Generation of Inducible BCL11B Knockout in TAL1/LMO1 Transgenic Mouse T Cell Leukemia/Lymphoma Model

Abstract

The B-cell CLL/lymphoma 11B gene (BCL11B) plays a crucial role in T-cell development, but its role in T-cell malignancies is still unclear. To study its role in the development of T-cell neoplasms, we generated an inducible BCL11B knockout in a murine T cell leukemia/lymphoma model. Mice, bearing human oncogenes TAL BHLH Transcription Factor 1 (TAL1; SCL) or LIM Domain Only 1 (LMO1), responsible for T-cell acute lymphoblastic leukemia (T-ALL) development, were crossed with BCL11B floxed and with CRE-ER/lox mice. The mice with a single oncogene BCL11Bflox/floxCREtg/tgTAL1tg or BCL11Bflox/floxCREtg/tgLMO1tg were healthy, bred normally, and were used to maintain the mice in culture. When crossed with each other, >90% of the double transgenic mice BCL11Bflox/floxCREtg/tgTAL1tgLMO1tg, within 3 to 6 months after birth, spontaneously developed T-cell leukemia/lymphoma. Upon administration of synthetic estrogen (tamoxifen), which binds to the estrogen receptor and activates the Cre recombinase, the BCL11B gene was knocked out by excision of its fourth exon from the genome. The mouse model of inducible BCL11B knockout we generated can be used to study the role of this gene in cancer development and the potential therapeutic effect of BCL11B inhibition in T-cell leukemia and lymphoma.

Keywords: BCL11B; CRE-ER/lox; LMO1; T-ALL; TAL1; TCL; mouse model.

Figure 1

Scheme of transgenic mice crossing to generate inducible BCL11B knockout in spontaneously developing T cell malignancies.

Figure 2

Establishment of BCL11B^flox/flox LMO1^tg and BCL11B^flox/flox TAL1^tg mice. (A) Generation of heterozygous BCL11B^flox/WTLMO1^tg and BCL11B^flox/WTTAL1^tg mice. Genotypes of the parents (F: female, M: male) are indicated on the top, the identification numbers of the progeny below, the photo the genotypes of the progeny below, the size of the DNA ladder in bp on the left, and the location of multiplex PCR products on the right. The BCL11B^flox/WTLMO1^tg and BCL11B^flox/WTTAL1^tg mice used for further crossing are in bold. MW 1: Molecular weight marker 1—GeneRuler 50 bp (Thermo Scientific, Waltham, MA, USA). (B) Generation of homozygous BCL11B^flox/foxLMO1^tg and BCL11B^flox/floxTAL1^tg mice. The homozygous BCL11B^flox/flox mice with either LMO1^tg or TAL1^tg oncogenes are in bold. The homozygous BCL11B^flox/flox mouse with both oncogenes is in bold and is indicated by an asterisk (*). MW 2: Molecular weight marker 2—GeneRuler 1 kb (Thermo Scientific).

Figure 3

Creation of the BCL11B^flox/delCRE^tg/tgTAL1^tg mice with germline hemizygous BCL11B knockout. Mice 643 and 645 are offspring of the tamoxifen-induced BCL11B knockout mother with LMO1^tg and untreated father. Mice 646 and 647 are offspring of the tamoxifen-induced BCL11B knockout mother and untreated father, both without oncogenes. MW 1: Molecular weight marker 1—GeneRuler 50 bp (Thermo Scientific).

Figure 4

Detection of tumors using magnetic resonance imaging. (A) Bilateral neck lymph node tumors, (B) abdominal lymph node tumor.

Figure 5

Peripheral blood smear at diagnosis. Magnification 1000×. (A) Normal lymphocyte, (B,C) lymphoblasts, (D) apoptotic cell.

Figure 6

Determination of malignant T cell immunophenotype by flow cytometry analysis. (A) Mouse 795 without signs of malignancy, (B) Mouse 742 with increased ratio of CD3+, CD4+/CD8+, CD25+/CD44-, and TdT+ cells. (C) Mouse 781 with increased ratio of CD3+, CD4+/CD8+, CD25+/CD44+ and NK-1.1 cells.