Physiological, pathological, and structural implications of non-enzymatic protein-protein interactions of the multifunctional human transglutaminase 2

Abstract

Transglutaminase 2 (TG2) is a ubiquitously expressed member of an enzyme family catalyzing Ca(2+)-dependent transamidation of proteins. It is a multifunctional protein having several well-defined enzymatic (GTP binding and hydrolysis, protein disulfide isomerase, and protein kinase activities) and non-enzymatic (multiple interactions in protein scaffolds) functions. Unlike its enzymatic interactions, the significance of TG2's non-enzymatic regulation of its activities has recently gained importance. In this review, we summarize all the partners that directly interact with TG2 in a non-enzymatic manner and analyze how these interactions could modulate the crosslinking activity and cellular functions of TG2 in different cell compartments. We have found that TG2 mostly acts as a scaffold to bridge various proteins, leading to different functional outcomes. We have also studied how specific structural features, such as intrinsically disordered regions and embedded short linear motifs contribute to multifunctionality of TG2. Conformational diversity of intrinsically disordered regions enables them to interact with multiple partners, which can result in different biological outcomes. Indeed, ID regions in TG2 were identified in functionally relevant locations, indicating that they could facilitate conformational transitions towards the catalytically competent form. We reason that these structural features contribute to modulating the physiological and pathological functions of TG2 and could provide a new direction for detecting unique regulatory partners. Additionally, we have assembled all known anti-TG2 antibodies and have discussed their significance as a toolbox for identifying and confirming novel TG2 regulatory functions.

Fig. 1

Scheme of TG2 with its domains along with its proposed functions. TG2 domains are color coded. Domain boundaries have been indicated by amino acid numbers. Calcium binding residues, catalytic residues, and GTP/GDP binding residues have also been indicated. Amino acid numbers have been provided for the catalytic residues and GTP/GDP binding residues. This figure is adapted from Mehta et al. [181]

Fig. 2

Interaction sites of TG2 binding proteins and ID regions in the TG2 crystal structure. The left panel and right panel show the front and back views of the TG2 crystal structure in the closed (upper half, 1kv3) and open form (lower half, 2q3z). Intrinsically disordered regions are shown in lime green. Interacting sites overlapping with disordered protein regions with linear interaction motifs are displayed: fibronectin (olive-green), syndecan (orange), SUMO 1 (dark yellow, light green), α1-adrenoceptor (beige)

Fig. 3

Scheme of TG2 and its non-enzymatic partners in regulating TG2 functions. a TG2 facilitates cell adhesion and migration by binding to cell-surface receptors, integrin, and syndecan-4 along with cell matrix protein fibronectin (FN). The interaction takes place in the extracellular matrix (ECM); b TG2 serves as a scaffolding protein by bridging integrin and platelet-derived growth factor receptor (PDGFR), thus participating in amplifying joint PDGFR/integrin signaling via activation of downstream signaling cascades (protein kinase B also called Akt, focal adhesion kinase (FAK), and extracellular signal regulated kinase (ERK); TG2 was shown to form a complex with integrin and low-density lipoprotein receptor related protein (LRP5/6), which in turn activates the β catenin pathway. TG2 was also shown to interact with LRP1, thus participating in LRP1-mediated cell adhesion and migration functions; c membrane metalloprotease (MT MMP) activates soluble metalloprotease (MMP2), which in turn binds to TG2 and mediates its degradation (solid black arrow represents activation), heat shock protein (HSP70) interacts with cytoplasmic (Cyto) TG2 and facilitates its migration to the cell periphery (dotted arrow represents migration of TG2 to the cell periphery), TG2 interacts with protein kinase A anchor protein 13 in the cytoplasm, which in turn interacts with RhoA, thereby potentially facilitating stress fiber formation, cancer cell polarity, and migration; d TG2 interacts with a tumor suppressor protein PTEN (phosphatase and tensin homologue deleted on chromosome 10) and facilitates its ubiquitination and simultaneous degradation by the proteasomal pathway, TG2 modulates Rac (Rho family of small GTPases) activity and its function by blocking the Rac binding site of GTPase-activating proteins such as Bcr/Abr; e TG2 interacts with retinoblastoma protein (Rb) and mediates the pro-apoptotic response when the interaction takes place in the cytoplasm but mediates the anti-apoptotic response when the interaction takes place in the nucleus. Similarly, TG2 mediates the pro-apoptotic response upon interacting with the Bcl2 family member Bax and 14-3-3, a serine/threonine binding protein; f calreticulin modulates TG2 activity by interacting with TG2 and downregulating its TGase and GTPase activities, TG2 facilitates transmembrane signaling by interacting with several transmembrane receptors like adrenoceptor (AR) and along with phospholipase C δ1 (PLCδ1), mediates the release of two second messengers, inositol 4,5 triphosphate (IP3) and diacylglycerol (DAG); g importin α3 interacts with TG2 and mediates its active transport into the nucleus where it can interact with histones, GDP and GTP-bound TG2 regulate the changes in the cellular localization of eukaryotic elongation factor elf5A; h calmodulin interacts with TG2 and increases its TGase activity. Activated TG2 simultaneously binds to tubulin and crosslinks huntingtin (Htt), thus contributing to Huntington’s disease pathology; i intracellular reactive oxygen species (ROS) mediates TG2 SUMOylation by facilitating interaction of TG2 with small ubiquitin-like modifiers 1 (SUMO1) (Fig. 2) and PIASy (a SUMO E3 ligase). ROS-mediated TG2 SUMOylation leads to an increase in TG2 activity, which subsequently affects inflammation by crosslinking and degrading PPARγ (peroxisome proliferator-activator receptor, a negative regulator of inflammation). TG2 was also shown to interact with Rac 1 and regulate inflammation by activating the NF-kβ pathway; j nuclear TG2 was shown to regulate ECM remodeling and metastasis by modulating MMP9 gene transcription. It does so by interacting with cJun, thereby inhibiting dimerization of cJun and cFos and subsequently blocking the AP1 transcription factor binding site on the MMP9 gene; k TG2 regulates metastasis by mediating phosphorylation of β catenin via the bound src. Phosphorylated β catenin dissociates from cadherin and translocates to the nucleus, where it activates Wnt signaling machinery; l TG2 regulates hypoxia by preventing the dimerization of hypoxia-inducible factor 1β and 1α (HIF1β, HIF1α), subsequently inhibiting functional HIF transcription factor binding to hypoxia response elements (HRE) present in the promoter of the Bnip3 gene