Tissue transglutaminase as a central mediator in inflammation-induced progression of breast cancer

Abstract

TGM2 is a stress-responsive gene that encodes a multifunctional and structurally complex protein called tissue transglutaminase (abbreviated as TG2 or tTG). TGM2 expression is frequently upregulated during inflammation and wounding. Emerging evidence indicates that TGM2 expression is aberrantly upregulated in multiple cancer cell types, particularly those selected for resistance to chemotherapy and radiation therapy and those isolated from metastatic sites. It is becoming increasingly evident that chronic expression of TG2 in epithelial cancer cells initiates a complex series of signaling networks which contributes to the development of drug resistance and an invasive phenotype. For example, forced or basal high expression of TG2 in mammary epithelial cells is associated with activation of nuclear transcription factor-kappa B (NF-κB), Akt, focal adhesion kinase, and hypoxia-inducible factor. All of these changes are considered hallmarks of aggressive tumors. TG2 expression is able to induce the developmentally regulated program of epithelial-to-mesenchymal transition (EMT) and to confer cancer stem cell (CSC) traits in mammary epithelial cells; both EMT and CSCs have been implicated in cancer metastasis and resistance to standard therapies. Importantly, TG2 expression in tumor samples is associated with poor disease outcome, increased drug resistance, and increased incidence of metastasis. These observations imply that TG2 plays a crucial role in promoting an aggressive phenotype in mammary epithelial cells. In this review, we discuss recent evidence that TG2-regulated pathways contribute to the aggressive phenotype in breast cancer.

Figure 1

Allosteric regulation of tissue transglutaminase (TG2) activity and functions. TG2 is a relatively non-specific crosslinking enzyme, and its activity in and outside the cell is regulated by Ca²⁺, guanine nucleotides, and the redox potential. Binding of Ca²⁺ (dissociation constant of approximately 60 μM) is essential for TG2 to acquire a catalytically active 'open' or 'extended' conformation. In contrast, binding of GTP/GDP (dissociation constant of approximately 1.6 μM) renders TG2 in a catalytically inactive 'closed' or 'compact' conformation. Under physiological conditions, high levels of GTP, low redox potential, and low free Ca²⁺ level keep TG2 in its catalytically inactive compact state. However, a calcium ion influx due to extreme stress or cell damage can induce the catalytically active or 'extended' conformation. In comparison with the intracellular environment, the extracellular matrix has a considerably lower GTP level and relatively high Ca²⁺ level. Therefore, the newly secreted TG2 can be expected to be in a catalytically active state. However, a large fraction of the extracellular TG2 in most organs is in an inactive form because of disulfide bonding. Thioredoxin 1 has been suggested to be a physiological activator of oxidized TG2. In the compact or catalytically inactive state, TG2 can act as a scaffold protein and result in the activation of various signaling pathways. In its extended and catalytically active state, TG2 catalyzes highly stable protein crosslinking, resulting in apoptotic death if inside the cell or stabilization of the matrix if outside the cell. ECM, extracellular matrix.

Figure 2

Tissue transglutaminase (TG2) expression induces epithelial-to-mesenchymal transition (EMT) and stem cellness in mammary epithelial cells. Stable expression of TG2 in mammary epithelial cells (shown are MCF-10A cells) resulted in appreciable morphologic and biochemical changes. Considered the hallmarks of the mesenchymal phenotype, these TG2-induced changes are associated with increased invasiveness, resistance to chemotherapeutic drugs, and acquisition of stem cell traits (increase in CD44⁺/CD24^low/− subpopulation, increased capability to form mammospheres and self-renewal ability, and cellular plasticity).

Figure 3

Tissue transglutaminase (TG2)-regulated inflammatory signaling promotes drug resistance and the metastatic phenotype. Owing to its binding and rapid degradation of the inhibitory protein IκBα, TG2 results in constitutive activation of the pro-inflammatory transcription factor NF-κB. Activated NF-κB (specifically the p65/RelA subunit), in complex with TG2, translocates to the nucleus, where it binds to HIF-1α promoter and results in its transcriptional regulation and protein expression even under normal oxygen levels. Increased expression of HIF-1α, in turn, induces the expression of transcription repressors such as Snail, Zeb, and Twist. Collectively, these TG2/NF-κB/HIF-1α-induced alterations result in acquisition of EMT and stem cell traits. Intracellular TG2 is also known to induce the synthesis and deposition of the ECM component proteins, whereas extracellular TG2 stabilizes the ECM by introducing proteolytic-resistant isopeptide bonds between its component proteins. TG2-induced changes in the ECM can alter cell attachment and cell motility functions. Similarly, TG2-catalyzed crosslinking of the ECM could further contribute to the aggressive phenotype of cancer cells by increasing shear stresses. Membrane-bound TG2, on the other hand, can interact with integrin and growth factor receptors (for example, PDGFR) to induce clustering and downstream signaling. Moreover, membrane-bound and integrin-associated TG2 can interact with fibronectin and further amplify integrin-mediated signaling. In a nutshell, aberrant expression of TG2 initiates reprogramming of the transcription machinery, which in turn initiates a whole new series of inside-out and outside-in signaling pathways to confer an aggressive phenotype to breast cancer cells. BMP7, bone morphogenetic protein 7; ECM, extracellular matrix; EMT, epithelial-to-mesenchymal transition; ERK1/2, extracellular signal-regulated kinase 1/2; FAK, focal adhesion kinase; HIF-1, hypoxia-induced factor-1; IκBα, inhibitory κBα; MMP, matrix metalloproteinase; NF-κB, nuclear transcription factor-kappa B; PDGF, platelet-derived growth factor; PDGFR, platelet-derived growth factor receptor; SPARC, secreted protein acidic and rich in cysteine; VEGF, vascular endothelial growth factor.