Characterization of a TUTase/RNase complex required for Drosophila gametogenesis

• Published in: RNA (Cambridge) Vol. 23; no. 3; pp. 284 - 296
• Main Authors: Lin, Ching-Jung, Wen, Jiayu, Bejarano, Fernando, Hu, Fuqu, Bortolamiol-Becet, Diane, Kan, Lijuan, Sanfilippo, Piero, Kondo, Shu, Lai, Eric C
• Format: Journal Article
• Language: English
• Published: United States Cold Spring Harbor Laboratory Press 01-03-2017
• Subjects: Animals; Drosophila; Drosophila melanogaster - genetics; Drosophila melanogaster - growth & development; Drosophila melanogaster - metabolism; Drosophila Proteins - genetics; Drosophila Proteins - metabolism; Exoribonucleases - genetics; Exoribonucleases - metabolism; Female; Infertility, Female - genetics; Infertility, Male - genetics; Male; Mutation; RNA Nucleotidyltransferases - genetics; RNA Nucleotidyltransferases - metabolism; RNA Processing, Post-Transcriptional; RNA, Untranslated - genetics; RNA, Untranslated - metabolism; Spermatogenesis - genetics; spermatogenesis; exoribonuclease; TUTase; Drosophila
• ISSN: 1355-8382; 1469-9001
• DOI: 10.1261/rna.059527.116

Post-transcriptional regulatory strategies that involve coupling between terminal uridyltransferase (TUTase) and exoribonuclease enzymes have recently been characterized in diverse species. Of note, the 3' exoribonuclease Dis3L2 has received substantial attention as a factor that metabolizes uridylated substrates in contexts such as general mRNA degradation, turnover of specific miRNAs, and quality control of noncoding RNAs. To date, most studies of Dis3L2 have focused on fungi and mammalian cells. Here, we introduce
as a system that permits analysis of molecular mechanisms as well as the ability to interrogate organismal phenotypes. We started with a structure-function analysis of the
TUTase Tailor, which we recently identified to inhibit biogenesis of splicing-derived miRNA hairpins. Next, we show that Tailor/Dis3L2 form a complex via N-terminal domains in the respective proteins that are distinct from their catalytic domains. In vitro, Dis3L2 has nuclease activity, but substrate oligouridylation by Tailor stimulates their degradation by Dis3L2, especially for structured substrates. We analyzed mutants of
and
, which are viable and lack overt morphological defects. Instead, these mutants exhibit defects in female and male fertility, implying specific requirements in the germline.
defects are more severe than
, and their requirements appear stronger in males than in females. In particular, loss of Dis3L2 completely blocks productive spermatogenesis, causing male sterility. RNA-seq analysis from single- and double-mutant testes reveals aberrant gene expression programs and suggests that noncoding RNAs may be preferentially affected by Dis3L2. Overall, our studies of a new tailing/trimming complex reveal unexpectedly specific requirements during gametogenesis.