No abstract

Transcriptional activation of the tal‐1 gene occurs in ‐30% of patients with T cell Acute Lymphoblastic Leukemia and is therefore likely to be involved in human T cell leukemogenesis. However, the TAL‐1 protein functional properties involved in this process have not been assessed so far. We have derived a clonal subline of the Jurkat T cell line which produced solely a mutant truncated form of TAL‐1 protein. Sequencing of genomic DNA and cDNAs showed that the only transcribed tal‐1 allele of this mutant subline harbored a G nucleotide insertion at codon 270. The resulting frameshift modifies TAL‐1 residues 272‐278 and creates a stop at codon 279. Although the deletion of the 53 carboxy‐terminal residues of the TAL‐1 protein did not directly affect the TAL‐1 basic helix‐loop‐helix domain (residues 185‐243), it had drastic effects on TAL‐1 functional properties, since the mutant subline exhibited a dramatic decrease of protein binding activity to the TAL‐1 DNA consensus sequence. Growth curves indicated that the mutant subline exhibited premature apoptosis upon medium depletion or serum reduction when compared with the parental cells. However, no difference between Jurkat and the mutant subline was observed in etoposide‐ or Fas/APO‐1‐triggered apoptosis. Stable expression of the mutant TAL‐1 protein in Jurkat cells resulted in a phenotype that was similar to that of the mutant Jurkat subline, indicating that the TAL‐1 mutant protein behaved like a dominant negative mutant and that the premature apoptosis of the mutant subline upon medium depletion was the consequence of the loss of TAL‐1 protein activity.

Rearrangement of the tal-1 gene (also known as SCL or TCL-5) occurs in at least 25% of T-cell acute lymphoblastic leukemias (T-ALLs) and results in the aberrant expression of tal-1 mRNA in the neoplastic cells. Also, tal-1 mRNA is constitutively expressed in erythroid precursors and megakaryocytes. This report describes a direct immunocytochemical study of the distribution and localization of TAL-1 protein in normal human tissues and cell lines using four monoclonal antibodies raised against recombinant TAL-1 proteins. One of these reagents recognizes a protein of 41 kD molecular weight in in vitro- translated TAL-1 proteins, two others recognize proteins of 39 and 41 kD molecular weight, and the fourth antibody also recognizes a TAL-1 protein of 22 kD in addition to the 39- and 41-kD proteins. These anti- TAL-1 antibodies label the nuclei of erythroid precursor cells and megakaryocytes in fetal liver and adult bone marrow. The punctate pattern of nuclear labeling suggests that TAL-1 may comprise part of a novel nuclear structure, similar to that recently found for the PML protein. The nuclei of T cell lines known to express mRNA encoding the full-length TAL-1 protein (eg, CCRF-CEM, RPMI 8402, and Jurkat) are also labeled. A study of normal human tissues (including thymus) showed labeling of smooth muscle, some tissue macrophages, and endothelial cells. TAL-1 protein is undetectable in other cell types. These reagents may play an important role in the diagnosis of T-ALL and could also be used in the context of lymphoma diagnosis on routinely fixed material.

SummaryA common feature ofT cell acute lymphoblastic leukemias (T-ALLs) is the presence of structural alteration of the 5' part of the tal-1 locus, localized on chromosomal band 1p32. These alterations consist of either a t(1;14)(p32;q11) chromosomal translocation (3% of T-ALLs) or tal d submicroscopic deletion (12-25% additional T-ALLs). We have characterized a case of T-ALL with t(1;14)(p32;q11) in which, unlike the majority of t (1;14), the recombination with the T cell receptor/~ elements affected the 3' side of the tat-1 locus. In this case, tal-1 transcription is initiated from a promoter located within the fourth exon similarly to the DU 528 cell line. In a T-ALL bearing a t(1;14) affecting the 5' part of ta/-1, two types of tal-1 transcripts were observed, namely those probably initiated from the D8 region juxtaposed to tal-1 by the translocation, and those from the exon 4 promoter. It is interesting that this exon 4 promotion was also found in leukemic T cell lines and T-ALL samples without apparent tal-1 genomic alteration. In contrast, no transcript initiated from the exon 4 promoter was found in T-ALL with tal dl or tal d2 deletion. In these cells, tal-1 is expressed via Sllctal-1 fused transcripts. Finally, this exon 4 initiation was detected neither in normal bone marrow, nor in malignant cells from the erythroid/megakaryocytic lineages. Taken as a whole, these data suggest that the exon 4 promoter is specifically active in T cell lineage.

The basic helix-loop-helix tal-1 gene plays a key role in hematopoiesis, and its expression is tightly controlled through alternative promoters and complex interactions of cis-acting regulatory elements. tal-1 is not expressed in normal T cells, but its transcription is constitutive in a large proportion of human T cell leukemias. We have previously described a downstream initiation of tal-1 transcription specifically associated with a subset of T cell leukemias that leads to the production of NH 2 -truncated TAL-1 proteins. In this study, we characterize the human promoter (promoter IV), embedded within a GC-rich region in exon IV, responsible for this transcriptional activity. The restriction of promoter IV usage is assured by a novel silencer element in the 3-unstranslated region of the human gene that represses its activity in erythroid but not in T cells. The silencer activity is mediated through binding of a tissue-specific nuclear factor to a novel protein recognition motif (designated tal-RE) in the silencer. Mutation of a single residue within the tal-RE abolishes both specific protein binding and silencing activity. Altogether, our results demonstrate that the tal-1 promoter IV is actively repressed in cells of the erythro-megakaryocytic lineage and that this repression is released in leukemic T cells, resulting in the expression of the tal-1 truncated transcript.