Deregulated expression of TCL1 causes T cell leukemia in mice

Abstract

The TCL1 oncogene on human chromosome 14q32.1 is involved in the development of T cell leukemia in humans. These leukemias are classified either as T prolymphocytic leukemias, which occur very late in life, or as T chronic lymphocytic leukemias, which often arise in patients with ataxia telangiectasia (AT) at a young age. The TCL1 oncogene is activated in these leukemias by juxtaposition to the alpha or beta locus of the T cell receptor, caused by chromosomal translocations t(14:14)(q11:q32), t(7:14)(q35:q32), or by inversions inv(14)(q11:q32). To show that transcriptional alteration of TCL1 is causally involved in the generation of T cell neoplasia we have generated transgenic mice that carry the TCL1 gene under the transcriptional control of the p56(lck) promoter element. The lck-TCL1 transgenic mice developed mature T cell leukemias after a long latency period. Younger mice presented preleukemic T cell expansions expressing TCL1, and leukemias developed only at an older age. The phenotype of the murine leukemias is CD4-CD8+, in contrast to human leukemias, which are predominantly CD4+CD8-. These studies demonstrate that transcriptional activation of the TCL1 protooncogene can cause malignant transformation of T lymphocytes, indicating the role of TCL1 in the initiation of malignant transformation in T prolymphocytic leukemias and T chronic lymphocytic leukemias.

Figure 1

(A) DNA construct used in generating human lck-TCL1 transgenic mice. A 0.35-kb cDNA fragment containing the entire human TCL1 coding region was inserted into the BamHI site of a vector containing the lck proximal promoter and the hGH 3′ untranslated region and poly(A) addition site. (B) Southern blot analysis of DNA from tail clips from the first transgenic progeny. Blots were hybridized with the same DNA fragment used for injection of zygotes. Lane 1, DNA from a positive founder mouse; lanes 2–4, DNA from nontransgenic progeny.

Figure 2

Western blot analysis of protein extracts from transgenic (TG) and nontransgenic (non-TG) mouse tissues. Proteins (100 μg) were loaded per lane: 1, kidney; 2, thymus; 3, liver; and 4, spleen. The control lane, C, contains extracts from the pre-B leukemic cell line 697, which expresses high levels of the Tcl1 protein (5).

Figure 3

Whole spleen specimen isolated from a leukemic mouse (Left, TG 39) and a control mouse (Right).

Figure 4

Immunocytochemical analysis of Tcl1 protein in transgenic mouse spleens. Normal mouse spleen (negative control) stained for Tcl1 protein using polyclonal anti-Tcl1 antibody (A). A young transgenic mouse (<1 year old) stained for Tcl1 protein (B). Arrow in B shows a small cluster of positively stained cells. Small numbers of stained cells can also be observed scattered throughout the specimen. Spleen specimen from a representative animal containing extensive tumor infiltration stained with secondary antibody alone (C) or together with anti-Tcl1 polyclonal antibodies (D). A low (×10) magnification (E) and high (×40) magnification (F) depicting a foci of Tcl1-stained cells in pre- (or early) leukemic mice.

Figure 5

Southern analysis of TCR gene rearrangements in leukemias from transgenic mice. (A) Clonal TCR rearrangements. Southern blot analysis of DNA isolated from the spleens or thymuses of leukemic and nonleukemic mice. DNAs were digested with EcoRI. Blots were hybridized with a probe specific for the TCR Jβ₂ gene segment. Lanes: 1, spleen TG25; 2, spleen non-TG littermate; 3, spleen TG28 (partial digest); 4, spleen TG 43; 5, spleen TG 45; 6, spleen TG 47; 7, thymus TG 47; 8, spleen TG 39; and 10, thymus TG39. The strong 2.2-kb bands represent the TCR Jβ₂ gene in its germ-line configuration; arrows point to weaker hybridizing bands indicating the presence of clonal TCR gene rearrangements. (B) Expression of a specific TCR-Vβ on the surface of mouse leukemic cells. Single-cell suspensions of splenocytes from a representative tumor were stained with anti-Vβ6 antibodies (black) or with control antibody (FITC control, gray) followed by fluorescence-activated cell sorter analysis.