A complementary transposon tool kit for Drosophila melanogaster using P and piggyBac

Thibault, Stephen T.; Singer, Matthew A.; Miyazaki, W Y; Milash, Brett; Dompe, Nicholas; Singh, Carol M.; Buchholz, Ross; Demsky, Madelyn; Fawcett, Robert; Francis-Lang, Helen; Ryner, Lisa; Cheung, Lai Man; Chong, Angela; Erickson, Cathy; Fisher, William W.; Greer, Kimberly J.; Hartouni, Stephanie R; Howie, Elizabeth; Jakkula, Lakshmi R.; Joo, Daniel M.; Killpack, Keith; Laufer, Alex; Mazzotta, Julie; Smith, Ronald D.; Stevens, Lynn M.; Stuber, Christiana; Tan, Lory R; Ventura, Richard; Woo, Alesa; Zakrajsek, Irena; Zhao, Lora; Chen, Feng; Swimmer, Candace; Kopczynski, Casey; Duyk, Geoffrey M.; Winberg, Margaret L.; Margolis, Jonathan

doi:10.1038/ng1314

Cited by 768 publications

(832 citation statements)

References 24 publications

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“…Gie/Arl8, the single Drosophila ortholog for mammalian ARL8A/B, also localizes to lysosomes [71], but a phenotypic description of Gie/Arl8 loss-of-function in flies is lacking. We found that Gie e00336 , caused by the insertion of a ~ 6 kb transposable element into the first intron of the Drosophila Gie / Arl8 gene [72], is pupal lethal in combination with the overlapping deficiency Df(3R)BSC222 , suggesting that it represents a strong loss-of-function allele. We therefore examined post-endocytic trafficking in garland cells from these hemizygotes, denoted Gie e00336 /Df .…”

Section: Resultsmentioning

confidence: 99%

Drosophila Rab2 controls endosome-lysosome fusion and LAMP delivery to late endosomes

2018

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Rab2 is a conserved Rab GTPase with a well-established role in secretory pathway function and phagocytosis. Here we demonstrate that Drosophila Rab2 is recruited to late endosomal membranes, where it controls the fusion of LAMP-containing biosynthetic carriers and lysosomes to late endosomes. In contrast, the lysosomal GTPase Gie/Arl8 is only required for late endosome-lysosome fusion, but not for the delivery of LAMP to the endocytic pathway. We also find that Rab2 is required for the fusion of autophagosomes to the endolysosomal pathway, but not for the biogenesis of lysosome-related organelles. Surprisingly, Rab2 does not rely on HOPS-mediated vesicular fusion for recruitment to late endosomal membranes. Our work suggests that Drosophila Rab2 is a central regulator of the endolysosomal and macroautophagic/autophagic pathways by controlling the major heterotypic fusion processes at the late endosome.

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Section: Resultsmentioning

confidence: 99%

Drosophila Rab2 controls endosome-lysosome fusion and LAMP delivery to late endosomes

2018

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“…Finally, the presence of a transposon at the target site may not be strictly required for gene conversion: an apparent instance of transposon-independent gene conversion has been obtained for the gene rol-6, albeit at an extremely low frequency 30 . An expansion of the available transposon insertion libraries 11-13 could conceivably tag every gene in C. elegans with a transposon and function as a basis for the detailed in vivo analysis of all genes by mutagenesis, as is currently underway in D. melanogaster 14,15 . In addition, intergenic regions that are not accessible by classical genetic analysis could be readily targeted.…”

Section: Discussionmentioning

confidence: 99%

“…An increase in these efforts could establish a C. elegans genome-wide transposon library. Such a project has been reported in Drosophila melanogaster, for which 450% of the genome has already been covered, and improved transposon-based techniques have been developed to address problems such as the nonrandom insertion bias of the transposons 14,15 .…”

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confidence: 99%

Targeted gene alteration in Caenorhabditis elegans by gene conversion

Barrett¹,

Fleming²,

Göbel³

2004

Nat Genet

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“…Likewise, genetic modifications of Minos-based transgenic lines require another transformation system. Transposons usually have their own bias of insertion sites, and using two or more transposons facilitates saturated mutagenesis (Thibault et al, 2004;Liang et al, 2009). Furthermore, transposons which have different characteristics from Minos will bring us another transgenic technology based on these characteristics, which could not be accomplished with Minos (Huet et al, 2002;Kokubu et al, 2009).…”

Section: Introductionmentioning

confidence: 99%

Germline transgenesis of the chordate Ciona intestinalis with hyperactive variants of sleeping beauty transposable element

Hozumi

Mita

Miskey

et al. 2012

Developmental Dynamics

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Background: Transposon-mediated transgenesis is an excellent method for creating stable transgenic lines and insertional mutants. In the chordate Ciona intestinalis, Minos is the only transposon that has been used as the tool for germline transformation. Adding another transposon system in this organism enables us to conduct genetic techniques which can only be realized with the use of two transposons. Results: In the present study, we found that another Tc1/mariner superfamily transposon, sleeping beauty (SB), retains sufficient activity for germline transformation of C. intestinalis. SB shows efficiencies of germline transformation, insertion into gene coding regions, and enhancer detection comparable to those of Minos. We have developed a system for the remobilization of SB copies in the C. intestinalis genome by using transgenic lines expressing SB transposase in the germ cells. With this system, we examined the manner of SB mobilization in the C. intestinalis genome. SB shows intrachromosomal transposition more frequently than Minos. Conclusions: SB-based germline transformation and the establishment of a new method that uses its frequent intrachromosomal transposition will result in breakthroughs in genetic approaches that use C. intestinalis together with Minos. Developmental Dynamics 242:30-43, 2013. V C 2012 Wiley Periodicals, Inc.Key words: ascidian; Ciona intestinalis; transposon; Tc1; mariner; sleeping beauty; Minos Key findings:In C. intestinalis, Minos is the only transposon that has been used as a tool for germline transformation. We found that another transposon, sleeping beauty (SB), retains sufficient activity for the germline transformation of C. intestinalis. SB-based germline transformation will result in breakthroughs in genetic approaches that use C. intestinalis together with Minos.

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A complementary transposon tool kit for Drosophila melanogaster using P and piggyBac

Cited by 768 publications

References 24 publications

Drosophila Rab2 controls endosome-lysosome fusion and LAMP delivery to late endosomes

Drosophila Rab2 controls endosome-lysosome fusion and LAMP delivery to late endosomes

Targeted gene alteration in Caenorhabditis elegans by gene conversion

Germline transgenesis of the chordate Ciona intestinalis with hyperactive variants of sleeping beauty transposable element

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