Identification and partial characterization of type 2 transglutaminase interactome in normal and autophagic conditions

Transglutaminase 2 (TG2) is a Ca+2 dependent enzyme that posttranslationally modifies glutamine residues of protein side chains by transamidation and deamidation. TG2 is the most abundant isoform in mammalian cells and ubiquitously expressed. As a pleiotropic enzyme, it is involved in a variety of cellular processes, including cell growth, apoptosis and autophagy as well as in several diseases such as neurodegenerative disorders. Autophagy is a self-degradative process in which cells remove worn-out or damaged components with replacement of new ones. During abnormal and pathological conditions, ubiquitinated misfolded and mutated proteins form insoluble aggregates which cannot be removed by proteasome. Accumulation of these ubiquitinated aggregates inside the cells severely impair cellular physiology and contributes to the pathogenesis of several common diseases. Autophagy plays a crucial role in the selective degradation of these aggregated proteins and mediates homeostasis. Previous studies have shown that TG2 is involved in autophagosome maturation as well as autophagy-dependent clearance of ubiquitinated proteins. In addition, cells lacking TG2 have been shown to display impaired autophagy/mitophagy and as a consequence shift their metabolism to glycolysis. To define the molecular partners of TG2 involved in these cellular processes, we carried out the analysis of the TG2 interactome under normal and autophagic conditions. We found that TG2 interacts with several groups of proteins. Among the different interactors, there is an interesting group of chaperones participating in the handling of ubiquitinated and misfolded proteins, such as HSPA1A, EF1A, Hsc70, HSP90 and others. HSPA1A interacts with another co-chaperone, BAG3, and with cargo proteins such as p62 for the clearance of ubiquitinated proteins through autophagy. Interestingly, the TG2 interaction with these proteins is much stronger in response to the cellular stress leading to the accumulation of ubiquitinated protein aggregates. In addition, we analysed the interaction of TG2 with PKM2, a rate limiting enzyme of glycolysis. Noteworthy, the cells lacking PKM2 resulted in a significant accumulation of p62 whereas microtubule-associated protein 1 light chain 3 cleaved isoform II (LC3 II), ULK1 and Beclin1 decreased significantly which is suggesting a block in the upstream region of autophagosome formation. These data also indicate a role for PKM2/TG2 interplay on autophagy regulation accompanied by the accumulation of damaged mitochondria.