The Groucho Corepressors, TLE1 and TLE4, Are Candidate Del(9q) AML Tumor Suppressor Genes That Can Complement AML1-ETO.

Abstract

Deletion of the long arm of chromosome 9, del(9q), is one of the most common cytogenetic abnormalities seen with t(8;21) AML. This close association suggests that that the loss of a critical gene(s) on chr9q cooperates with the AML1-ETO (RUNX1-MTG8) fusion gene produced by t(8;21) in leukemogenesis. We recently mapped the commonly deleted region of del(9q) AML to less than 2.4 Mb and identified all the genes mapping to this region. Extensive sequence analysis of these genes failed to reveal obvious mutations in del(9q) AML samples although the expression of a number of these genes appeared low specifically in del(9q) samples (Genes Chr and Ca 44, 279–291). To help determine the critical affected genes in the del(9q) CDR we used an siRNA expression library directed against all del(9q) CDR gene transcripts in an in vitro complementation assay with AML1-ETO. Inducible expression of AML1-ETO in U937 cells (U937T-A/E, Mol Cell Biol 21, 5577–5590) leads to cell cycle arrest and apoptosis. We introduced the del(9q) CDR siRNA library into U937T-A/E cells with the goal of finding siRNAs that rescued these cells after induction of AML1-ETO expression. The most commonly recovered siRNA using flanking PCR primers was directed against TLE1. Transducin-like enhancer of split (TLE) 1 and TLE4, are two members of the Groucho family of co-repressors that map adjacent to the del(9q) CDR. We subsequently demonstrated that knockdown of either TLE1 or TLE4 with specific siRNAs were able to overcome the induced cell cycle arrest in U937T-A/E cells. We further demonstrate that knockdown of either TLE in the AML1-ETO+ Kasumi-1 cell line increased cell division, increased expression of cyclin D1 and Ki67 and reduced the G0/G1 fraction, while expression of TLE1 or TLE4 in Kasumi-1 cells induced cell cycle arrest and apoptosis. Understanding how various mutations work cooperatively to produce a malignant phenotype is one of the great challenges in oncology. The TLEs are known to inhibit NFKB as well as Wnt signaling, two pathways implicated leukemic stem cell expansion. TLE inhibition may represent an important cooperating mutation with AML11-ETO in AML that links Wnt signaling and core binding protein transcription factors. Studies are currently underway to demonstrate this cooperativity and more directly evaluate the role of TLE inhibition in leukemogenesis.