Molecular analysis of acute promyelocytic leukemia breakpoint cluster region on chromosome 17

Acute promyelocytic leukemia (APL; FAB M3) is characterized by a predominance of malignant promyelocytes that carry a reciprocal translocation between the long arms of chromosomes 15 and 17, t(15;17) (q22;q11.2-q12). This translocation has become diagnostic for APL, as it is present in almost 100 percent of cases. A Not I linking clone was used to detect this translocation initially on pulsed-field gel electrophoresis and subsequently with conventional Southern (DNA) analysis. The breakpoints in ten APL cases examined were shown to cluster in a 12-kb region of chromosome 17, containing two CpG-rich islands. The region is the first intron of the retinoic acid receptor alpha gene (RARA).     

The role of the c-mos gene in the 8;21 translocation in human acute myeloblastic leukemia

The human c-mos proto-oncogene is located on chromosome 8 at band q22, close to the breakpoint in the t(8;21) (q22;q22) chromosome rearrangement. This translocation is associated with acute myeloblastic leukemia, subgroup M2. The c-myc gene, another proto-oncogene, has been mapped to 8q24. The breakpoint at 8q22 separates these genes, as determined by in situ hybridization of c-mos and c-myc probes. The c-mos gene remains on the 8q-chromosome and the c-myc gene is translocated to the 21q+ chromosome. Southern blot analysis of DNA from bone marrow cells of four patients with this translocation showed no rearrangement of c-mos.     

Gene for human insulin receptor: localization to site on chromosome 19 involved in pre-B-cell leukemia

Consistent chromosomal translocations in neoplastic cells may alter the expression of proto-oncogenes that are located near the breakpoints. The complementary DNA sequence of the human insulin receptor is similar to those of the EGF receptor (erbB oncogene) and products of the src family of oncogenes. With in situ hybridization and Southern blot analysis of somatic cell hybrid DNA, the human insulin receptor gene was mapped to the distal short arm of chromosome 19 (bands p13.2----p13.3), a site involved in a nonrandom translocation in pre-B-cell acute leukemia.     

The human gene encoding GM-CSF is at 5q21-q32, the chromosome region deleted in the 5q- anomaly

Human granulocyte-macrophage colony-stimulating factor (GM-CSF) is a 22,000-dalton glycoprotein that stimulates the growth of myeloid progenitor cells and acts directly on mature neutrophils. A full-length complementary DNA clone encoding human GM-CSF was used as a probe to screen a human genomic library and isolate the gene encoding human GM-CSF. The human GM-CSF gene is approximately 2.5 kilobase pairs in length with at least three intervening sequences. The GM-CSF gene was localized by somatic cell hybrid analysis and in situ hybridization to human chromosome region 5q21-5q32, which is involved in interstitial deletions in the 5q- syndrome and acute myelogenous leukemia. An established, human promyelocytic leukemia cell line, HL60, contains a rearranged, partially deleted GM-CSF allele and a candidate 5q- marker chromosome, indicating that the truncated GM-CSF allele may reside at the rejoining point for the interstitial deletion on the HL60 marker chromosome.     

Chromosomal localization of the human homolog (c-sis) of the simian sarcoma virus onc gene

Nonrandom chromosome rearrangements of chromosome 22 have been identified in different human malignancies. As a result of Southern blot hybridization of a c-sis probe to DNA's from mouse-human somatic cell hybrids, the human homolog (c-sis) of the transforming gene of simian sarcoma virus was assigned to chromosome 22. Hybrids between thymidine kinase-deficient mouse cells and human fibroblasts carrying a translocation of the region q11-qter of chromosome 22 to chromosome 17 were also analyzed. These studies demonstrate that the human c-sis gene is on region 22q11 greater than qter.     

Hu-ets-1 and Hu-ets-2 genes are transposed in acute leukemias with (4;11) and (8;21) translocations

Human probes identifying the cellular homologs of the v-ets gene, Hu-ets-1 and Hu-ets-2, and two panels of rodent-human cell hybrids were used to study specific translocations occurring in acute leukemias. The human ets-1 gene was found to translocate from chromosome 11 to 4 in the t(4;11)(q21;23), a translocation characteristic of a subtype of leukemia that represents the expansion of a myeloid/lymphoid precursor cell. Similarly, the human ets-2 gene was found to translocate from chromosome 21 to chromosome 8 in the t(8;21)(q22;q22), a nonrandom translocation commonly found in patients with acute myeloid leukemia with morphology M2 (AML-M2). Both translocations are associated with expression different from the expression in normal lymphoid cells of ets genes, raising the possibility that these genes play a role in the pathogenesis of these leukemias.     

