title: miR-15a and miR-16-1 inhibit the proliferation of leukemic cells by down-regulating WT1 protein level.
abstract: BACKGROUND
  miR-15a and miR-16-1(miR-15a/16-1) have been implicated as tumor suppressors in chronic lymphocytic leukemia, multiple myeloma, and acute myeloid leukemic cells. However the mechanism of inhibiting the proliferation of leukemic cells is poorly understood.
  
  
  METHODS
  K562 and HL-60 cells were transfected with pRS-15/16 or pRS-E, cell growth were measured by CCK-8 assay and direct cell count. Meanwhile WT1 protein and mRNA level were measured by Western blotting and quantitative real-time PCR.
  
  
  RESULTS
  In this study we found that over-expression of miR-15a/16-1 significantly inhibited K562 and HL-60 cells proliferation. Enforced expression of miR-15a/16-1 in K562 and HL-60 cells significantly reduced the protein level of WT1 but not affected the mRNA level. However enforced expression of miR-15a/16-1 can not reduce the activity of a luciferase reporter carrying the 3'-untranslated region(3'UTR) of WT1. Silencing of WT1 by specific siRNA suppressed leukemic cells proliferation resembling that of miR-15a/16-1 over-expression. Anti-miR-15a/16-1 oligonucleotides (AMO) reversed the expression of WT1 in K562 and HL-60 cells. Finally, we found a significant inverse correlation between miR-15a or miR-16-1 expression and WT1 protein levels in primary acute myeloid leukemia (AML) blasts and normal controls.
  
  
  CONCLUSIONS
  These data suggest that miR-15a/16-1 may function as a tumor suppressor to regulate leukemic cell proliferation potentially by down-regulating the WT1 oncogene. However WT1 is not directly targeted by miR-15a/16-1 through miRNA-mRNA base pairing, therefore more study are required to understand the mechanism by which miR-15a/16-1 downregulate WT1.
authors: Laboratory of Internal Medicine, The First Affiliated Hospital of Wenzhou Medical College, China.
authors: Shen-meng
authors: SM
authors: Gao
authors: Chong-yun
authors: CY
authors: Xing
authors: Chi-qi
authors: CQ
authors: Chen
authors: Si-si
authors: SS
authors: Lin
authors: Pei-hong
authors: PH
authors: Dong
authors: Fu-jun
authors: FJ
authors: Yu
mesh_terms: D020413:3' Untranslated Regions; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D015536:Down-Regulation; D015972:Gene Expression Regulation, Neoplastic; D018922:HL-60 Cells; D006801:Humans; D020014:K562 Cells; D007938:Leukemia / Q000235:genetics* / Q000378:metabolism*; D035683:MicroRNAs / Q000378:metabolism*; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger / Q000378:metabolism; D025721:WT1 Proteins / Q000378:metabolism*
chemical_list: D020413:3' Untranslated Regions; C505362:MIRN16 microRNA, human; D035683:MicroRNAs; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D025721:WT1 Proteins