TEL/AML1 Shows Dominant-Negative Effects over TEL as Well as AML1.
Authors: Waga Kazuo , Gunji Hisako , Nakamura Fumihiko , Maki Kazuhiro , Sasaki Ko , Nakamura Yuichi , Mitani Kinuko 
Published: October 2019

The TEL/AML1 chimeric gene is generated by the t(12;21) translocation in pre-B cell acute lymphoblastic leukemia. TEL/AML1 consists of the helix-loop-helix (HLH) dimerization domain from TEL and almost the entire of AML1, but loses the ETS DNA-binding domain from TEL. Dominant-negative effects of TEL/AML1 over wild-type-AML1 are believed to trigger the development of this type of leukemia. However, it could also be possible that TEL/AML1 affects wild-type-TEL’s molecular and tumor suppressive functions through the HLH domain. To test this hypothesis, we investigated molecular and biological effects of TEL/AML1 on wild-type-TEL.

Since TEL/AML1 contains the HLH domain from TEL, we first tested whether TEL/AML1 associates with wild-type-TEL by immunoprecipitation analysis. Wild-type-TEL was co-immunoprecipitated with TEL/AML1, but not with a TEL/AML1 mutant lacking the HLH domain. This suggests that TEL/AML1 associates with wild-type-TEL depending on the HLH domain and might affect its functions. To clarify functional alterations of TEL/AML1, we compared molecular function of wild-type-TEL and TEL/AML1 in DNA-binding and transcriptional reporter assays. Wild-type-TEL bound to ETS-binding site (EBS) and showed transcriptional repression on stromelysin-1 promoter containing EBS. On the other hand, TEL/AML1 failed to bind to EBS and showed no transcriptional repression on it. Moreover, TEL/AML1 showed a dominant-negative effect over the wild-type-TEL-mediated transcriptional repression. These data suggest that TEL/AML1 loses wild-type-TEL’s molecular functions, but works as a dominant-negative molecule over wild-type-TEL. To further investigate TEL/AML1’s biological effects over wild-type-TEL, we established NIH3T3 stable cell lines expressing wild-type-AML1, wild-type-TEL or TEL/AML1 and performed colony assays. TEL/AML1-expressing cells produced approximately the same numbers of colonies as the mock cells. Wild-type-AML1-expressing cells generated much more colonies, while wild-type-TEL-expressing cells generated less colonies. We further introduced TEL/AML1 cDNA into the wild-type-AML1- and the wild-type-TEL-expressing cells. Co-expression of TEL/AML1 releaved the wild-type-TEL’s growth-suppression as well as the wild-type-AML1’s growth-stimulation. Finally, we counted cell numbers of the same set of NIH3T3 stable cell lines in liquid culture and obtained the similar results as in soft agar culture.

In this study we demonstrate that TEL/AML1 dominantly inhibits both molecular and biological functions of wild-type-TEL, possibly through heterodimerizing with it. Therefore, appearance of the t(12;21) translocation could lead to not only inactivation of wild-type-AML1 but also inactivation of wild-type-TEL. As TEL is a tumor suppressor, functional loss of wild-type-TEL seems to be one hit for the development of leukemia carrying the t(12;21) translocation. However, it has been reported that expression of TEL/AML1 is not sufficient to fully transform hematopoietic cells in vitro and in vivo and requires other hits to cause leukemia. Since deletion of the non-translocated TEL allele is associated in the vast majority of leukemia with the t(12;21) translocation, complete inactivation of wild-type-TEL by the allelic loss could be necessary to be the second hit that provides a further proliferative advantage to leukemic cells.