The 26S proteasome in Schistosoma mansoni: Bioinformatics analysis, developmental expression, and RNA interference (RNAi) studies

Abstract Comprehensive understanding of pleiotropic roles of RNAi machinery highlighted the conserved chromosomal functions of RNA interference. The consequences of the evolutionary variation in the core RNAi pathway genes are mostly unknown, but may lead to the species-specific functions associated with gene silencing. The two-spotted spider mite, Tetranychus urticae, is a major polyphagous chelicerate pest capable of feeding on over 1100 plant species and developing resistance to pesticides used for its control. A well annotated genome, susceptibility to RNAi and economic importance, make T. urticae an excellent candidate for development of an RNAi protocol that enables high-throughput genetic screens and RNAi-based pest control. Here, we show that the length of the exogenous dsRNA critically determines its processivity and ability to induce RNAi in vivo. A combination of the long dsRNAs and the use of dye to trace the ingestion of dsRNA enabled the identification of genes involved in membrane transport and 26S proteasome degradation as sensitive RNAi targets. Our data demonstrate that environmental RNAi can be an efficient reverse genetics and pest control tool in T. urticae. In addition, the species-specific properties together with the variation in the components of the RNAi machinery make T. urticae a potent experimental system to study the evolution of RNAi pathways.

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Environmental RNA interference in two-spotted spider mite, Tetranychus urticae, reveals dsRNA processing requirements for efficient RNAi response

10.1101/2020.05.10.087429 ◽

2020 ◽

Cited By ~ 1

Author(s):

Nicolas Bensoussan ◽

Sameer Dixit ◽

Midori Tabara ◽

David Letwin ◽

Maja Milojevic ◽

...

Keyword(s):

Rna Interference ◽

Tetranychus Urticae ◽

Pest Control ◽

Spider Mite ◽

26S Proteasome ◽

Rnai Pathway ◽

Rnai Machinery ◽

Two Spotted Spider Mite ◽

Species Specific

AbstractComprehensive understanding of pleiotropic roles of RNAi machinery highlighted the conserved chromosomal functions of RNA interference. The consequences of the evolutionary variation in the core RNAi pathway genes are mostly unknown, but may lead to the species-specific functions associated with gene silencing. The two-spotted spider mite, Tetranychus urticae, is a major polyphagous chelicerate pest capable of feeding on over 1,100 plant species and developing resistance to pesticides used for its control. A well annotated genome, susceptibility to RNAi and economic importance, make T. urticae an excellent candidate for development of an RNAi protocol that enables high-throughput genetic screens and RNAi-based pest control. Here, we show that the length of the exogenous dsRNA critically determines its processivity and ability to induce RNAi in vivo. A combination of the long dsRNAs and the use of dye to trace the ingestion of dsRNA enabled the identification of genes involved in membrane transport and 26S proteasome degradation as sensitive RNAi targets. Our data demonstrate that environmental RNAi can be an efficient reverse genetics and pest control tool in T. urticae. In addition, the species-specific properties together with the variation in the components of the RNAi machinery make T. urticae a potent experimental system to study the evolution of RNAi pathways.

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Molecular characterization and developmental expression of Ras-related nuclear protein in the oriental river prawn Macrobrachium nipponense and the effects of RNA interference on ovarian maturation

Journal of Fishery Sciences of China ◽

10.3724/sp.j.1118.2017.16365 ◽

2017 ◽

Vol 24 (3) ◽

pp. 459

Author(s):

Zongyuan BU ◽

Hongtuo FU ◽

Shengming SUN ◽

Hui QIAO ◽

Shubo JIN ◽

...

Keyword(s):

Rna Interference ◽

Molecular Characterization ◽

Nuclear Protein ◽

Developmental Expression ◽

Ovarian Maturation ◽

Macrobrachium Nipponense ◽

Oriental River Prawn

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Reverse Genetic Analysis of the Caenorhabditis elegans 26S Proteasome Subunits by RNA Interference

Biological Chemistry ◽

10.1515/bc.2002.140 ◽

2002 ◽

Vol 383 (7-8) ◽

pp. 1263-1266 ◽

Cited By ~ 23

Author(s):

M. Takahashi ◽

H. Iwasaki ◽

H. Inoue ◽

K. Takahashi

Keyword(s):

Rna Interference ◽

Caenorhabditis Elegans ◽

Genetic Analysis ◽

Postembryonic Development ◽

26S Proteasome ◽

Reverse Genetic ◽

Proteasome Subunit ◽

Proteasome Subunits ◽

Regulatory Particle

Abstract Reverse genetic analysis was performed on the Caenorhabditis elegans 26S proteasome subunit genes by doublestranded RNAmediated interference (RNAi). Embryonic and postembryonic lethality was caused by interference of all of the eight tested 20S core subunits and all of the 19S regulatory particle subunits except for CeRpn9, CeRpn10, and Ce Rpn12, where RNAi caused no abnormality. However, synthetic suppression of CeRpn10 and CeRpn12 was lethal, whereas neither the combination of Ce Rpn9 with CeRpn10 nor with CeRpn12 resulted in abnormalities in RNAi. These results indicate that the 26S proteasome is indispensable for embryogenesis and postembryonic development, although Ce Rpn9, CeRpn10, and CeRpn12 are not essential, at least under the conditions used. CeRpn10 and Ce Rpn12 are considered to compensate for the suppression of each other.

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Use of RNA Interference and Complementation To Study the Function of the Drosophila and Human 26S Proteasome Subunit S13

Molecular and Cellular Biology ◽

10.1128/mcb.23.15.5320-5330.2003 ◽

2003 ◽

Vol 23 (15) ◽

pp. 5320-5330 ◽

Cited By ~ 49

Author(s):

Josefin Lundgren ◽

Patrick Masson ◽

Claudio A. Realini ◽

Patrick Young

Keyword(s):

Cell Cycle ◽

Rna Interference ◽

Cell Cycle Progression ◽

Degradation Pathway ◽

26S Proteasome ◽

Wild Type ◽

Proteasome Subunit ◽

Rnai Treatment ◽

Regulatory Complex

ABSTRACT The S13 subunit (also called Pad1, Rpn11, and MPR1) is a component of the 19S complex, a regulatory complex essential for the ubiquitin-dependent proteolytic activity of the 26S proteasome. To address the functional role of S13, we combined double-stranded RNA interference (RNAi) against the Drosophila proteasome subunit DmS13 with expression of wild-type and mutant forms of the homologous human gene, HS13. These studies show that DmS13 is essential for 26S function. Loss of the S13 subunit in metazoan cells leads to increased levels of ubiquitin conjugates, cell cycle defects, DNA overreplication, and apoptosis. In vivo assays using short-lived proteasome substrates confirmed that the 26S ubiquitin-dependent degradation pathway is compromised in S13-depleted cells. In complementation experiments using Drosophila cell lines expressing HS13, wild-type HS13 was found to fully rescue the knockdown phenotype after DmS13 RNAi treatment, while an HS13 containing mutations (H113A-H115A) in the proposed isopeptidase active site was unable to rescue. A mutation within the conserved MPN/JAMM domain (C120A) abolished the ability of HS13 to rescue the Drosophila cells from apoptosis or DNA overreplication. However, the C120A mutant was found to partially restore normal levels of ubiquitin conjugates. The S13 subunit may possess multiple functions, including a deubiquitinylating activity and distinct activities essential for cell cycle progression that require the conserved C120 residue.

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