DDX3X interacts with the translational machinery via the 38–44 residues. (A) Diagram of the DDX3X protein structure. DDX3X contains a helicase core, highly conserved across the DEAD-box helicase protein family, composed of two RecA-like domains, denoted as Domain I and Domain II (denoted in blue). In addition to the helicase core, the functional core of the protein also includes the NTE and CTE, which have been shown to be necessary for the RNA unwinding activity of DDX3X. Outside of the functional core, there are the N and C termini, which are less conserved across the protein family, but contain regions conserved across the Ded1/DDX3X subfamily (denoted in gray). (B) Immunoprecipitation of DDX3X mutants. HEK 293T cells were lentivirally transduced with FLAG-tagged DDX3X WT or several mutants, including the helicase defective mutant R534H and three N- and C-termini mutations in sites conserved across the DDX3X/Ded1 subfamily. Immunoprecipitation was conducted for FLAG and run on western blot, staining for ribosome-related proteins and controls. Note that for DDX3X 14–21, the DDX3X antibody signal is lowered because the DDX3X epitope is affected by the 14–21 deletion. (C) DDX3X coimmunoprecipitation with translation machinery proteins. HEK 293T cells were transduced with FLAG-tagged DDX3X WT, R534H, or 38–44ala. Immunoprecipitation was conducted for FLAG and immunoblotted for ribosome-related proteins and controls.
