The endonuclease activity of Mili fuels piRNA amplification that silences LINE1 elements

TY - JOUR

T1 - The endonuclease activity of Mili fuels piRNA amplification that silences LINE1 elements

AU - De Fazio, S.

AU - Di Giacomo, M.

AU - Sankar, A.

AU - Moreira, P.N.

AU - O'carroll, D.

AU - Bartonicek, N.

AU - Abreu-Goodger, C.

AU - Enright, A.J.

AU - Funaya, C.

AU - Antony, C.

PY - 2011/12/8

Y1 - 2011/12/8

N2 - Piwi proteins and Piwi-interacting RNAs (piRNAs) have conserved functions in transposon silencing. The murine Piwi proteins Mili and Miwi2 (also called Piwil2 and Piwil4, respectively) direct epigenetic LINE1 and intracisternal A particle transposon silencing during genome reprogramming in the embryonic male germ line. Piwi proteins are proposed to be piRNA-guided endonucleases that initiate secondary piRNA biogenesis; however, the actual contribution of their endonuclease activities to piRNA biogenesis and transposon silencing remain unknown. To investigate the role of Piwi-catalysed endonucleolytic activity, we engineered point mutations in mice that substitute the second aspartic acid to an alanine in the DDH catalytic triad of Mili and Miwi2, generating the Mili DAH and Miwi2 DAH alleles, respectively. Analysis of Mili-bound piRNAs from homozygous Mili DAH fetal gonadocytes revealed a failure of transposon piRNA amplification, resulting in the marked reduction of piRNA bound within Miwi2 ribonuclear particles. We find that Mili-mediated piRNA amplification is selectively required for LINE1, but not intracisternal A particle, silencing. The defective piRNA pathway in Mili DAH mice results in spermatogenic failure and sterility. Surprisingly, homozygous Miwi2 DAH mice are fertile, transposon silencing is established normally and no defects in secondary piRNA biogenesis are observed. In addition, the hallmarks of piRNA amplification are observed in Miwi2-deficient gonadocytes. We conclude that cycles of intra-Mili secondary piRNA biogenesis fuel piRNA amplification that is absolutely required for LINE1 silencing.

AB - Piwi proteins and Piwi-interacting RNAs (piRNAs) have conserved functions in transposon silencing. The murine Piwi proteins Mili and Miwi2 (also called Piwil2 and Piwil4, respectively) direct epigenetic LINE1 and intracisternal A particle transposon silencing during genome reprogramming in the embryonic male germ line. Piwi proteins are proposed to be piRNA-guided endonucleases that initiate secondary piRNA biogenesis; however, the actual contribution of their endonuclease activities to piRNA biogenesis and transposon silencing remain unknown. To investigate the role of Piwi-catalysed endonucleolytic activity, we engineered point mutations in mice that substitute the second aspartic acid to an alanine in the DDH catalytic triad of Mili and Miwi2, generating the Mili DAH and Miwi2 DAH alleles, respectively. Analysis of Mili-bound piRNAs from homozygous Mili DAH fetal gonadocytes revealed a failure of transposon piRNA amplification, resulting in the marked reduction of piRNA bound within Miwi2 ribonuclear particles. We find that Mili-mediated piRNA amplification is selectively required for LINE1, but not intracisternal A particle, silencing. The defective piRNA pathway in Mili DAH mice results in spermatogenic failure and sterility. Surprisingly, homozygous Miwi2 DAH mice are fertile, transposon silencing is established normally and no defects in secondary piRNA biogenesis are observed. In addition, the hallmarks of piRNA amplification are observed in Miwi2-deficient gonadocytes. We conclude that cycles of intra-Mili secondary piRNA biogenesis fuel piRNA amplification that is absolutely required for LINE1 silencing.

UR - http://www.scopus.com/inward/record.url?scp=83055180736&partnerID=8YFLogxK

U2 - 10.1038/nature10547

DO - 10.1038/nature10547

M3 - Journal article

C2 - 22020280

AN - SCOPUS:83055180736

SN - 0028-0836

VL - 480

SP - 259

EP - 263

JO - Nature

JF - Nature

IS - 7376

ER -