title: The Caenorhabditis elegans HEN1 ortholog, HENN-1, methylates and stabilizes select subclasses of germline small RNAs.
abstract: Small RNAs regulate diverse biological processes by directing effector proteins called Argonautes to silence complementary mRNAs. Maturation of some classes of small RNAs involves terminal 2'-O-methylation to prevent degradation. This modification is catalyzed by members of the conserved HEN1 RNA methyltransferase family. In animals, Piwi-interacting RNAs (piRNAs) and some endogenous and exogenous small interfering RNAs (siRNAs) are methylated, whereas microRNAs are not. However, the mechanisms that determine animal HEN1 substrate specificity have yet to be fully resolved. In Caenorhabditis elegans, a HEN1 ortholog has not been studied, but there is evidence for methylation of piRNAs and some endogenous siRNAs. Here, we report that the worm HEN1 ortholog, HENN-1 (HEN of Nematode), is required for methylation of C. elegans small RNAs. Our results indicate that piRNAs are universally methylated by HENN-1. In contrast, 26G RNAs, a class of primary endogenous siRNAs, are methylated in female germline and embryo, but not in male germline. Intriguingly, the methylation pattern of 26G RNAs correlates with the expression of distinct male and female germline Argonautes. Moreover, loss of the female germline Argonaute results in loss of 26G RNA methylation altogether. These findings support a model wherein methylation status of a metazoan small RNA is dictated by the Argonaute to which it binds. Loss of henn-1 results in phenotypes that reflect destabilization of substrate small RNAs: dysregulation of target mRNAs, impaired fertility, and enhanced somatic RNAi. Additionally, the henn-1 mutant shows a weakened response to RNAi knockdown of germline genes, suggesting that HENN-1 may also function in canonical RNAi. Together, our results indicate a broad role for HENN-1 in both endogenous and exogenous gene silencing pathways and provide further insight into the mechanisms of HEN1 substrate discrimination and the diversity within the Argonaute family.
authors.affiliation: Life Sciences Institute, University of Michigan, Ann Arbor, Michigan, United States of America.
authors.forename: Allison C
authors.initials: AC
authors.lastname: Billi
authors.forename: Amelia F
authors.initials: AF
authors.lastname: Alessi
authors.forename: Vishal
authors.initials: V
authors.lastname: Khivansara
authors.forename: Ting
authors.initials: T
authors.lastname: Han
authors.forename: Mallory
authors.initials: M
authors.lastname: Freeberg
authors.forename: Shohei
authors.initials: S
authors.lastname: Mitani
authors.forename: John K
authors.initials: JK
authors.lastname: Kim
mesh_terms: D000818:Animals; D060565:Argonaute Proteins / Q000235:genetics / Q000378:metabolism; D017173:Caenorhabditis elegans / Q000235:genetics* / Q000378:metabolism; D029742:Caenorhabditis elegans Proteins / Q000235:genetics / Q000378:metabolism; D005260:Female; D005786:Gene Expression Regulation; D020868:Gene Silencing; D005854:Germ Cells / Q000254:growth & development / Q000378:metabolism; D008297:Male; D008745:Methylation; D008780:Methyltransferases / Q000235:genetics / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D012342:RNA, Small Nuclear / Q000235:genetics* / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D060565:Argonaute Proteins; D029742:Caenorhabditis elegans Proteins; C548334:ERGO-1 protein, C elegans; D035683:MicroRNAs; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D012342:RNA, Small Nuclear; D016601:RNA-Binding Proteins; C548335:T22B3.2 protein, C elegans; C575114:HENN-1 protein, C elegans; D008780:Methyltransferases