Abstract
BackgroundTransglutaminase 2 (TG2) and its phosphorylation have been consistently found to be upregulated in a number of cancer cell types. At the molecular level, TG2 has been associated with the activation of nuclear factor-kappa B (NF-κB), protein kinase B (PKB/Akt) and in the downregulation of phosphatase and tensin homologue deleted on chromosome 10 (PTEN). However, the underlying mechanism involved is not known. We have reported that protein kinase A (PKA) induced phosphorylation of TG2 at serine-216 (Ser216) regulates TG2 function and facilitates protein-protein interaction. However, the role of TG2 phosphorylation in the modulation of NF-κB, Akt and PTEN is not explored.MethodsIn this study we have investigated the effect of TG2 phosphorylation on NF-κB, Akt and PTEN using embryonic fibroblasts derived from TG2 null mice (MEFtg2-/-) overexpressing native TG2 or mutant-TG2 (m-TG2) lacking Ser216 phosphorylation site with and without dibutyryl cyclic-AMP (db-cAMP) stimulation. Functional consequences on cell cycle and cell motility were determined by fluorescence activated cell sorting (FACS) analysis and cell migration assay respectively.ResultsPKA activation in TG2 overexpressing MEFtg2-/- cells resulted in an increased activation of NF-κB and Akt phosphorylation in comparison to empty vector transfected control cells as determined by the reporter-gene assay and immunoblot analysis respectively. These effects were not observed in MEFtg2-/- cells overexpressing m-TG2. Similarly, a significant downregulation of PTEN at both, the mRNA and protein levels were found in cells overexpressing TG2 in comparison to empty vector control and m-TG2 transfected cells. Furthermore, Akt activation correlated with the simultaneous activation of NF-κB and a decrease in PTEN suggesting that the facilitatory effect of TG2 on Akt activation occurs in a PTEN-dependent manner. Similar results were found with MCF-7 and T-47D breast cancer cells overexpressing TG2 and m-TG2 further supporting the role of TG2 phosphorylation in NF-κB activation and in the downregulation of PTEN.ConclusionsCollectively, these data suggest that phosphorylation of TG2 at Ser216 plays a role in TG2 mediated activation of NF-κB, Akt and in the downregulation of PTEN. Blocking TG2 phosphorylation may provide a novel strategy to attenuate NF-κB activation and downregulation of PTEN in TG2 overexpressing cancers.
Highlights
Transglutaminase 2 (TG2) and its phosphorylation have been consistently found to be upregulated in a number of cancer cell types
We have investigated the functional impact of loss of TG2 phosphorylation at Ser216 on TG2 mediated modulation of Nuclear factor-kappa B (NF-κB), PTEN and Protein kinase B/Akt (Akt) in embryonic fibroblasts derived from TG2 null mice (MEFtg2-/-) and in breast cancer cells
Since TG2 activates NF-κB, we explored whether protein kinase A (PKA) induced phosphorylation of TG2 at Ser216 play a role in TG2 mediated activation of NF-κB and subsequent downregulation of PTEN
Summary
Transglutaminase 2 (TG2) and its phosphorylation have been consistently found to be upregulated in a number of cancer cell types. TG2 has been associated with the activation of nuclear factor-kappa B (NF-κB), protein kinase B (PKB/Akt) and in the downregulation of phosphatase and tensin homologue deleted on chromosome 10 (PTEN). We have reported that protein kinase A (PKA) induced phosphorylation of TG2 at serine-216 (Ser216) regulates TG2 function and facilitates protein-protein interaction. TG2 can bind and hydrolyze GTP and ATP, and functions as a G-protein in cell signaling processes [3,4]. We have reported that TG2 has intrinsic kinase activity and phosphorylates a number of proteins involved in cell proliferation and/ or apoptosis [7,8,9]. We have shown that TG2 undergoes phosphorylation in response to PKA activation and PKA induced phosphorylation of TG2 modulates TG function and facilitates protein-protein interaction [10]. Emerging evidence suggests that transamidation function of TG2 is not involved in this process [2]
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