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. 2015 Dec 14;291(7):3145–3157. doi: 10.1074/jbc.M115.703058

FIGURE 9.

FIGURE 9.

The TspanC8s bind differentially to the disintegrin (D), cysteine-rich (C), and stalk (S) regions of ADAM10. A, HEK-293T cells were mock transfected (−) or transfected with FLAG-tagged mouse TspanC8s or CD9, and co-transfected with the pDisplay vector containing HA-tagged human ADAM10DCS. Cell lysates were produced in 1% digitonin lysis buffer and immunoprecipitated with an anti-FLAG antibody. Immunoprecipitated proteins were blotted with anti-HA tag antibody (top panel) or anti-FLAG antibody (lower panel). Whole cell lysates were probed with the anti-Myc tag antibody (middle panel). B, data from panel A (upper panel) were quantitated and presented as the amount of immunoprecipitated ADAM10DCS relative to the Tspan14 immunoprecipitation, which was arbitrarily set to 100. Data were normalized by log transformation and statistically analyzed using a one-way ANOVA with a Dunnett's multiple comparison test compared with the CD9 control. All TspanC8s bound significantly to ADAM10DCS (p < 0.001). Error bars represent the standard error of the mean from three experiments. C and D, these experiments were carried out as described for panels A and B except using HA-tagged human ADAM10CS. All TspanC8s bound significantly to ADAM10DCS (p < 0.0001). E and F, these experiments were carried out as for panels A and B except using HA-tagged human ADAM10S (****, p < 0.0001; **, p < 0.01; *, p < 0.05).