The Study Of TAL1 In IM-induced CML Cell Apoptosis And Erythroid Differentiation

Objective: Chronic myeloid leukemia(CML)is myeloproliferative neoplasia of hematopoietic stem cells that are characterized by the Philadelphia chromosome(Ph).Although tyrosine kinase inhibitors(TKIs)have dramatically improved survival in the treatment of CML patients,it failed to eliminate leukemia stem cells(LSCs),and CML patients usually relapse upon treatment discontinuation.Besides,some CML patients have drug resistance problems caused by other tumor-related gene mutations or long-term TKI treatment.T cell acute lymphoblastic leukemia 1(TAL1)is a transcription factor that is essential for hematopoietic process.TAL1 was downregulated during the maturation of hematopoietic stem/progenitor cells to T cells and eventually silenced in mature T cells.Abnormal expression in mature T cells is related to the occurrence of T-Acute lymphocytic leukemia(T-ALL).In this study,based on the characteristic of abnormal expression of TAL1 in CML found by the database,we analyze the expression of TAL1 in CMLLSCs and IM-induced CML cells;Next,we explore the mechanism of TAL1 in apoptosis and erythroid differentiation to understand the regulatory network of TAL1 and find new therapeutic targets for CML.Methods:(1)GEO database was used to analyze the expression of TAL1 in peripheral blood mononuclear cells(PBMCs)from CML patients and healthy individuals(HI).(2)LinCD34+CD38-CD26+ cells and Lin-CD34+CD38-CD26-cells were sorted from the bone marrow of de novo chronic phase CML by multicolor flow cytometry.(3)RNA interference(RNAi)was used to knock-down gene expression in K562,KBM5,KU812,JURKAT,TOM1,and SUPB15 cell lines.(4)Enrichment for signal pathway and the changes of related genes were described by RNA sequencing technology and DAVID(The Database for Annotation,Visualization and Integrated Discovery)bioinformatics resources.(5)q RT-PCR was used to detect the expression of TAL1,PTEN,lnc RNA-BGLT3,BCR-ABL,NFE2,γ-goblin,CD71,and CD235 A.(6)Western Blot was used to detect the protein levels of TAL1,PTEN,AKT,and p-AKT in samples with different treatments.(7)The effects of genes and drugs on cell apoptosis was detected by flow cytometry.Results:(1)In GEO databases,the expression of TAL1 in PBMCs of patients with newly diagnosed CML was significantly lower than that of healthy individuals.(2)The BCR-ABL fusion gene was positive,and the expression of TAL1 was low in Lin-CD34+CD38-CD26+ cells of patients with chronic CML.(3)Knocking down of the BCR-ABL fusion gene and IM treatment in K562 cells could up-regulate the expression of TAL1.(4)Down-regulation of TAL1 expression in K562 cells could significantly inhibit apoptosis and the reduction of AKT phosphorylation induced by IM.(5)The PTEN and lnc RNA-BGLT3 were up-regulated by IM treatment.(6)Ablation of TAL1 represses the expression of PTEN and lnc RNA-BGLT3 in K562 and KU812 cells,and there was significantly increased expression of PTEN and lnc RNA-BGLT3 in TAL1-overexpressing K562 cells.(7)IM-induced apoptosis was inhibited by down-regulation of PTEN or lnc RNA-BGLT3 in K562 cells.(8)Inhibition of the PI3K-AKT pathway by RNA interference or inhibitors could significantly promoted IM-induced apoptosis.(9)Different from CML cells,the expression of PTEN and lnc RNA-BGLT3 was inhibited by TAL1 in ALL cells.(10)The expression of TAL1,PTEN and lnc RNA-BGLT3 were up-regulated during the erythroid differentiation of cord blood CD34+ cells(CB-CD34+).(11)In K562 cells,the gene expression of NFE2,CD71,CD235 A and γ-goblin were up-regulated by the effect of low concentration of IM.(12)Loss of TAL1 or lnc RNA-BGLT3 in CB-CD34+ cells significantly inhibited the expression of CD71 and γ-goblin,but not CD235 A.(13)Knocking down of TAL1 or lnc RNABGLT3 in K562 cells significantly inhibited the IM-induced expression of NFE2,CD71,CD235 A and γ-goblin.Conclusion: In this study,we first discovered that TAL1 was low expressed in CML-LSCs and inhibited by the BCR-ABL fusion gene.We demonstrated that transcription factor TAL1 was down-regulated in CML-LSCs by BCR-ABL fusion gene.TAL1 knock-down could activated PI3K-AKT pathway by down-regulation of PTEN,leading to inhibition of IM-induced apoptosis;PTEN and lnc RNA-BGLT3 were potential target genes of TAL1 in CML;The sensitivity of CML cells to IM was increased by inhibiting PI3K-AKT pathway;Knocking-down TAL1 or lnc RNABGLT3 inhibited the erythroid differentiation of CB-CD34+ cells and K562 cells.This study reveals an unexpected contribution of TAL1 in CML and demonstrate that TAL1 may regulate PTEN expression and lead to inhibition of the PI3K-AKT pathway in TKI response.These results indicate that this may be a novel mechanism for the pathogenesis and treatment of CML,and this will provide new ideas for the treatment of CML.