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Transcription factor E2F3a regulates CASP8AP2 transcription and enhances sensitivity to chemotherapeutic drugs in acute lymphoblastic leukemia.

ABSTRACT: Background:Low expression of E2F3a and caspase 8 associated protein 2 (CASP8AP2) are associated with poor prognosis of childhood acute lymphoblastic leukemia (ALL). Methods:Dual-luciferase reporter assay and wild type as well as four mutated types of reporter plasmids were used to demonstrate the activation of E2F3a on CASP8AP2 transcription. The direct binding of E2F3a with the promoter of CASP8AP2 was shown by Chromatin Immunoprecipitation (ChIP). Cell proliferation activity and cell cycle were determined by MTS and flow cytometry in leukemic cells after treating with common chemotherapeutic drugs vincristine and daunorubicin. Results:In this study, we found that up-regulation of E2F3a in leukemic cells led to increased fraction of cells in S and G2/M phase, accelerated proliferation, and enhanced sensitivity to vincristine and daunorubicin. ChIP and luciferase assay indicated that E2F3a could directly bind to two fragments in the wild type of CASP8AP2 promotor (-?206 to -?69 and -?677 to -?507), and activate its transcription activity which was reduced in mutated promotors. The effect of E2F3a on chemotherapeutic sensitivity of leukemic cells could be reversed by down-regulating CASP8AP2. Conclusions:E2F3a could promote transcription and expression of CASP8AP2. The effect of E2F3a on chemotherapeutic sensitivity of ALL cells was implemented by regulating CASP8AP2 expression to a great extent.

SUBMITTER: Liu FF

PROVIDER: S-EPMC5859744 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Transcription factor E2F3a regulates <i>CASP8AP2</i> transcription and enhances sensitivity to chemotherapeutic drugs in acute lymphoblastic leukemia.

Liu Fei-Fei FF Wang Kai-Ling KL Deng Li-Ping LP Liu Xiao X Wu Min-Yuan MY Wang Tian-You TY Cui Lei L Li Zhi-Gang ZG

Cancer cell international 20180320

<h4>Background</h4>Low expression of <i>E2F3a</i> and caspase 8 associated protein 2 (<i>CASP8AP2</i>) are associated with poor prognosis of childhood acute lymphoblastic leukemia (ALL).<h4>Methods</h4>Dual-luciferase reporter assay and wild type as well as four mutated types of reporter plasmids were used to demonstrate the activation of E2F3a on <i>CASP8AP2</i> transcription. The direct binding of E2F3a with the promoter of <i>CASP8AP2</i> was shown by Chromatin Immunoprecipitation (ChIP). Cel ...[more]

PMID: 29568235

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Project description:Acute lymphoblastic leukemia (ALL) is the most prevalent of pediatric cancers. Neuroepithelial cell-transforming 1 (NET1) has been associated with malignancy in a number of cancers, but the role of NET1 in ALL development is unclear. In the present study, we investigated the effect of NET1 gene in ALL cell proliferation and chemoresistance. We analyzed GEO microarray data comparing bone marrow expression profiles of pediatric B-cell ALL samples and those of age-matched controls. MTT and colony formation assays were performed to analyze cell proliferation. ELISA assays, Western blot analyses, and TUNEL staining were used to detect chemoresistance. We confirmed that NET1 was targeted by miR-206 using Western blot and luciferase reporter assays. We identified NET1 gene as one of the most significantly elevated genes in pediatric B-ALL. MTT and colony formation assays demonstrated that NET1 overexpression increases B-ALL cell proliferation in Nalm-6 cells. ELISA assays, Western blot analyses, and TUNEL staining showed that NET1 contributes to ALL cell doxorubicin resistance, whereas NET1 inhibition reduces resistance. Using the TargetScan database, we found that several microRNAs (miRNAs) were predicted to target NET1, including microRNA-206 (miR-206), which has been shown to regulate cancer development. To determine whether miR-206 targets NET1 in vitro, we transfected Nalm-6 cells with miR-206 or its inhibitor miR-206-in. Western blot assays showed that miR-206 inhibits NET1 expression and miR-206-in increases NET1 expression. Luciferase assays using wild-type or mutant 3'-untranslated region (3'-UTR) of NET1 confirmed these findings. We ultimately found that miR-206 inhibits B-ALL cell proliferation and chemoresistance induced by NET1. Taken together, our results provide the first evidence that NET1 enhances proliferation and chemoresistance in B-ALL cells and that miR-206 regulates these effects by targeting NET1. This study therefore not only contributes to a greater understanding of the molecular mechanisms underlying B-ALL progression but also opens the possibility for developing curative interventions.

Dataset Information

Transcription factor E2F3a regulates CASP8AP2 transcription and enhances sensitivity to chemotherapeutic drugs in acute lymphoblastic leukemia.

Publications

Transcription factor E2F3a regulates <i>CASP8AP2</i> transcription and enhances sensitivity to chemotherapeutic drugs in acute lymphoblastic leukemia.

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