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Endo-siRNAs production requires the RNA-dependent RNA polymerase, Rdp1, and two partially redundant Dicer enzymes, Dcr1 and Dcr2, but is independent of histone H3 lysine 9 methylation. We describe here an insertional mutagenesis screen for factors required to suppress the mobilization of the C. neoformans HARBINGER family DNA transposon HAR1. Validation experiments uncovered five novel genes (RDE1-5) required for HAR1 suppression and global production of suppressive endo-siRNAs. The RDE genes do not impact transcript levels, suggesting the endo-siRNAs do not act by impacting target transcript synthesis or turnover. RDE3 encodes a non-Dicer RNase III related to S. cerevisiaeRnt1, RDE4 encodes a predicted terminal nucleotidyltransferase, while RDE5 has no strongly predicted encoded domains. Affinity purification-mass spectrometry studies suggest that Rde3 and Rde5 are physically associated. RDE1 encodes a G-patch protein homologous to the S. cerevisiaeSqs1\/Pfa1, a nucleolar protein that directly activates the essential helicase Prp43 during rRNA biogenesis. Rde1 copurifies Rde2, another novel protein obtained in the screen, as well as Ago1, a homolog of Prp43, and numerous predicted nucleolar proteins. We also describe the isolation of conditional alleles of PRP43, which are defective in RNAi. This work reveals unanticipated requirements for a non-Dicer RNase III and presumptive nucleolar factors for endo-siRNA biogenesis and transposon mobilization suppression in C. neoformans.<\/jats:p>","DOI":"10.1534\/g3.119.400330","type":"journal-article","created":{"date-parts":[[2019,5,16]],"date-time":"2019-05-16T00:05:31Z","timestamp":1557965131000},"page":"2235-2244","source":"Crossref","is-referenced-by-count":27,"title":["A Non-Dicer RNase III and Four Other Novel Factors Required for RNAi-Mediated Transposon Suppression in the Human Pathogenic Yeast\n                    <i>Cryptococcus neoformans<\/i>"],"prefix":"10.1093","volume":"9","author":[{"given":"Jordan E","family":"Burke","sequence":"first","affiliation":[{"name":"Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94158"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Adam D","family":"Longhurst","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94158"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Prashanthi","family":"Natarajan","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94158"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Beiduo","family":"Rao","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94158"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"John","family":"Liu","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94158"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jade","family":"Sales-Lee","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94158"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yasaman","family":"Mortensen","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94158"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"James J","family":"Moresco","sequence":"additional","affiliation":[{"name":"Salk Institute for Biological Studies, La Jolla, CA 92037"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jolene K","family":"Diedrich","sequence":"additional","affiliation":[{"name":"Salk Institute for Biological Studies, La Jolla, CA 92037"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"suffix":"III","given":"John R","family":"Yates","sequence":"additional","affiliation":[{"name":"Department of Molecular Medicine, the Scripps Research Institute, La Jolla, CA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hiten D","family":"Madhani","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94158, Chan-Zuckerberg Biohub, San Francisco, CA 94158"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"286","published-online":{"date-parts":[[2019,7,1]]},"reference":[{"key":"2021042014040144100_bib1","doi-asserted-by":"publisher","first-page":"3389","DOI":"10.1093\/nar\/25.17.3389","article-title":"Gapped BLAST and PSI-BLAST: a new generation of protein database search programs.","volume":"25","author":"Altschul","year":"1997","journal-title":"Nucleic Acids Res."},{"key":"2021042014040144100_bib2","doi-asserted-by":"publisher","first-page":"166","DOI":"10.1093\/bioinformatics\/btu638","article-title":"HTSeq\u2013a Python framework to work with high-throughput sequencing data.","volume":"31","author":"Anders","year":"2015","journal-title":"Bioinformatics"},{"key":"2021042014040144100_bib3","doi-asserted-by":"publisher","first-page":"561","DOI":"10.1007\/s10577-013-9388-2","article-title":"RNAi function, diversity, and loss in the fungal kingdom.","volume":"21","author":"Billmyre","year":"2013","journal-title":"Chromosome Res."},{"key":"2021042014040144100_bib4","doi-asserted-by":"publisher","first-page":"707","DOI":"10.1016\/j.cell.2007.03.038","article-title":"RNAi-Dependent and -Independent RNA Turnover Mechanisms Contribute to Heterochromatic Gene Silencing.","volume":"129","author":"B\u00fchler","year":"2007","journal-title":"Cell"},{"key":"2021042014040144100_bib5"},{"key":"2021042014040144100_bib6","doi-asserted-by":"publisher","first-page":"15548","DOI":"10.1073\/pnas.1522127112","article-title":"Conserved factor Dhp1\/Rat1\/Xrn2 triggers premature transcription termination and nucleates heterochromatin to promote gene silencing.","volume":"112","author":"Chalamcharla","year":"2015","journal-title":"Proc. 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