The coiled coil domain of DEF6 facilitates formation of large vesicle-like, cytoplasmic aggregates that trap the P-body marker DCP1 and exhibit prion-like features

Abstract

DEF6, also known as SLAT and IBP, is critical for the development of autoimmune disease and cancer. In T cells, DEF6 participates in TCR-mediated signalling determining T helper cell-mediated immune responses. In addition, DEF6 acts as a guanine nucleotide exchange factor for Rho GTPases facilitating F-actin assembly and stabilisation of the immunological synapse. However, DEF6 is also a component of mRNA processing bodies (P-bodies) linking it to mRNA metabolism. DEF6 can adopt multiple conformations that result in different cellular localisations and functions. Post translational modifications such as phosphorylation result in conformational change liberating functional domains that are masked in the native stage of DEF6. ITK phosphorylation of Try210/222 liberates the N-terminal end and to a certain extend also the C-terminal coiled coil domain of DEF6 resulting in P-body colocalisation. In fact, the N-terminal 45 amino acids of DEF6 that encode a Ca2+-binding EF hand are sufficient to target P-bodies. Mutant proteins that unleashed the C-terminal coiled coil domain of DEF6 spontaneously aggregated forming large vesicle-like, cytoplasmic structures. These aggregates trapped proteins such as the P-body component DCP1 altering its cytoplasmic localisation. However, cellular stress reversed aggregate formation in mutant DEF6 proteins that contained ITAM and PH domain in conjunction with the coiled coil domain resulting in colocalisation with DCP1. Furthermore, coiled coil-mediated aggregates appeared to function like prions enforcing conformational change onto wild type DEF6 protein.