FIG. 7.

The GPPX3Y motif of TEX14 is essential for binding to the hinge region of CEP55. (A and B) Summary of the modified mammalian-two-hybrid assays (A) Three kinds of transfection vectors were made. One protein coding sequence (“X”) was fused to a transcriptional activation domain sequence (VP16-AD), and the other protein coding sequence (“Y”) was fused to a DNA-binding domain sequence (GAL4-BD). (B) When proteins “X” and “Y” interact, transcriptional activation of the mCherry gene occurs, which is detected as red fluorescence (B, top right). The GAL4-BD-Y vector expresses the Renilla reniformis luciferase, allowing for normalization of transfections. The relative interaction of protein X and Y is determined by the mCherry/Renilla reniformis luciferase ratio. (C to F) Mammalian-two-hybrid interactions of chimeric VP16-AD-X and GAL4-BD-Y proteins in transfected HEK293T cells is shown (see Fig. 6 and Table 2 for additional details).