Journal: Proceedings of the Royal Society B: Biological Sciences

Article Title: Individual differences in boldness influence patterns of social interactions and the transmission of cuticular bacteria among group-mates

doi: 10.1098/rspb.2016.0457

Figure Lengend Snippet: Spiders under long-wave UV light: (a) exposed to sterile PBS (control); (b) 4 h after exposure to CNK02; and (c) 48 h after exposure to CNK02. (d) An unexposed spider that has interacted with an exposed spider for 24 h. Areas of green fluorescence, suggesting the presence of viable CNK02 cells, are pointed out with arrows. (Online version in colour.)

Article Snippet: After 24 h, we removed the susceptible individual and vortexed it in 1 ml sterile LB amp/ara broth for 10 s. We removed the spider, transferred 40 μl of this solution onto LB amp/ara agar, and incubated this plate and the remaining solution (960 μl) at 30°C for 20 h. We visually counted the number of LB amp/ara broth solutions that fluoresced under a long-wave UV light pen (BioRad) to assess successful transmission, and counted the number of colony forming units (CFUs) that grew and fluoresced on each LB amp/ara plate to approximate the relative bacterial load that had been transferred to susceptible individuals.

Techniques: Sterility, Control, Fluorescence

Journal: Journal of Biological Chemistry

Article Title: WAVE Forms Hetero- and Homo-oligomeric Complexes at Integrin Junctions in Drosophila Visualized by Bimolecular Fluorescence Complementation

doi: 10.1074/jbc.m110.139337

Figure Lengend Snippet: FIGURE 3. Visualization of WAVE-Abi and WAVE-WAVE interactions in wing imaginal discs. A, Wing imaginal discs expressing the indicated Split-YFP construct combinations in the en-Gal4 pattern. Expression of transgenes is verified by antibody staining as indicated. Anterior is to the left. Co-expression of WAVE-NYFP and Abi-CYFP or Abi-NYFP and WAVE-CYFP leads to reconstitution of YFP, whereas co-expression of WAVE-NYFP and CYFP or NYFP and Abi- CYFP does not show YFP fluorescence. B, Wing imaginal discs expressing the indicated Split-YFP construct combinations in the en-Gal4 pattern. Deletion of the N terminus of WAVE disrupts the interaction between WAVE and Abi and no fluorescence can be observed upon co-expression of WAVEN-NYFP and Abi-CYFP. Co-expression of WAVE-NYFP and WAVE-CYFP leads to the reconstitution of YFP, which is also dependent on the N terminus of WAVE, because co-expression of WAVEN-NYFP and WAVE-CYFP does not yield fluorescence. No fluorescence complementation is observed when Abi-NYFP and Abi-CYFP are co-expressed. Scale bar, 50 m. C, co-immunoprecipitation of HA-WAVE with Myc-WAVE. HA-tagged full-length WAVE or HA-tagged WAVEN con- structs were co-transfected with Myc-tagged full-length WAVE into S2R cells and cell lysates were immunoprecipitated with a control antiserum against Slit or an anti-Myc antibody. The immunoprecipitates were probed with an anti-HA antibody. D, gel filtration profiles of endogenous WAVE and WAVE- NYFP-WAVE-CYFP complexes from heads of adult flies. WAVE-WAVE BiFC complexes co-fractionate with high molecular weight complexes at 500–700 kDa sizes. The elution profile of proteins of known molecular mass is indicated at the bottom.

Article Snippet: Primary antibodies were used at the following dilutions: -WAVE 1:5000 (23), mouse -HA 1:1000 (clone 16B12, Covance), rabbit -Myc 1:1000 (A-14, Santa Cruz Biotechnology), -Chaoptin (24B10) 1:40 (Developmental Studies Hybridoma Bank).

Techniques: Expressing, Construct, Staining, Fluorescence, Immunoprecipitation, Transfection, Control, Filtration, High Molecular Weight

Journal: Journal of Biological Chemistry

Article Title: WAVE Forms Hetero- and Homo-oligomeric Complexes at Integrin Junctions in Drosophila Visualized by Bimolecular Fluorescence Complementation

doi: 10.1074/jbc.m110.139337

Figure Lengend Snippet: FIGURE 4. Subcellular localization of WAVE complexes in wing imaginal discs. A, schematic representation of third instar larvae wing imaginal disc. A: anterior, P: posterior. Cross-section reveals the bilayered structure consisting of a columnar monolayer epithelium covered by a squamous peripodial epi- thelium. B, top: detail of a wing imaginal disc (wing pouch) expressing waveRNAi in the en-Gal4 pattern, stained for endogenous WAVE protein (green) and F-actin (red). Note the strong reduction of WAVE protein in the posterior compartment of the wing disc. Bottom: orthogonal section of a wild type wing imaginal discs, stained for endogenous WAVE protein (green) and F-actin (red). C, detail of (wing pouch) co-expressing WAVE-NYFP and Abi-CYFP (left) and WAVE-NYFP and WAVE-CYFP (right) stained for Myc (red) and HA (blue). The strongest YFP signal is observed at the basal side of the wing disc, whereas the strongest antibody staining for the transgenes is at the apical side (orthogonal view). D, detail of wing imaginal discs (wing pouch) co-expressing Myc- WAVE and Abi-HA-CYFP in the posterior compartment detected by an adapted epitope-tagged PLA; PLA signal in red; F-actin (phalloidin) in green. The strongest PLA signal is observed at the basal side of the wing disc. Scale bar, 50 m.

Article Snippet: Primary antibodies were used at the following dilutions: -WAVE 1:5000 (23), mouse -HA 1:1000 (clone 16B12, Covance), rabbit -Myc 1:1000 (A-14, Santa Cruz Biotechnology), -Chaoptin (24B10) 1:40 (Developmental Studies Hybridoma Bank).

Techniques: Expressing, Staining