Tools allowing independent visualization and genetic manipulation of Drosophila melanogaster macrophages and surrounding tissues Journal Article


Author(s): Gyoergy, Attila; Roblek, Marko; Ratheesh, Aparna; Valoskova, Katarina; Belyaeva, Vera; Wachner, Stephanie; Matsubayashi, Yutaka; Sanchez-Sanchez, Besaiz J; Stramer, Brian; Siekhaus, Daria E
Article Title: Tools allowing independent visualization and genetic manipulation of Drosophila melanogaster macrophages and surrounding tissues
Affiliation IST Austria
Abstract: Drosophila melanogaster plasmatocytes, the phagocytic cells among hemocytes, are essential for immune responses, but also play key roles from early development to death through their interactions with other cell types. They regulate homeostasis and signaling during development, stem cell proliferation, metabolism, cancer, wound responses and aging, displaying intriguing molecular and functional conservation with vertebrate macrophages. Given the relative ease of genetics in Drosophila compared to vertebrates, tools permitting visualization and genetic manipulation of plasmatocytes and surrounding tissues independently at all stages would greatly aid in fully understanding these processes, but are lacking. Here we describe a comprehensive set of transgenic lines that allow this. These include extremely brightly fluorescing mCherry-based lines that allow GAL4-independent visualization of plasmatocyte nuclei, cytoplasm or actin cytoskeleton from embryonic Stage 8 through adulthood in both live and fixed samples even as heterozygotes, greatly facilitating screening. These lines allow live visualization and tracking of embryonic plasmatocytes, as well as larval plasmatocytes residing at the body wall or flowing with the surrounding hemolymph. With confocal imaging, interactions of plasmatocytes and inner tissues can be seen in live or fixed embryos, larvae and adults. They permit efficient GAL4-independent FACS analysis/sorting of plasmatocytes throughout life. To facilitate genetic analysis of reciprocal signaling, we have also made a plasmatocyte-expressing QF2 line that in combination with extant GAL4 drivers allows independent genetic manipulation of both plasmatocytes and surrounding tissues, and a GAL80 line that blocks GAL4 drivers from affecting plasmatocytes, both of which function from the early embryo to the adult.
Journal Title: G3: Genes, Genomes, Genetics
Volume: 8
Issue 3
ISSN: 2160-1836
Publisher: Genetics Society of America  
Publication Place: United States
Date Published: 2018-03-01
Start Page: 845
End Page: 857
Copyright Statement: CC BY
URL:
DOI: 10.1534/g3.117.300452
Notes: We thank J. Brennecke , K. Brü ckner, P. Duchek, J. Heinisch, H.Gilbert, L. Luo, C. Potter, J. Sekelsky , K. Senti, and the Drosophila Genomics Resource Center, supported by NIH grant 2P40OD010949-10A1, for plasmids, F. Mauri and J. Knöblich for cell lines , and K. Brueckner, P. Duchek , P. Martin, R. Reuter, the Bloomington Drosophila Stock Center supported by NIH grant P40OD018537 and the Vienna Drosophila Resource Center for fly stocks. We utilized an antibody contributed by U. Banerjee, and produced by the Developmental Studies Hybridoma Bank, which was created by the NICHD of the NIH and is maintained at the University of Iowa. We thank the Life Scientific Service Units at IST Austria for technical support and assistance with microscopy and FACS analysis. We are grateful to T. Hurd and P. Rangan for comments on the manuscript. A. György and A. Ratheesh were supported by the Austrian Science Fund grant DASI_FWF01_P29638S, and A. Ratheesh also by Marie Curie IIF GA-2012-32950BB:DICJI, Marko Roblek by grant LSC16_021 from the NÖ Forschungs und Bildungsges.m.b.H. (NFB) and the provincial government of Lower Austria, K. Valoskova and S. Wachner by DOC Fellowships from the Austrian Academy of Sciences, and D.E. Siekhaus by Marie Curie CIG 334077/IRTIM.
Open access: yes (OA journal)