Skip to main content
. 2017 Feb 1;28(3):429–439. doi: 10.1091/mbc.E16-11-0813

FIGURE 1:

FIGURE 1:

Interactions of IFT-A subunits demonstrated by all-by-all and subtractive VIP assays. (A) Schematic representation of the structure and domain organization of IFT-A proteins. IAB, IFT-A–binding sequence; WD40, WD40 repeat domain; TPR, tetratricopeptide repeat domain; Tubby, Tubby-like domain. (B, C) All-by-all VIP assay. HEK293T cells cultured in six-well plates were transfected with a combination of expression vectors for EGFP-fused and mChe-fused IFT-A proteins as indicated and incubated for 24 h. After confirmation of the expression of the EGFP and mChe fusion proteins in transfected cells under a microscope, lysates were prepared from the cells and precipitated with GST-tagged anti–GFP Nb prebound to glutathione–Sepharose beads. The green (B) and red (C) fluorescence signals on the precipitated beads were observed, and images of the beads were acquired using a BZ-8000 microscope. (D–F) Subtractive VIP assays. HEK293T cells were cotransfected with an expression vector for TULP3 (D), IFT140 (E), or IFT139 (F) fused to EGFP and expression vectors for all but one (as indicated) of the other IFT-A subunits fused to mChe, and lysates prepared from the transfected cells were processed for the VIP assay as described.

HHS Vulnerability Disclosure