miR-16 family induces cell cycle arrest by regulating multiple cell cycle genes.
MicroRNAs (miRNAs) are a class of small regulatory RNAs that are thought to be involved in diverse biological processes by regulating gene expression. Numerous miRNAs have been identified in various species, and many more miRNAs remain to be detected. Generally, hundreds of mRNAs have been predicted to be potential targets of one miRNA, so it is a great challenge to identify the genuine miRNA targets. Here, we generated the cell lines depleted of Drosha protein and screened dozens of transcripts (including Cyclin D1) regulated potentially by miRNA-mediated RNA silencing pathway. On the basis of miRNA expressing library, we established a miRNA targets reverse screening method by using luciferase reporter assay. By this method, we found that the expression of Cyclin D1 (CCND1) was regulated by miR-16 family directly, and miR-16 induced G1 arrest in A549 cells partially by CCND1. Furthermore, several other cell cycle genes were revealed to be regulated by miR-16 family, including Cyclin D3 (CCND3), Cyclin E1 (CCNE1) and CDK6. Taken together, our data suggests that miR-16 family triggers an accumulation of cells in G0/G1 by silencing multiple cell cycle genes simultaneously, rather than the individual target.
Liu
Qin
Q
Beijing Institute of Radiation Medicine, 27 Taiping Road, Beijing 100850, People's Republic of China.
Fu
Hanjiang
H
Sun
Fang
F
Zhang
Haoming
H
Tie
Yi
Y
Zhu
Jie
J
Xing
Ruiyun
R
Sun
Zhixian
Z
Zheng
Xiaofei
X
D002453:Cell Cycle / Q000235:genetics; D002460:Cell Line; D019938:Cyclin D1 / Q000235:genetics; D056743:Cyclin D3; D019927:Cyclin E / Q000235:genetics; D051361:Cyclin-Dependent Kinase 6 / Q000235:genetics; D016213:Cyclins / Q000235:genetics; D016193:G1 Phase / Q000235:genetics; D018816:Genes, cdc; D006801:Humans; D008156:Luciferases / Q000032:analysis; D035683:MicroRNAs / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis; D015513:Oncogene Proteins / Q000235:genetics; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D043244:Ribonuclease III / Q000235:genetics / Q000378:metabolism
C534696:CCND3 protein, human; C475820:CCNE1 protein, human; D056743:Cyclin D3; D019927:Cyclin E; D016213:Cyclins; C505362:MIRN16 microRNA, human; D035683:MicroRNAs; D015513:Oncogene Proteins; D012333:RNA, Messenger; D019938:Cyclin D1; D008156:Luciferases; D051361:Cyclin-Dependent Kinase 6; C517918:DROSHA protein, human; D043244:Ribonuclease III