title: TALENs-mediated gene disruption of FLT3 in leukemia cells: Using genome-editing approach for exploring the molecular basis of gene abnormality.
abstract: Novel analytic tools are needed to elucidate the molecular basis of leukemia-relevant gene mutations in the post-genome era. We generated isogenic leukemia cell clones in which the FLT3 gene was disrupted in a single allele using TALENs. Isogenic clones with mono-allelic disrupted FLT3 were compared to an isogenic wild-type control clone and parental leukemia cells for transcriptional expression, downstream FLT3 signaling and proliferation capacity. The global gene expression profiles of mutant K562 clones and corresponding wild-type controls were compared using RNA-seq. The transcriptional levels and the ligand-dependent autophosphorylation of FLT3 were decreased in the mutant clones. TALENs-mediated FLT3 haplo-insufficiency impaired cell proliferation and colony formation in vitro. These inhibitory effects were maintained in vivo, improving the survival of NOD/SCID mice transplanted with mutant K562 clones. Cluster analysis revealed that the gene expression pattern of isogenic clones was determined by the FLT3 mutant status rather than the deviation among individual isogenic clones. Differentially expressed genes between the mutant and wild-type clones revealed an activation of nonsense-mediated decay pathway in mutant K562 clones as well as an inhibited FLT3 signaling. Our data support that this genome-editing approach is a robust and generally applicable platform to explore the molecular bases of gene mutations.
authors.affiliation: Department of hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Jue
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Tongjuan
authors.initials: T
authors.lastname: Li
authors.affiliation: Department of hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Mi
authors.initials: M
authors.lastname: Zhou
authors.affiliation: Cancer Biology Research Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Zheng
authors.initials: Z
authors.lastname: Hu
authors.affiliation: Department of hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Xiaoxi
authors.initials: X
authors.lastname: Zhou
authors.affiliation: Department of hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Shiqiu
authors.initials: S
authors.lastname: Zhou
authors.affiliation: Department of hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Na
authors.initials: N
authors.lastname: Wang
authors.affiliation: Department of hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Liang
authors.initials: L
authors.lastname: Huang
authors.affiliation: Department of hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Zhao
authors.affiliation: Department of hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Cao
authors.affiliation: Department of hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Min
authors.initials: M
authors.lastname: Xiao
authors.affiliation: Cancer Biology Research Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Ding
authors.initials: D
authors.lastname: Ma
authors.affiliation: Wuhan YZY Bio-Pharma Co., Ltd., Wuhan, Hubei, China.
authors.forename: Pengfei
authors.initials: P
authors.lastname: Zhou
authors.affiliation: Department of hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Zhen
authors.initials: Z
authors.lastname: Shang
authors.affiliation: Department of hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.|Cancer Biology Research Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors.forename: Jianfeng
authors.initials: J
authors.lastname: Zhou
mesh_terms: D000818:Animals; D018389:Codon, Nonsense; D057895:Haploinsufficiency; D006801:Humans; D019169:Jurkat Cells; D020014:K562 Cells; D007938:Leukemia / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D015398:Signal Transduction / Q000235:genetics*; D051941:fms-Like Tyrosine Kinase 3 / Q000235:genetics* / Q000378:metabolism
chemical_list: D018389:Codon, Nonsense; C497970:FLT3 protein, human; D051941:fms-Like Tyrosine Kinase 3