Person: Villalta, Christians
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Villalta
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Christians
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Villalta, Christians
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Publication The Twin Spot Generator for Differential Drosophila Lineage Analysis(Springer Science and Business Media LLC, 2009-07-26) Griffin, Ruth; Sustar, Anne; Bonvin, Marianne; Binari, Richard; del Valle Rodriguez, Alberto; Hohl, Amber M; Bateman, Jack R; Villalta, Christians; Heffern, Elleard; Grunwald, Didier; Bakal, Chris; Desplan, Claude; Schubiger, Gerold; Wu, C-ting; Perrimon, NorbertIn Drosophila, widely-used mitotic recombination-based strategies generate mosaic flies with positive readout for only one daughter cell after division. To differentially label both daughter cells, we developed the Twin Spot Generator technique (TSG) and demonstrate that through mitotic recombination, TSG generates green and red twin spots in internal fly tissues, visible even as single cells. We discuss the wide applications of TSG to lineage and genetic mosaic studies.Publication Exploiting Position Effects and the Gypsy Retrovirus Insulator to Engineer Precisely Expressed Transgenes(Springer Science and Business Media LLC, 2008-03-02) Markstein, Michele; Pitsouli, Chrysoula; Villalta, Christians; Celniker, Susan E.; Perrimon, NorbertA major obstacle to creating precisely expressed transgenes lies in the epigenetic effects of the host chromatin that surrounds them. Here we present a strategy to overcome this problem, employing a Gal4-inducible luciferase assay to systematically quantify position effects of host chromatin and the ability of insulators to counteract these effects at phiC31 integration loci randomly distributed throughout the Drosophila genome. We identify loci that can be exploited to deliver precise doses of transgene expression to specific tissues. Moreover, we uncover a previously unrecognized property of the gypsy retrovirus insulator to boost gene expression to levels severalfold greater than at most or possibly all un-insulated loci, in every tissue tested. These findings provide the first opportunity to create a battery of transgenes that can be reliably expressed at high levels in virtually any tissue by integration at a single locus, and conversely, to engineer a controlled phenotypic allelic series by exploiting several loci. The generality of our approach makes it adaptable to other model systems to identify and modify loci for optimal transgene expression.Publication Vector and Parameters for Targeted Transgenic RNA Interference in Drosophila melanogaster(Springer Nature, 2008-01) Ni, Jian-Quan; Markstein, Michele; Binari, Richard; Pfeiffer, Barret D.; Liu, Lu-Ping; Villalta, Christians; Booker, Matthew; Perkins, Lizabeth; Perrimon, NorbertThe conditional expression of hairpin constructs in Drosophila melanogaster has emerged in recent years as a method of choice in functional genomic studies. To date, upstream activating site–driven RNA interference constructs have been inserted into the genome randomly using P-element–mediated transformation, which can result in false negatives due to variable expression. To avoid this problem, we have developed a transgenic RNA interference vector based on the phiC31 site-specific integration method.Publication Hierarchical Rules for Argonaute Loading in Drosophila(Elsevier BV, 2009-11) Czech, Benjamin; Zhou, Rui; Erlich, Yaniv; Brennecke, Julius; Binari, Richard; Villalta, Christians; Gordon, Assaf; Perrimon, Norbert; Hannon, Gregory J.Drosophila Argonaute-1 and Argonaute-2 differ in function and small RNA content. AGO2 binds to siRNAs, whereas AGO1 is almost exclusively occupied by microRNAs. MicroRNA duplexes are intrinsically asymmetric, with one strand, the miR strand, preferentially entering AGO1 to recognize and regulate the expression of target mRNAs. The other strand, miR*, has been viewed as a byproduct of microRNA biogenesis. Here, we show that miR*s are often loaded as functional species into AGO2. This indicates that each microRNA precursor can potentially produce two mature small RNA strands that are differentially sorted within the RNAi pathway. miR* biogenesis depends upon the canonical microRNA pathway, but loading into AGO2 is mediated by factors traditionally dedicated to siRNAs. By inferring and validating hierarchical rules that predict differential AGO loading, we find that intrinsic determinants, including structural and thermodynamic properties of the processed duplex, regulate the fate of each RNA strand within the RNAi pathway.Publication An Integrative Analysis of the InR/PI3K/Akt Network Identifies the Dynamic Response to Insulin Signaling(Elsevier BV, 2016-09-13) Vinayagam, Arunachalam; Kulkarni, Meghana; Sopko, Richelle; Sun, Xiaoyun; Hu, Yanhui; Nand, Ankita; Villalta, Christians; Moghimi, Ahmadali; Yang, Xuemei; Mohr, Stephanie; Hong, Pengyu; Asara, John; Perrimon, NorbertInsulin regulates an essential conserved signaling pathway affecting growth, proliferation and metabolism. To expand our understanding of the insulin pathway, we combine biochemical, genetic and computational approaches to build a comprehensive Drosophila InR/PI3K/Akt network. First, we map the dynamic protein-protein interaction network surrounding the insulin core pathway using bait-prey interactions connecting 566 proteins. Combining RNA interference screening and phospho-specific antibodies, we find that 47% of interacting proteins affect pathway activity, and using quantitative phosphoproteomics, we demonstrate that ~10% of interacting proteins are regulated by insulin stimulation at the level of phosphorylation. Next, we integrate these orthogonal datasets to characterize the structure and dynamics of the insulin network at the level of protein complexes, and validate our method by identifying regulatory roles for the Protein Phosphatase 2A (PP2A) and Reptin-Pontin chromatin-remodeling complexes as negative and positive regulators of ribosome biogenesis, respectively. Altogether, our study represents a comprehensive resource for study of the evolutionary conserved insulin network.