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G3 (Bethesda). 2019 May 15. pii: g3.400330.2019. doi: 10.1534/g3.119.400330. [Epub ahead of print]

A Non-Dicer RNase III and Four Other Novel Factors Required for RNAi-Mediated Transposon Suppression in the Human Pathogenic Yeast Cryptococcus neoformans.

Author information

1
University of California, San Francisco.
2
Salk Institute for Biological Studies.
3
The Scripps Research Institute.
4
University of California, San Francisco; Chan-Zuckerberg Biohub hitenmadhani@gmail.com.

Abstract

The human pathogenic yeast Cryptococcus neoformans silences transposable elements using endo-siRNAs and an Argonaute, Ago1. 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. cerevisiae Rnt1, 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. cerevisiae Sqs1/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.

KEYWORDS:

RNA surveillance; RNAi; transposon

PMID:
31092606
DOI:
10.1534/g3.119.400330
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