A Baculovirus Superinfection System: Efficient Vehicle for Gene Transfer into
            <i>Drosophila</i>
            S2 Cells

Lee, Dung‐Fang; Chen, Chun-Chen; Hsu, Tsu‐An; Juang, Jyh‐Lyh

doi:10.1128/jvi.74.24.11873-11880.2000

Cited by 61 publications

(36 citation statements)

References 40 publications

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“…Flagtagged D-elp1protein was expressed in Sf9 cells using a modified baculovirus vector containing the heat shock inducible HSP70 promoter (17). High level recombinant D-elp1 (rD-elp1) expression from the polh promoter in the standard baculovirus vector was toxic to Sf9 cells.…”

Section: Resultsmentioning

confidence: 99%

RETRACTED: Identification of an RNA-dependent RNA polymerase in Drosophila involved in RNAi and transposon suppression

Lipardi

Paterson

2009

Proc. Natl. Acad. Sci. U.S.A.

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The authors wish to note the following: "In our article, we reported that recombinant subunit 1 of the Drosophila elongator complex, D-elp1, had RNA-dependent RNA polymerase (RdRP) activity and was involved in RNAi, transposon suppression, and endo-siRNA production, but not miRNA targeting. RdRP activity was identified using three assays to interrogate the reaction products: Dcr2 digestion, RNase sensitivity, and nearest neighbor analysis. Although we do see differential Dcr2 cleavage and RNase resistance, the gold standard for second strand RNA synthesis is nearest neighbor analysis, as described previously for the Neurospora crassa RdRP, QDE-1 (1). Subsequent studies revealed that the nearest neighbor analysis was misinterpreted because of two factors unknown to us at the time: First, T7 run-off transcripts used as single-stranded RNA templates have heterogeneous 3' termini (2, 3); Second, the Flag-tag affinity purified recombinant D-elp1 protein preparations used in our study contain a ribonucleotide terminal transferase activity able to catalyze the addition of single a-labeled ribonucleotide triphosphates to the 3' hydroxyl terminus of the template RNA. Therefore, nearest neighbor analysis measured the heterogeneous 3' termini of the template RNA and not second strand RNA synthesis, as initially reported. Given this result, we wish to rescind the interpretation that D-elp1 can be considered as an RNA-dependent RNA polymerase. The basis for the ribonucleotide terminal transferase activity is under investigation. The interpretation of the results does not detract from the fact that D-elp1 is involved in RNAi, endo-siRNA production, and transposon suppression, but not miRNA targeting as shown in Figs. 3 and 4. By clarifying this issue, we hope to promote further productive investigation into the role of elongator subunit 1 in RNAi and transposon suppression. We apologize for any difficulties our original interpretation may have caused other investigators and hereby retract the paper." Concetta Lipardi Bruce M. Paterson

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Section: Resultsmentioning

confidence: 99%

RETRACTED: Identification of an RNA-dependent RNA polymerase in Drosophila involved in RNAi and transposon suppression

Lipardi

Paterson

2009

Proc. Natl. Acad. Sci. U.S.A.

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“…The first is a widely used method of stable transfection in cells, expressing the protein under the CuSO 4 -inducible Metallothionein (Mtn) promoter (20). The second method uses a baculovirus superinfection system (24). Baculovirus, which infects and replicates in lepidopteran cells, can also infect Drosophila S2 cells but is unable to replicate (24).…”

