CCM1 and ICAP-1 proteins are destabilized upon CCM1 and CCM2 loss. (A) ICAP-1, CCM1, and CCM2 protein content in total lysates of KD HUVECs was analyzed by Western blot. (B) Quantification of the three proteins normalized to actin in silenced HUVEC; Error bars are ± SEM (n = 3). ICAP-1 and CCM1 proteins were strongly reduced in the three conditions. (C) Q-PCR measurements show that knock-down of one protein had not effect on the expression of the two other partners. Error bars are means ± SEM (n = 3). (D) Western blot of CCM2 +/+ and −/− embryo lysates showing the loss of CCM1 and ICAP-1 proteins upon CCM2 depletion. Remaining CCM2 comes from maternal material. (E) Western blot of ICAP-1 andCCM1 in liver lysates from ICAP-1+/+ and ICAP-1−/− mice. One fourth of CCM1 protein remains in ICAP-1−/− mice (see Fig. S1 C for densitometric analysis). Black lines indicate that intervening lanes have been spliced out for presentation purposes. (F) Overexpressed ICAP-1 protein in CHO is stabilized by the coexpression of CCM1 but not of the N192-Y195 non-interacting CCM1 mutant. Cycloheximide was added at t = 0 to block protein synthesis with or without the proteasomal inhibitor MG132. Results are representative of three independent experiments. (G) Quantification of ICAP-1 protein level over a time-course after addition of cycloheximide when expressed alone or in different combinations with CCM1 and/or CCM2. Results are representative of two independent experiments.