Figure 7. Quantitative mass spectrometry: Drep-2 and FMRP were found in a common protein complex.
(A) Strategy for the identification of Drep-2 interactors by quantitative mass spectrometry. UAS-Drep-2GFP was overexpressed using the pan-neural driver line elavc155-Gal4. (B) Volcano plot showing proteins from Drep-2GFP flies binding to anti-GFP and/or plain control beads. A hyperbolic curve (set at an FDR of 1%) separates GFP-enriched proteins (light pink) from background (grey). Proteins enriched in the control are shown in blue. Proteins that were significantly enriched, both in Drep-2GFP flies and in independent control experiments with wild-type flies, are colored magenta (n = 35). Drep-2 and GFP are shown as green dots. (C) Classification of the 35 core network proteins; multiple counts were allowed. (D) Network of the 35 proteins that were significantly and reproducibly enriched in GFP pulldown experiments (at an FDR of 1%, magenta-colored dots in B). Additional putative interactors of the core network (FDR set at 10%) are shown in white (Supplementary file 2). The circle (node) and font size correspond to the rank within the results (indicated in Supplementary files 1 and 2). The line (edge) width and shade correspond to the number of interactions each of the significantly enriched proteins has with others. The line/edge length is arbitrary. (E) Anti-FMRP probing confirmed the specific presence of FMRP in Drep-2GFP complexes. Head extracts of flies expressing Drep-2GFP or the presynaptic protein Syd-1GFP were processed in parallel. FMRP was only enriched in preparations of Drep-2GFP extracts. Immunoprecipitations were performed using either GFP-Trap-A beads (lanes labeled IP) or blocked agarose beads as binding control (labeled Blank beads).
Figure 7—figure supplement 1. Drep-2GFP colocalizes with endogenous Drep-2.
