TY - JOUR
T1 - Global analysis of AGO2-bound RNAs reveals that miRNAs induce cleavage of target RNAs with limited complementarity
AU - Jung, Eunsun
AU - Seong, Youngmo
AU - Jeon, Bohyun
AU - Song, Hoseok
AU - Kwon, Young Soo
N1 - Funding Information:
This work was supported by Ministry of Education, Science and Technology through the National Research Foundation of Korea [ NRF-2011-0024962 and NRF-2009-0068994 to Y.-S.K.; 2014R1A1A2058610 to H.S.], and Ministry of Health and Welfare [ HI15C2835 to H.S.].
Publisher Copyright:
© 2017 Elsevier B.V.
PY - 2017/11
Y1 - 2017/11
N2 - Among the four Argonaute family members in mammals, only AGO2 protein retains endonuclease activity and facilitates cleavage of target RNAs base-pairing with highly complementary guide RNAs. Despite the deeply conserved catalytic activity, only a small number of targets have been reported to extensively base pair with cognate miRNAs to be cleaved by AGO2. Here, we analyzed AGO2-bound RNAs by CrossLinking ImmunoPrecipitation (CLIP) of genetically modified cells that express epitope-tagged AGO2 from the native genomic locus. We found that HMGA2 mRNA is cleaved by AGO2 loaded with let-7 and miR-21. In contrast to the generally accepted notion, the base-pairing from the seed region to the cleavage site, rather than perfect or near perfect complementarity, was required for cleavage of the target mRNA in cells. Non-templated addition of nucleotides at the 3′ end of the cleaved RNA was observed, further supporting the AGO2-mediated cleavage. Based on the observation that the limited complementarity is the minimum requirement for cleavage, we found that AGO2-mediated cleavage of targets is more common than previously thought. Our result may explain the vital role of endonuclease activity in controlling miRNA-mediated gene regulation.
AB - Among the four Argonaute family members in mammals, only AGO2 protein retains endonuclease activity and facilitates cleavage of target RNAs base-pairing with highly complementary guide RNAs. Despite the deeply conserved catalytic activity, only a small number of targets have been reported to extensively base pair with cognate miRNAs to be cleaved by AGO2. Here, we analyzed AGO2-bound RNAs by CrossLinking ImmunoPrecipitation (CLIP) of genetically modified cells that express epitope-tagged AGO2 from the native genomic locus. We found that HMGA2 mRNA is cleaved by AGO2 loaded with let-7 and miR-21. In contrast to the generally accepted notion, the base-pairing from the seed region to the cleavage site, rather than perfect or near perfect complementarity, was required for cleavage of the target mRNA in cells. Non-templated addition of nucleotides at the 3′ end of the cleaved RNA was observed, further supporting the AGO2-mediated cleavage. Based on the observation that the limited complementarity is the minimum requirement for cleavage, we found that AGO2-mediated cleavage of targets is more common than previously thought. Our result may explain the vital role of endonuclease activity in controlling miRNA-mediated gene regulation.
KW - AGO2 catalytic activity
KW - CrossLinking ImmunoPrecipitation
KW - HMGA2
KW - MicroRNA
KW - RNA cleavage
UR - http://www.scopus.com/inward/record.url?scp=85032872933&partnerID=8YFLogxK
U2 - 10.1016/j.bbagrm.2017.10.001
DO - 10.1016/j.bbagrm.2017.10.001
M3 - Article
C2 - 29031931
AN - SCOPUS:85032872933
SN - 1874-9399
VL - 1860
SP - 1148
EP - 1158
JO - Biochimica et Biophysica Acta - Gene Regulatory Mechanisms
JF - Biochimica et Biophysica Acta - Gene Regulatory Mechanisms
IS - 11
ER -