Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2019 Nov 29;10:5463. doi: 10.1038/s41467-019-13237-8

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

PMC Copyright notice

Fig. 4 — Cisd2 interacts with IRP1A and AGBE. a Protein–protein interaction map. Lines carrying knock-in alleles encoding Flag-tagged AGBE, IRP1A and IRP1B (yellow boxes, Supplementary Fig. 4) were used to produce bait (circle) for immunoprecipitation followed by mass spectrometry (MS) to identify physically bound proteins to the bait. Whole-body (WB, black) and prothoracic gland samples (PG) were used. Red: detected in both PG and WB samples. Dashed line: Only WB samples were tested for AGBE. H2Av, H2A, H2B and H4 are histone proteins. GlyS = Glycogen Synthase. b Venn diagram depicting overlaps of immunoprecipitated proteins from endogenously tagged proteins (WB samples). H4 & GlyS see A. c Co-transfection of Schneider 2 cells with plasmids encoding Myc-tagged AGBE, Flag-tagged IRP1A and HA-tagged Cisd2, followed by immunoprecipitation via anti-Myc or anti-Flag antibodies and Western blotting. Names shown in red indicate the protein used as bait. Myc-tagged and Flag-tagged enhanced GFP (eGFP^M and eGFP^F, respectively) served as negative controls. Presence of co-immunoprecipitated proteins were tested with anti-HA antibodies and anti-Myc antibodies. d Quantification of immunoprecipitated Cisd2 in the triple co-transfection experiment shown above in C. Graph shows relative fold change of co-immunoprecipitated Cisd2 with Flag-IRP1A as bait in the presence or absence of AGBE. Data was normalized to the amount of Cisd2 protein in the absence of co-transfected AGBE. The asterisk indicates a P-value < 0.05 according to the Student’s t test. Error bars represent standard deviation based on three biological replicates. e Survival rates of Cisd2^IR-RNAi animals and Cisd2^G6528 mutants on fly food ± BPS. nd = not detected. Error bars represent standard deviation from three biological replicates (each sample contained 50 individuals). f Autofluorescence/protoporphyrin accumulation in prothoracic glands (PG) of PG > Cisd2^IR and Cisd2^G6528 larvae reared on fly food ± BPS. Scale bar = 250 μm. g Genetic interaction between Cisd2 and IRP1A on regular (=iron-replete) fly food based on autofluorescening PGs and survival of the corresponding RNAi lines. All lines express RNAi via a PG-specific Gal4 driver (phm22-Gal4 = PG >). Scale bar = 250 μm. h. Subcellular localization of Flag-tagged IRP1A and IRP1B proteins expressed from knock-in alleles (Supplementary Table 1) in Cisd2^G6528 mutants reared on fly food ± BPS. For control larvae, see Supplementary Fig. 10. Scale bar = 500 μm. Source data are provided as a Source Data file.