Evidence for the involvement of GM-CSF and FMS in the deletion (5q) in myeloid disorders

By in situ chromosomal hybridization, the GM-CSF and FMS genes were localized to human chromosome 5 at bands q23 to q31, and at band 5q33, respectively. These genes encode proteins involved in the regulation of hematopoiesis, and are located within a chromosome region frequently deleted in patients with neoplastic myeloid disorders. Both genes were deleted in the 5q-chromosome from bone marrow cells of two patients with refractory anemia and a del(5)(q15q33.3). The GM-CSF gene alone was deleted in a third patient with acute nonlymphocytic leukemia (ANLL) who has a smaller deletion, del(5)(q22q33.1). Leukemia cells from a fourth patient who has ANLL and does not have a del(5q), but who has a rearranged chromosome 5 that is missing bands q31.3 to q33.1 [ins(21;5)(q22;q31.3q33.1)] were used to sublocalize these genes; both genes were present on the rearranged chromosome 5. Thus, the deletion of one or both of these genes may be important in the pathogenesis of myelodysplastic syndromes or of ANLL.     

Interferon and c-ets-1 genes in the translocation (9;11)(p22;q23) in human acute monocytic leukemia

Gene probes for interferons alpha and beta 1 and v-ets were hybridized to metaphase chromosomes from three patients with acute monocytic leukemia who had a chromosomal translocation, t(9;11)(p22;q23). The break in the short arm of chromosome 9 split the interferon genes, and the interferon-beta 1 gene was translocated to chromosome 11. The c-ets-1 gene was translocated from chromosome 11 to the short arm of chromosome 9 adjacent to the interferon genes. No DNA rearrangement was observed when these probes were hybridized to genomic DNA from leukemic cells of two of the patients. The results suggest that the juxtaposition of the interferon and c-ets-1 genes may be involved in the pathogenesis of human monocytic leukemia.     

A common mechanism of chromosomal translocation in T- and B-cell neoplasia

The chromosomal breakpoint involved in the t(8;14)(q24;q11) chromosome translocation in the SKW-3 cell line, which directly involves the 3' flanking region of the c-myc gene, was cloned and sequenced. The breakpoint on chromosome 8 mapped to a position 3 kb 3' of c-myc while the chromosome 14 breakpoint occurred 36 kb 5' of the gene for the constant region of the alpha chain of the T-cell receptor (TCR). The translocation resulted in a precise rearrangement of sequences on chromosome 8 and what appears to be a functional J alpha segment on chromosome 14. Signal sequences for V-J joining occurred at the breakpoint positions on both chromosomes 14 and 8, suggesting that the translocation occurs during TCR gene rearrangement and that it is catalyzed by the enzymatic systems involved in V-J joining reactions. The involvement of c-myc in the translocation and the association of joining signals at the breakpoints provides a parallel to the situation observed in the translocations involving c-myc and the immunoglobulin loci in B-cell neoplasms and suggests that common mechanisms of translocation and oncogene deregulation are involved in B- and T-cell malignancies.     

Detection of bcr-abl fusion in chronic myelogeneous leukemia by in situ hybridization

Chronic myelogeneous leukemia (CML) is genetically characterized by fusion of the bcr and abl genes on chromosomes 22 and 9, respectively. In most cases, the fusion involves a reciprocal translocation t(9;22)(q34;q11), which produces the cytogenetically distinctive Philadelphia chromosome (Ph1). Fusion can be detected by Southern (DNA) analysis or by in vitro amplification of the messenger RNA from the fusion gene with polymerase chain reaction (PCR). These techniques are sensitive but cannot be applied to single cells. Two-color fluorescence in situ hybridization (FISH) was used with probes from portions of the bcr and abl genes to detect the bcr-abl fusion in individual blood and bone marrow cells from six patients. The fusion event was detected in all samples analyzed, of which three were cytogenetically Ph1-negative. One of the Ph1-negative samples was also PCR-negative. This approach is fast and sensitive, and provides potential for determining the frequency of the abnormality in different cell lineages.     

Clustering of leukocyte and fibroblast interferon genes of human chromosome 9

At least ten leukocyte interferon genes and the single known fibroblast interferon gene have been localized on the pter leads to q12 region of human chromosome 9. Gene mapping was accomplished by blot hybridization of cloned interferon complementary DNA to DNA from human-mouse cell hybrids with a translocation involving human chromosome 9. Supporting evidence suggests these genes are clustered.     

