Protease-mediated processing of Argonaute proteins controls small RNA association

•DPF-3, a DPPIV family protease, protects C. elegans fertility•Loss of DPF-3 alters cellular endo-siRNA profiles and desilences transposons•DPF-3-mediated processing WAGO-1/-3 Argonaute proteins promotes siRNA sorting•A combination processing-refractive WAGO-1 WAGO-3 phenocopies loss Small RNA pathways defend the germlines animals against selfish genetic elements, yet pathway activities need to be contained prevent silencing self genes. Here, we reveal proteolytic mechanism that controls endogenous small interfering (22G) activity in Caenorhabditis germline protect genome integrity maintain fertility. We find DPF-3, P-granule-localized N-terminal dipeptidase orthologous mammalian dipeptidyl peptidase (DPP) 8/9, processes unusually proline-rich N termini (Ago) proteins. Without activity, these WAGO lose their proper complement 22G RNAs. Desilencing repeat-containing transposon-derived transcripts, DNA damage, acute sterility ensue. These phenotypes are recapitulated when rendered resistant DPF-3-mediated processing, identifying them as critical substrates DPF-3. conclude Ago regulates elements by ensuring association with appropriate Members IV (DPPIV) serine protease have important roles animal pathophysiology. DPP4 functions regulator glucose homeostasis, its pharmaceutical inhibition is clinically exploited treat type 2 diabetes (Deacon, 2019Deacon C.F. Physiology Pharmacology DPP-4 Glucose Homeostasis Treatment Type Diabetes.Front. Endocrinol. (Lausanne). 2019; 10: 80Crossref PubMed Google Scholar), whereas paralogous 8 DPP9 play role immune system function malignancies, inflammation, beyond (Zhang et al., 2013Zhang H. Chen Y. Keane F.M. Gorrell M.D. Advances understanding expression 9.Mol. Cancer Res. 2013; 11: 1487-1496Crossref Scopus (48) Scholar). (DPF) proteases cleave off two amino-acid-long peptides sequence Xaa-Pro or Xaa-Ala from terminus (Keane 2011Keane Nadvi N.A. Yao T.W. 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Although most mutant appeared overtly wild under standard culture conditions, noticed unfertilized oocytes plates dpf-3(xe68) null (hereafter called dpf-3?). Moreover, at 25°C, had reduced brood size relative wild-type 1A). DPP8/9. understand whether reproduction, used editing change 784, part presumed (Ajami 2003Ajami K. Abbott C.A. Obradovic Gysbers V. Kähne McCaughan G.W. Structural requirements catalysis, expression, dimerization CD26/DPIV family.Biochemistry. 42: 694-701Crossref (58) alanine, generating dpf-3(xe71[S784A]) animals, henceforth dpf-3(S784A). strain, additionally FLAG-hemagglutinin (HA) epitope tagged. dpf-3(S784A) accumulated levels S1D), exhibited notable decrease 1B). For both point strains, observed temperature dependence phenotype. decreased increasing complete occurred 26.5°C 1B), was maintained functional copy expressed dpf-3? (dpf-3 rescue, Figures S1E S1F). Biochemical experiments shown later validate an extensive animals. mutations residues caused fully penetrant dpf-3(D861A) dpf-3(H893A) S1E). particular identify cause sterility, examined differential interference contrast (DIC) microscopy. When grown 20°C, adult either genotype fertilized eggs multicellular embryos uterus. At 26.5°C, still true not Instead embryos, carried uterus 1C). assess apparent fertilization defect consequence defects, oocyte both, performed crosses between hermaphrodites males. mated males 20°C yielded viable progeny S1G). could sire, albeit inefficiently, S1G), (n > 100). elevated temperature. Using his-70::gfp reporter visualize progenitor cells mature (Delaney 2018Delaney Mailler Wenda Gabus Steiner F.A. Differential Expression Histone H3.3 Their Role Modulating Temperature Stress Response elegans.Genetics. 209: 551-565Crossref (23) found types were readily detectable fourth larval (L4) stage, spermatogenesis occurs, irrespective growth By contrast, visible undetectable 1D). Thus, supports production temperatures. Wild-type sired fewer crossed Combined evidence later, indicates female Because serial hermaphrodite produces only limited number last stage before switching production, continues throughout adulthood (L’Hernault, 2009L’Hernault S.W. genetics cell biology nematode 306: 59-65Crossref dominate. data thus far support germline. inserted encoding 3xFLAG::TEV::GFP::HiBit tag just stop codon gene. dpf-3(xe246) fertile confirming functionality tagged Imaging revealed various tissues S2A–S2D), 2A) spermatozoa 2B). although distributed diffusely cytoplasm cells, perinuclear punctae pachytene-stage germ proximal gonad, maturing 2B; Figure S2E). 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