MIB1 promotes RYK protein turnover. (A) Western blot of cell lysates showing that MIB1 cDNA overexpression results in a decrease in full-length RYK but not RYKICD or MYR-RYKICD. (B) Western blot of cell lysates showing that MIB1 cDNA overexpression results in a decrease in full-length mouse RYK and noncleavable RYK-RC but not mouse RYKICD. (C) Western blot of cell lysates showing that MIB1 cDNA overexpression results in a decrease in full-length mouse RYK in the presence of γ-secretase inhibitors. (D) Western blot of cell lysates showing that MIB1-dependent degradation of RYK can be reversed by inhibition of the proteasome with MG132 or inhibition of the lysosome with chloroquine. (E) Western blot of cell lysates demonstrating that the inhibition of the expression of TSG101, a member of the ESCRT pathway, reverses the ability of MIB1 to degrade RYK. (F) Western blot of cell lysates demonstrating that the turnover of full-length RYK in the presence of translation inhibition by cycloheximide can be reversed with proteasome or lysosome inhibition. (G) Western blot of cell lysates demonstrating that the turnover of full-length RYK in the presence of cycloheximide can be reversed by knockdown of MIB1 with siRNAs. Blots represent n = 3.