Locus of the alpha-chain of the T-cell receptor is split by chromosome translocation in T-cell leukemias

Mouse lymphoma cells were hybridized with two human acute T-cell leukemias with a t(11;14) (p13;q11) translocation and the segregated hybrids were examined for the presence of the DNA segments coding for the constant (C) and the variable (V) regions of the alpha chain (C alpha and V alpha) of the T-cell receptor. The C alpha segment was translocated to the involved chromosome 11 (11p+) while the V alpha segment remained on the involved chromosome 14 (14q-). The data indicate that the locus for the alpha chain of the T-cell receptor is split by the chromosomal breakpoint between the V alpha and the C alpha gene segments, and that the V alpha segments are proximal to the C alpha segment within chromosome band 14q11.2.     

Physical mapping of a translocation breakpoint in neurofibromatosis

The gene for von Recklinghausen neurofibromatosis (NF1), one of the most common autosomal-dominant disorders of humans, was recently mapped to chromosome 17 by linkage analysis. The identification of two NF1 patients with balanced translocations that involved chromosome 17q11.2 suggests that the disease can arise by gross rearrangement of the NF1 locus, and that the NF1 gene might be identified by cloning the region around these translocation breakpoints. To further define the region of these translocations, a series of chromosome 17 Not I-linking clones has been mapped to proximal 17q and studied by pulsed-field gel electrophoresis. One clone, 17L1 (D17S133), clearly identifies the breakpoint in an NF1 patient with a t(1;17) translocation. A 2.3-megabase pulsed-field map of this region was constructed and indicates that the NF1 breakpoint is only 10 to 240 kilobases away from 17L1. This finding prepares the way for the cloning of NF1.     

Molecular analysis of the t(2;14) translocation of childhood chronic lymphocytic leukemia

Two rare cases of chronic lymphocytic leukemia (CLL) in children have been studied; both are associated with a previously undescribed chromosomal translocation [t(2;14) (p13;q32)]. In one patient the translocation was reciprocal and the breakpoint on chromosome 14 occurred just 5' of the C gamma 2 region on the productive immunoglobulin heavy-chain allele. The breakpoint on chromosome 2 does not involve the K locus but lies within an uncharacterized region that coincides with the position of a constitutive fragile site that occurs within normal lymphocytes. Data on the second patient are consistent with these findings and suggest that these cases represent a rare but distinct subgroup of CLL's with a specific cytogenetic change.     

The human homologs of the raf (mil) oncogene are located on human chromosomes 3 and 4

Two human genes that are homologous to both the murine transforming gene (oncogene) v-raf and the chicken transforming gene v-mil have been mapped by means of human-rodent somatic cell hybrids to human chromosomes previously devoid of known oncogenes. One gene, c-raf-2, which appears to be a processed pseudogene, is located on chromosome 4. The other gene, c-raf-1, which appears to be the active gene, is located on chromosome 3 and has been regionally mapped by chromosomal in situ hybridization to 3p25. This assignment correlates with specific chromosomal abnormalities associated with certain human malignancies.     

Gene for T-cell growth factor: location on human chromosome 4q and feline chromosome B1

T-cell growth factor (TCGF) or interleukin-2 (IL-2), an immunoregulatory lymphokine, is produced by lectin- or antigen-activated mature T lymphocytes and in a constitutive manner by certain T-cell lymphoma cell lines. By means of a molecular clone of human TCGF and DNA extracted from a panel of somatic cell hybrids (rodent cells X normal human lymphocytes), the TCGF structural gene was identified on human chromosome 4. In situ hybridization of the TCGF clone to human chromosomes resulted in significant labeling of the midportion of the long arm of chromosome 4, indicating that the TCGF gene was located at band q26-28. Genomic DNA from a panel of hybrids prepared with HUT-102 B2 cells was examined with the same molecular clone. In this clone of cells, which produces human T-cell leukemia virus, the TCGF gene was also located on chromosome 4 and was apparently not rearranged. The homologous TCGF locus in the domestic cat was assigned to chromosome B1 by using a somatic cell hybrid panel that segregates cat chromosomes. Linkage studies as well as high-resolution G-trypsin banding indicate that this feline chromosome is partially homologous to human chromosome 4.     

Mapping of the human Blym-1 transforming gene activated in Burkitt lymphomas to chromosome 1

Blym-1, a transforming gene detected by transfection of NIH 3T3 cells with DNA from Burkitt lymphomas, was mapped to the short arm of chromosome 1 (1p32) by chromosomal in situ hybridization. The Blym-1 gene was not physically linked to the cellular myc oncogene or to any of the immunoglobulin gene loci implicated in the characteristic chromosomal translocations in Burkitt lymphoma.