Section: Resultsmentioning

confidence: 99%

The COMPASS Family of H3K4 Methylases in Drosophila

Mohan

Herz

Smith

et al. 2011

Molecular and Cellular Biology

205

222

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Methylation of histone H3 lysine 4 (H3K4) in Saccharomyces cerevisiae is implemented by Set1/COMPASS, which was originally purified based on the similarity of yeast Set1 to human MLL1 and Drosophila melanogaster Trithorax (Trx). While humans have six COMPASS family members, Drosophila possesses a representative of the three subclasses within COMPASS-like complexes: dSet1 (human SET1A/SET1B), Trx (human MLL1/2), and Trr (human MLL3/4). Here, we report the biochemical purification and molecular characterization of the Drosophila COMPASS family. We observed a one-to-one similarity in subunit composition with their mammalian counterparts, with the exception of LPT (lost plant homeodomains [PHDs] of Trr), which copurifies with the Trr complex. LPT is a previously uncharacterized protein that is homologous to the multiple PHD fingers found in the N-terminal regions of mammalian MLL3/4 but not Drosophila Trr, indicating that Trr and LPT constitute a split gene of an MLL3/4 ancestor. Our study demonstrates that all three complexes in Drosophila are H3K4 methyltransferases; however, dSet1/COMPASS is the major monoubiquitination-dependent H3K4 di-and trimethylase in Drosophila. Taken together, this study provides a springboard for the functional dissection of the COMPASS family members and their role in the regulation of histone H3K4 methylation throughout development in Drosophila.Histone H3 lysine 4 methylation (H3K4me) is associated with the transcriptionally active regions of the genome in yeast, flies, and mammals (3,23,35). Set1 was identified as a component of a macromolecular protein complex named COMPASS (complex of proteins associated with Set 1), as the first H3K4 methylase, and it is responsible for all mono-, di-, and trimethylation of H3K4 in yeast (22,31,40,52). In Drosophila melanogaster, four SET domain-containing proteins, namely, Trithorax (Trx), Trithorax-related (Trr), dSet1, and Ash1, have been reported to implement H3K4 methylation (10). All but Ash1, which has subsequently been demonstrated to be an H3K36 methyltransferase (49,59), are related to subunits of the six COMPASS and COMPASS-like complexes in mammals. trx was originally characterized as a gene that when mutated caused homeotic transformations (6, 18). Detailed genetic and molecular analyses showed that Trx is required to maintain activation states of its target genes throughout development and counteracts the repressive effects of the Polycomb group proteins (PcG) (39, 41). Trr was identified based on sequence similarity to Trx but was shown to function in the regulation of hormone-responsive gene expression (42). dSet1 was identified based on sequence homology to the Saccharomyces cerevisiae and mammalian Set1 proteins (53, 58).In mammals, there are at least six SET1-related proteins that form COMPASS-like complexes, namely, SET1A, SET1B, and MLL1 to MLL4. SET1A and SET1B are orthologous to dSet1; MLL1 and MLL2 are orthologous to Drosophila Trx; MLL3 and MLL4 (also known as ALR) are orthologous to Drosophila Trr (33,43,45). All of the m...

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“…However, cell lysis-associated proteolysis (if the protein of interest is a secreted protein, proteinases from lysed cells could severely compromise the yield of protein production the restricted host range [97]. Furthermore, the restricted host range (invertebrates) obviously makes baculovirus expression inconvenient for some applications.…”

Section: Non-integrating Viral Vectorsmentioning

confidence: 99%

RNA Interference with Special Reference to Combating Viruses of Crustacea

Fauce

Owens

2012

Indian J. Virol.

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RNA interference has evolved from being a nuisance biological phenomenon to a valuable research tool to determine gene function and as a therapeutic agent. Since pioneering observations regarding RNA interference were first reported in the 1990s from the nematode worm, plants and Drosophila, the RNAi phenomenon has since been reported in all eukaryotic organisms investigated from protozoans, plants, arthropods, fish and mammals. The design of RNAi therapeutics has progressed rapidly to designing dsRNA that can specifically and effectively silence disease related genes. Such technology has demonstrated the effective use of short interfering as therapeutics. In the absence of a B cell lineage in arthropods, and hence no long term vaccination strategy being available, the introduction of using RNA interference in crustacea may serve as an effective control and preventative measure for viral diseases for application in aquaculture.

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A Baculovirus Superinfection System: Efficient Vehicle for Gene Transfer into Drosophila S2 Cells

Cited by 61 publications

References 40 publications

RETRACTED: Identification of an RNA-dependent RNA polymerase in Drosophila involved in RNAi and transposon suppression

RETRACTED: Identification of an RNA-dependent RNA polymerase in Drosophila involved in RNAi and transposon suppression

The COMPASS Family of H3K4 Methylases in Drosophila

RNA Interference with Special Reference to Combating Viruses of Crustacea

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