of their respective loci, on chromosomes 2 and 10, respectively. HOX11L2 expression was associated with a chromosome 5q abnormality, the location of the HOX11L2 locus in each case tested. Finally, our data show that HOX11L2 expression was a suitable marker for minimal residual disease follow-up and was significantly associated with relapse (P ‫؍‬ .02). (Blood. 2002;100:991-997)

The constitutive androstane receptor (CAR, NR1I3) transcriptionally activates cytochrome P450 2B6, 2C9, and 3A4 when activated by xenobiotics, such as phenobarbital. Information on the human CAR promoter was obtained by searching the NCBI human genome database. A contig (NT026945) corresponding to a fragment of chromosome 1q21 was found to contain the complete CAR gene. These data were confirmed using chromosomal in situ hybridization. Both primer extension and 5'-rapid amplification of the cDNA end PCR analysis were carried out to determine the transcriptional start site of human CAR, which was found to be 32 nucleotides downstream of a potential TATA box (CATAAAA). In addition, we found that the 5'-untranslated region of CAR mRNA is 110 nucleotides shorter than previously reported. Using genomic PCR, we amplified and cloned approximately 4.9 kb (-4711/+144) of the CAR gene promoter. The activity of this promoter was measured by transient transfection. Deletion analysis suggested the presence of a glucocorticoid responsive element in its distal region (-4477/-4410). From cotransfection experiments, mutagenesis, and gel shift assays, we identified a glucocorticoid response element at -4447/-4432 that was recognized and transactivated by the human glucocorticoid receptor. Finally, using the chromatin immunoprecipitation assay, we demonstrated that the glucocorticoid receptor binds to the distal region of CAR promoter in cultured hepatocytes only in the presence of dexamethasone. Identification of this functional element provides a rational mechanistic basis for CAR induction by glucocorticoids. CAR appears to be a primary glucocorticoid receptor-response gene.

The TEL͞ETV6 gene is located at 12p13 and encodes a member of the ETS family of transcription factors. Translocated ETS leukemia (TEL) is frequently involved in chromosomal translocations in human malignancies, usually resulting in the expression of fusion proteins between the amino-terminal part of TEL and either unrelated transcription factors or protein tyrosine kinases. We have characterized a t(1;12)(q21;p13) translocation in an acute myeloblastic leukemia (AML-M2). At the protein level, the untranslocated TEL copy and, as a result of the t(1;12) translocation, a fusion protein between TEL and essentially all of aryl hydrocarbon receptor nuclear translocator (ARNT) are expressed. The involvement of ARNT in human leukemogenesis has not been previously described. The ARNT protein belongs to a subfamily of the ''basic region helix-loop-helix'' (bHLH) protein that shares an additional region of similarity called the PAS (Per, ARNT, SIM) domain. ARNT is the central partner of several heterodimeric transcription factors, including those containing the aryl hydrocarbon (dioxin) receptor (AhR) and the hypoxia-inducible factor 1␣ (HIF1␣). Our results show that the TEL-ARNT fusion protein is the crucial product of the translocation and suggest that interference with the activity of AhR or HIF1␣ can contribute to leukemogenesis. C hromosomal abnormalities have been identified as an important step toward malignancy. Studies of solid and hematopoietic tumors have shown that chromosomal translocations usually result in the creation of chimeric genes and subsequent expression of fusion proteins. The translocated ETS leukemia (TEL) (also known as ETV6) gene, which encodes a member of the ETS family of transcription factors, is affected in more than half of the abnormalities of the short arm of chromosome 12 in various hematopoietic malignancies, and in solid tumors. Some of these abnormalities are specific for a leukemia subtype, whereas others are not. Perhaps the most intriguing characteristics of TEL-derived fusion proteins is that the contribution of TEL appears to be variable and the fusion partners are not functionally homogeneous because they encode protein tyrosine kinases or various unrelated transcription factors (for review see ref. 1).More precisely, following the description of the fusion of TEL to the platelet-derived growth factor receptor ␤ chain (PDGFR␤) gene, which is specific for chronic myelomonocytic leukemia (CMML) (2), the amino-terminal part of TEL has been found to be fused with other tyrosine kinases, such as ABL in atypical chronic myelocytic leukemia (aCML) and B acute lymphoblastic leukemia (ALL) (3, 4), JAK2 in T or B ALL and aCML (5, 6), and NTRK3 in acute myeloid leukemia (AML) and in congenital fibrosarcomas and renal tumors (7,8). In those situations, either 154 or 336 amino acids of the amino-terminal part of TEL, including a powerful oligomerization domain, are fused to these tyrosine kinases. The resulting fusion proteins exhibit constitutive tyrosine kinase activity and transforming pr...

Translocation t(1;22)(p13;q13) is associated with a peculiar subtype of acute megakaryocytic leukemia (M7) occurring in infants. We have recently characterized a fusion gene, OTT-MAL, resulting from this translocation. We now report three additional cases and show that this gene fusion is present in all five t(1;22) cases studied to date. Nucleotide sequence analysis of two translocation breakpoints suggests a nonhomologous end joining mechanism in the genesis of this translocation and reveals a noncanonical topoisomerase II-like consensus sequence within the OTT gene. FISH and PCR techniques described in this work are useful for identifying t(1;22) associated with M7.

Fluorescence in situ hybridization (FISH) studies were performed in three cases of acute lymphoblastic leukemia (ALL) with marker chromosomes to analyze the contribution of chromosome 21 in these markers. FISH with a chromosome 21 painting probe confirmed that chromosome 21 was involved in all three cases. FISH with YAC probes showed that the number of extra copies varied according to their location on chromosome 21. Attention was focused on the AML1 gene, which was present as five copies in most of the cells exhibiting the marker chromosomes. As controls, 11 cases of childhood ALL were studied with PAC probes covering AML1. The results agreed with the banded karyotypes in 10 patients. FISH uncovered a clone with four copies of AML1 which were only observed by FISH analysis of interphase nuclei in one patient. No point mutation was detected in exons 3-5, encoding the runt domain of AML1, in the three cases, suggesting an oncogenic role of wild-type AML1 amplification.

Cytogenetic and fluorescence in situ hybridization (FISH) analysis of 10 patients with various hematopoietic malignancies revealed the presence of dicentric chromosomes or pericentric chromosome rearrangements. Dicentrics were only ascertained by FISH studies in six patients. Two types of pericentric chromosome rearrangements have been observed: 'classical' dicentrics with two clearly separated centromeric regions, and more unusual rearrangements with a breakpoint within the centromeric or heterochromatic area, but outside the alphoid domain. FISH analysis of partial chromosome 1 q duplications present in three Burkitt lymphoma cell lines confirmed the partial involvement of the non-alphoid centromeric domain in the duplicated chromosome segment. The incidence of centromeric and pericentromeric rearrangements in hematopoietic malignancies may be higher than hitherto admitted. The chromosomal localization of these rearrangements suggests several mechanisms possibly involved in the malignant process and deserves more systematic study.

The TEL gene is involved in several chromosomal abnormalities of human hematopoietic malignancies. The chromosome 12 breakpoints frequently lie within the fifth intron of the gene, particularly in the most frequent translocation involving TEL, the t(12;21)(p13;q22). In order to search for a peculiar mechanism involved in the genesis of these translocations, we have established the sequence of two t(12;21) and a t(9;12)(q24;p13) breakpoints. Our data do not reveal the involvement of VDJ recombinase activity or Alu sequences but favor the occurrence of staggered breaks and DNA repair activity in the genesis of these translocations.