Type 2 transglutaminase-selective recruitment of proteins into exosomes under stressful cellular conditions

Numerous studies are revealing a role of exosomes in intercellular communication, and growing evidence indicates an important function of these vesicles in the progression and pathogenesis of cancer and neurodegenerative diseases. However, the biogenesis process of exosomes is still unclear. Tissue transglutaminase (TG2) is a multifunctional enzyme with different subcellular localizations, also present in the extracellular matrix, but the mechanism(s) by which TG2 is released outside of the cells are still not known and require further investigation. Therefore, the goal of the present study was to determine whether exosomes might be a vehicle for TG2 to reach the extracellular space, and if TG2 could be involved in the biogenesis of exosomes. To address this issue, exosomes derived from cells in basal conditions and upon proteasome impairment, either expressing TG2 or not, were isolated and analyzed, as well as exosomes obtained from the same cells also expressing a mutated form of huntingtin (mHtt) containing 84 polyglutamine repeats. Our results show that TG2 is present in the exosomes only upon proteasome blockage, a condition in which TG2 interacts with TSG101 and ALIX, two proteins involved in exosomes biogenesis. These data indicate that exosomes are a way by which cells release TG2 to the extracellular space under stressful stimuli. Moreover, we found that TG2 favors the assembly of a protein complex including mHtt, ALIX, TSG101 and BAG3, a co-chaperone involved in the clearance of mHtt. The formation of this complex is paralleled by the selective recruitment of mHtt and BAG3 only in the exosomes derived from TG2 expressing cells. These data strongly indicate TG2 as an important player in the biogenesis of exosomes controlling the selectivity of their cargo.