LRR domains are not sufficient for interaction with KIFC1 tail sequence. (A) A schematic is shown of the KIFC1 interacting clone with the 357 amino acid open reading frame indicated with an arrow. The LRR domains are indicated with black boxes and below are shown the regions tested for interaction with the KIFC1 bait plasmid and the results of interaction tests. (B) Plate assay of interaction of TLRR deletion constructs with KIFC1bait. The plate on the left selects only for yeast harboring both bait and prey plasmids; all cotransformants are able to grow. The plate on the right also lacks adenine and histidine and is therefore selective for transactivation of the reporter genes. Transactivation of the MEL1 reporter gene is detected by incorporation of X-α-gal into the media resulting in blue colonies. Positive control for protein interaction (+C) is yeast cotransformed with pGADT7-T and pGBKT7-53 while cells containing pGADT7 and pGBKT7-Lam (-C) is negative control. (C) Liquid assay of interaction of TLRR deletion constructs with KIFC1bait. The β-galactosidase activity of cultures of TLRR/KIFC1 bait cotransformants was determined as described in Methods. Each assay was repeated at least three times. Standard error of the mean for each transformant is indicated by bracketed lines in panel C.
Wang and Sperry BMC Cell Biology 2008 9:9 doi:10.1186/1471-2121-9-9